WO2025260285A1

WO2025260285A1 - Antibody drug conjugate

Info

Publication number: WO2025260285A1
Application number: PCT/CN2024/100198
Authority: WO
Inventors: Wei Gu; Yang Qiu; Rong Shi; Zhongyuan Zhu
Original assignee: Duality Biologics Suzhou Co Ltd; Biontech SE
Current assignee: Duality Biologics Suzhou Co Ltd; Biontech SE
Priority date: 2024-06-19
Filing date: 2024-06-19
Publication date: 2025-12-26
Anticipated expiration: 2026-12-19

Abstract

The invention relates to antibody drug conjugates (ADCs) comprising an antibody or fragment thereof that specifically binds to B7-H3, for use in the treatment of cancers, in particular castration-resistant prostate cancer and small-cell lung cancer. The invention also relates to compositions containing such antibody-drug conjugates, for use in the treatment of cancers.

Description

ANTIBODY DRUG CONJUGATE

FIELD OF THE INVENTION

BACKGROUND TO THE INVENTION

Prostate cancer is the second leading cause of cancer-related death among men worldwide, and incidence continues to rise. At the time of initial diagnosis, most prostate cancer patients exhibit progressive or metastatic disease, with androgen-deprivation therapy (ADT) being the treatment of choice for most patients with metastatic prostate cancer. While initially highly effective, the median time that patients respond well to ADT is just 18-24 months, after which the patient will progress to develop castration-resistant prostate cancer (CRPC) . CRPC is an advanced form of prostate cancer with a low survival rate, as CRPC patients only survive for 9-13 months on average. New hormonal agents (NHA) (such as abiraterone acetate and enzalutamide) are potent, orally available treatment options with a favourable tolerability profile and have replaced docetaxel as preferred choice of first-line therapy for mCRPC. Both of these agents have demonstrated robust improvements on PFS and OS and have also shown a significantly prolonged time to initiation of cytotoxic chemotherapy. However, once mCRPC patients have failed NHA, the benefit from approved therapeutic options appears substantially diminished and no clear single standard of care exists as none of the currently approved agents have been developed for a post-NHA setting. Taxanes docetaxel and cabazitaxel are administered per i. v. infusion and have boxed warnings for hypersensitivity reactions as well as neutropenia/neutropenic deaths. Unless symptomatic or with high tumour burden, few patients are expected to immediately receive docetaxel. Efficacy of docetaxel appears markedly reduced when given after NHA as compared to first line use, likely attributable to at least partial cross-resistance between taxanes and NHA. Cabazitaxel appears to have a low real-world usage and is approved only after failure of docetaxel. Limited data are available for patients who failed both NHA and at least one taxane-based chemotherapy. In general, response to treatment decreases with more lines of prior therapies. There is thus a high unmet medical need for mCRPC patients who have failed prior NHA (s) and prior taxane (s) .

Small-cell lung cancer (SCLC) is an aggressive lung cancer subtype with neuroendocrine differentiation diagnosed in more than 150,000 people worldwide each year. The 3-year survival rate for patients with extensive stage (ES) SCLC is 6%. The addition of immune checkpoint inhibitors atezolizumab or durvalumab to platinum and etoposide chemotherapy followed by maintenance therapy with checkpoint inhibitor alone as first-line treatment for SCLC has led to approximately 30%reduction in the risk of death and durable but modest survival gains for a small subset of patients with ES-SCLC. Limited therapies are available for the majority of patients with SCLC who relapse. Topotecan, the most widely used second-line agent globally, has limited efficacy and an unfavourable safety profile. In 2020, lurbinectedin was approved as a second-line agent on the basis of an objective response rate (ORR) of 35%; however, a randomized study failed to demonstrate OS benefit. No agent is specifically approved for third-line treatment of relapsed SCLC. Therefore, there is a significant need for improved novel treatment options for patients with ES-SCLC.

B7-H3, otherwise known as CD276, is a type I transmembrane protein. Although B7-H3 mRNA is expressed in most tissues, B7-H3 protein has a very limited expression on normal tissues because of its post-transcriptional regulation by microRNAs. However, high expression levels, as detected by immunohistochemistry, have been reported in both CRPC and SCLC, as well as non-small cell lung cancer (NSCLC) , oesophageal squamous cell carcinoma (ESCC) , melanoma, hepatocellular carcinoma (HCC) , cervical cancer, and other solid tumours. B7-H3 is an immune checkpoint molecule with pro-tumorigenic properties including promoting cell proliferation, migration, invasion, angiogenesis, metastatic capacity and anti-cancer drug resistance.

Antibody-drug conjugates (ADCs) comprise biologically active small molecule compounds conjugated to monoclonal antibodies or antibody fragments by chemical methods, so as to fully utilize antibodies’ binding specificity to normal cell and to tumour cell surface antigens, and small molecule’s high anti-tumour biological activity, while avoiding defects such as the low specific efficacy of the former as well as toxic side effects of the latter. Compared with traditional chemotherapeutic or targeted drugs, antibody-drug conjugates can more accurately bind to tumour cells and reduce their effects on normal cells.

There remains an unmet need for safe and effective treatments for B7-H3-positive cancers, in particular cancers such as SCLC and CRPC.

SUMMARY OF THE INVENTION

The present inventors have shown that an anti-B7-H3 antibody-drug conjugate is surprisingly effective for the treatment of B7-H3-expressing cancers with high unmet medical needs, such as ESCC, SCLC, prostate cancer and melanoma. In particular, when administered at a dose of 6 mg/kg or 9 mg/kg once every three weeks.

The present invention provides an antibody-drug conjugate of formula (I) :

or a tautomer, a mesomer, a racemate, an enantiomer or a diastereoisomer thereof, or a pharmaceutically acceptable salt thereof,

wherein:

Ab is an anti-B7-H3 antibody or a fragment thereof, wherein the anti-B7-H3 antibody or a fragment thereof comprises heavy chain complementarity determining regions (HCDRs) 1-3 and light chain complementarity determining regions (LCDRs) 1-3, defined:

(a) according to the Kabat numbering scheme, wherein:

HCDR1 comprises or consists of an amino acid sequence according to SEQ ID NO: 1, HCDR2 comprises or consists of an amino acid sequence according to SEQ ID NO: 2, HCDR3 comprises or consists of an amino acid sequence according to SEQ ID NO: 3, LCDR1 comprises or consists of an amino acid sequence according to SEQ ID NO: 4, LCDR2 comprises or consists of an amino acid sequence according to SEQ ID NO: 5, and LCDR3 comprises or consists of an amino acid sequence according to SEQ ID NO: 6; and/or

(b) according to the IMGT numbering scheme, wherein:

HCDR1 comprises or consists of an amino acid sequence according to SEQ ID NO: 7, HCDR2 comprises or consists of an amino acid sequence according to SEQ ID NO: 8, HCDR3 comprises or consists of an amino acid sequence according to SEQ ID NO: 9, LCDR1 comprises or consists of an amino acid sequence according to SEQ ID NO: 10, LCDR2 comprises or consists of an amino acid sequence according to SEQ ID NO: 11, and LCDR3 comprises or consists of an amino acid sequence according to SEQ ID NO: 12;

n is a connection number, and is an integer from 1 to 10;

L is -L_a-L_b-L_c-;

L_a-is

-L_b-is:

-L_c-is –CH₂-;

R¹ is selected from the group consisting of: -O-, - (R²) N-, -P (=O) (R²) -and -S-;

X is -L¹-CH₂-C (O) -;

L¹ is - (C (R^3a) (R^3b) ) _m-, wherein 0 or at least 1 methylene unit of L¹ is independently replaced by -C (O) -, -C (=S) -, -C (=NR^4b) -or -C (=N₂) -;

wherein each R², each R^3a, each R^3b and each R^4b are each independently hydrogen, protium, deuterium, tritium, halogen, -NO₂, -CN, -OR, -SR, -N (R^a) (R^b) , -C (O) R, -CO₂R, -C (O) C (O) R, -C (O) CH₂C (O) R, -S (O) R, -S (O) ₂R, -C (O) N (R^a) (R^b) , -SO₂N (R^a) (R^b) , -OC (O) R, -N (R) SO₂R, or a C_1-6 aliphatic group optionally substituted with R;

wherein each R, each R^a and each R^b are each independently hydrogen, protium, deuterium, tritium, halogen, -NO₂, -CN, -OH, -SH, -NH₂, -C (O) H, -CO₂H, -C (O) C (O) H, -C (O) CH₂C (O) H, -S (O) H, -S (O) ₂H, -C (O) NH₂, -SO₂NH₂, -OC (O) H, -N (H) SO₂H or a C_1-6 aliphatic group;

m is selected from the group consisting of integers ≥ 0;

when R¹ is -O-or -HN-, at least 1 methylene unit of L¹ is independently replaced by -C (O) -, -C (=S) -, -C (=NR^4b) -or -C (=N₂) -, or each R^3a and each R^3b are not both hydrogen;

for use in treating cancer in a subject.

The invention also provides a composition comprising an antibody-drug conjugate of formula (I’) :

or a tautomer, a mesomer, a racemate, an enantiomer or a diastereoisomer thereof, or a pharmaceutically acceptable salt thereof, or a mixture of any thereof,

wherein:

(a) according to the Kabat numbering scheme, wherein:

(b) according to the IMGT numbering scheme, wherein:

n’ is an average connection number, and is an integer or a decimal from 1 to 10;

L is -L_a-L_b-L_c-;

L_a-is

-L_b-is:

-L_c-is –CH₂-;

X is -L¹-CH₂-C (O) -;

m is selected from the group consisting of integers ≥ 0;

for use in treating cancer in a subject.

The invention also provides a method of treating cancer in a subject comprising administering to the subject an antibody-drug conjugate of formula (I) :

wherein:

(a) according to the Kabat numbering scheme, wherein:

(b) according to the IMGT numbering scheme, wherein:

n is a connection number, and is an integer from 1 to 10;

L is -L_a-L_b-L_c-;

L_a-is

-L_b-is:

-L_c-is –CH₂-;

X is -L¹-CH₂-C (O) -;

m is selected from the group consisting of integers ≥ 0;

when R¹ is -O-or -HN-, at least 1 methylene unit of L¹ is independently replaced by -C (O) -, -C (=S) -, -C (=NR^4b) -or -C (=N₂) -, or each R^3a and each R^3b are not both hydrogen.

The invention also provides a method of treating cancer in a subject comprising administering to the subject a composition comprising an antibody-drug conjugate of formula (I’) :

wherein:

(a) according to the Kabat numbering scheme, wherein:

(b) according to the IMGT numbering scheme, wherein:

L is -L_a-L_b-L_c-;

L_a-is

-L_b-is:

-L_c-is –CH₂-;

X is -L¹-CH₂-C (O) -;

m is selected from the group consisting of integers ≥ 0;

The invention also provides for the use of an antibody-drug conjugate of formula (I) :

wherein:

(a) according to the Kabat numbering scheme, wherein:

(b) according to the IMGT numbering scheme, wherein:

n is a connection number, and is an integer from 1 to 10;

L is -L_a-L_b-L_c-;

L_a-is

-L_b-is:

-L_c-is –CH₂-;

X is -L¹-CH₂-C (O) -;

m is selected from the group consisting of integers ≥ 0;

in the manufacture of a medicament for the treatment of cancer.

The invention also provides for the use of a composition comprising an antibody-drug conjugate of formula (I’) :

wherein:

(a) according to the Kabat numbering scheme, wherein:

(b) according to the IMGT numbering scheme, wherein:

L is -L_a-L_b-L_c-;

L_a-is

-L_b-is:

-L_c-is –CH₂-;

X is -L¹-CH₂-C (O) -;

m is selected from the group consisting of integers ≥ 0;

in the manufacture of a medicament for the treatment of cancer.

In some embodiments, X is -L¹-CH₂-C (O) -, L¹ is - (C (R^3a) (R^3b) ) _m-, m is not 0, and each R^3a and each R^3b are not both hydrogen.

In some embodiments, m is 1, and L¹ is -C (R^3a) (R^3b) -.

In some embodiments, R^3a is a C_1-6 aliphatic group, and R^3b is hydrogen.

In some embodiments, R^3a is methyl, and R^3b is hydrogen.

In some embodiments, R¹ is -O-.

In some embodiments, the antibody-drug conjugate comprises the following structure:

In some embodiments, the antibody-drug conjugate is an antibody-drug conjugate of formula (II) :

or a pharmaceutically acceptable salt thereof,

wherein:

(a) according to the Kabat numbering scheme, wherein:

(b) according to the IMGT numbering scheme, wherein:

n is the connection number, and is an integer from 1 to 10.

In some embodiments, the composition comprises an antibody-drug conjugate of formula (II’ ) :

or a pharmaceutically acceptable salt thereof,

wherein:

(a) according to the Kabat numbering scheme, wherein:

(b) according to the IMGT numbering scheme, wherein:

n’ is an average connection number, and is an integer or a decimal from 1 to 10.

In some embodiments, n is an integer from 3 to 9, such as 4 to 8, such as 5 to 7. In some embodiments, n is 6.

In some embodiments, n’ is an integer or decimal from 3 to 9, such as 4 to 8, such as 5 to 7. In some embodiments, n’ is about 6.

The anti-B7-H3 antibody or fragment thereof may comprise a heavy chain variable region comprising or consisting of an amino acid sequence according to SEQ ID NO: 13, or a variant having at least 80%sequence identity thereto, and a light chain variable region comprising or consisting of an amino acid sequence according to SEQ ID NO 14, or a variant having at least 80%sequence identity thereto.

The anti-B7-H3 antibody may comprise a heavy chain comprising or consisting of an amino acid sequence according to SEQ ID NO: 15, or a variant having at least 80%sequence identity thereto, and a light chain comprising or consisting of an amino acid sequence according to SEQ ID NO 16, or a variant having at least 80%sequence identity thereto.

The cancer may be selected from the group consisting of non-small cell lung cancer (NSCLC) , small cell lung cancer (SCLC) , castration-resistant prostate cancer (CRPC) , oesophageal squamous cell carcinoma (ESCC) , melanoma, hepatocellular carcinoma (HCC) , cervical cancer, and solid tumour.

The cancer may be advanced, unresectable, and/or metastatic.

In some embodiments, the antibody-drug conjugate is administered to the subject at a dose in the range of about 3 mg/kg to about 15 mg/kg, such as in the range of about 6 mg/kg to about 12 mg/kg, such as in the range of about 6 mg/kg to about 9 mg/kg. The antibody-drug conjugate may be administered to the subject at a dose of about 6 mg/kg or about 9 mg/kg.

The antibody-drug conjugate or composition may be administered to the subject once every three weeks (Q3W) . The antibody-drug conjugate or composition may be administered to the subject in the form of a pharmaceutical composition. The antibody-drug conjugate or composition may be administered to the subject intravenously.

In some embodiments, the subject is a human. In some embodiments, the subject has received prior treatment for cancer, such as chemotherapy, immunotherapy and/or targeted therapy.

BRIEF DESCRIPTION OF THE DRAWINGS

Figure 1 –Percentage change in tumour size from baseline in subjects receiving at least one dose of Compound 1 at 6 mg/kg, 9 mg/kg or 12 mg/kg, Q3W.

Figure 2 –Percentage change in tumour size from baseline in subjects having SCLC and receiving Compound 1 (6 mg/kg, 9 mg/kg or 12 mg/kg, Q3W) .

Figure 3 –Percentage change in tumour size from baseline in subjects having CRPC and receiving Compound 1 (6 mg/kg or 9 mg/kg, Q3W) .

Figure 4 –Tumour volume in female BALB/c Nude mice bearing KYSE-150 human ESCC tumours and receiving three injections of either negative control, compound 1 (1/2 mg/kg) , compound 1 (3/6 mg/kg) , ADC-1 (1/2 mg/kg) , or ADC-1 (3/6 mg/kg) .

Figure 5 –Tumour volume in female NCG mice bearing DMS 53 human SCLC tumours and receiving two injections of either negative control, compound 1 (3 mg/kg) , compound 1 (6 mg/kg) , or ADC-1 (6 mg/kg) .

Figure 6 –Tumour volume in female BALB/c Nude mice bearing Calu-6 human lung cancer cell tumours and receiving one injection of either negative control, compound 1 (1 mg/kg) , compound 1 (3 mg/kg) , ADC-1 (1 mg/kg) , or ADC-1 (3 mg/kg) .

Figure 7 –Tumour volume in female NOD/SCID mice bearing A375 human melanoma tumours and receiving one injection of either negative control, compound 1 (1 mg/kg) , compound 1 (3 mg/kg) , compound 1 (10 mg/kg) , ADC-1 (3 mg/kg) or ADC-1 (10 mg/kg) .

Figure 8 –Tumour volume in female BALB/c Nude mice bearing PC-3 human prostate cancer cell tumours and receiving one injection of either negative control, compound 1 (2 mg/kg) , compound 1 (6 mg/kg) , ADC-1 (2 mg/kg) or ADC-1 (6 mg/kg) .

Figure 9 –Tumour volume in male mice bearing prostate cancer patient-derived xenografts and receiving two injections of compound 1 (10 mg/kg) . A) Patient-derived PR9586 in NPG mice. B) Patient derived PR9587 in NOG mice.

Figure 10 –Cell viability (relative luminescence unit) in a prostate cancer micro patient-derived xenograft mouse model treated with negative control or compound 1 (3 mg/kg) . Patient-derived prostate cancer transplantation models A) LD1-2027-410644 and B) LD1-2034-362055.

DETAILED DESCRIPTION OF THE INVENTION

General terms and definitions

This disclosure is not limited by the exemplary methods and materials disclosed herein, and any methods and materials similar or equivalent to those described herein can be used in the practice or testing of embodiments of this disclosure. Numeric ranges are inclusive of the numbers defining the range. Unless otherwise indicated, any nucleic acid sequences are written left to right in 5'to 3'orientation; amino acid sequences are written left to right in amino to carboxy orientation, respectively.

In the context of the present disclosure, the term "about" denotes an interval of accuracy that the person of ordinary skill will understand to still ensure the technical effect of the feature in question. The term typically indicates deviation from the indicated numerical value by ±10%, such as ±9%, ±8%, ±7%, ±6%, ±5%, ±4%, ±3%, ±2%, ±1%, ±0.9%, ±0.8%, ±0.7%, ±0.6%, ±0.5%, ±0.4%, ±0.3%, ±0.2%, ±0.1%, ±0.05%, and for example ±0.01%. As will be appreciated by the person of ordinary skill, the specific such deviation for a numerical value for a given technical effect will depend on the nature of the technical effect. For example, a natural or biological technical effect may generally have a larger such deviation than one for a man-made or engineering technical effect. The term “about” allows to account for technical measurement uncertainty and variations when referring for example to a particular quantity or concentration.

The term “polypeptide” is used in the conventional sense to mean a series of amino acids, typically L-amino acids, connected one to the other, typically by peptide bonds between the α-amino and carboxyl groups of adjacent amino acids. The term “polypeptide” is used interchangeably with the terms “amino acid sequence” , “peptide” and/or “protein” . The term “residues” is used to refer to amino acids in an amino acid sequence.

The term "variant" refers to a polypeptide that has an equivalent function to the amino acid sequences described herein, but which includes one or more amino acid substitutions, insertions or deletions.

The sequence may have one or more deletions, insertions or substitutions of amino acid residues which produce a silent change and result in a functionally equivalent molecule. These sequences are encompassed by the present invention. Deliberate amino acid substitutions may be made on the basis of similarity in polarity, charge, solubility, hydrophobicity, hydrophilicity, and/or the amphipathic nature of the residues as long as the activity is retained.

For example, negatively charged amino acids include aspartic acid and glutamic acid; positively charged amino acids include lysine and arginine; and amino acids with uncharged polar head groups having similar hydrophilicity values include leucine, isoleucine, valine, glycine, alanine, asparagine, glutamine, serine, threonine, phenylalanine, and tyrosine.

As used herein, “variant” is synonymous with “mutant” and refers to a polynucleotide or amino acid sequence which differs in comparison to the corresponding wild-type sequence. The term “wild-type” is used to mean a gene or protein having a polynucleotide or amino acid sequence respectively, which is identical with the native gene or protein respectively.

The nucleic acid sequence may be an RNA or DNA sequence or a variant thereof. The term "polynucleotide" includes an RNA or DNA sequence. It may be single or double stranded. It may, for example, be genomic, recombinant, mRNA or cDNA.

The terms “selectively binds/selectively binding” and “specifically binds/specifically binding” may be used interchangeably herein.

Where a range of values is provided, it is understood that each intervening value, to the tenth of the unit of the lower limit unless the context clearly dictates otherwise, between the upper and lower limits of that range is also specifically disclosed. Each smaller range between any stated value or intervening value in a stated range and any other stated or intervening value in that stated range is encompassed within this disclosure. The upper and lower limits of these smaller ranges may independently be included or excluded in the range, and each range where either, neither or both limits are included in the smaller ranges is also encompassed within this disclosure, subject to any specifically excluded limit in the stated range. Where the stated range includes one or both of the limits, ranges excluding either or both of those included limits are also included in this disclosure.

It must be noted that as used herein and in the appended claims, the singular forms "a" , "an" , and "the" include plural referents unless the context clearly dictates otherwise.

The terms "comprising" , "comprises" and "comprised of' as used herein are synonymous with "including" , "includes" or "containing" , "contains" , and are inclusive or open-ended and do not exclude additional, non-recited members, elements or method steps. The terms "comprising" , "comprises" and "comprised of' also include the term "consisting of' .

The terms “identity” and “%sequence identity” as used herein, may refer to the proportion of nucleotides or amino acids (expressed in percent) of a contiguous nucleotide sequence or contiguous amino acid sequence respectively which across the sequence, are identical to a reference sequence. The identity is calculated by counting the number of aligned nucleobases or amino acids that are identical (aMatch) between the sequence of interest and a reference sequence, and dividing that number by the total number of nucleotides amino acids respectively and multiplying by 100. Therefore, Percentage of Identity = (Matches x 100) /Length of aligned region. Insertions and deletions are not allowed in the calculation the percentage of identity. Chemical modifications of nucleotides may be disregarded provided that the functional capacity to form Watson-Crick base pairing is retained.

Identity comparisons can be conducted by eye, or more usually, with the aid of readily available sequence comparison programs. These commercially available computer programs can calculate %identity between two or more sequences. A suitable computer program for carrying out such an alignment is the GCG Wisconsin Bestfit package (University of Wisconsin, U.S.A.; Devereux et al., 1984, Nucleotide sequences Research 12: 387) . Examples of other software than can perform sequence comparisons include, but are not limited to, the BLAST package (see Ausubel et al., 1999 ibid –Chapter 18) , FASTA (Atschul et al., 1990, J. Mol. Biol., 403-410) and the GENEWORKS suite of comparison tools. Both BLAST and FASTA are available for offline and online searching. For example, the percentage identity between two polypeptide sequences may be readily determined by BLAST which is freely available at http: //blast. ncbi. nlm. nih. gov.

Once the software has produced an optimal alignment, it is possible to calculate %identity. The software typically does this as part of the sequence comparison and generates a numerical result.

The term "alkyl" refers to a monoradical of a saturated straight or branched hydrocarbon. Preferably, the alkyl group comprises from 1 to 40, i.e., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39 or 40, carbon atoms, such as 1 to 30, such as 1 to 20 carbon atoms, such as 1 to 12 carbon atoms, such as 1 to 10 carbon atoms, such as 1 to 8 carbon atoms, such as 1 to 6 or 1 to 4 carbon atoms. Exemplary alkyl groups include methyl, ethyl, propyl, iso-propyl (also called 2-propyl or 1 methylethyl) , butyl, iso-butyl, tert-butyl, n-pentyl, iso-pentyl, sec-pentyl, neo-pentyl, 1, 2-dimethylpropyl, iso-amyl, n-hexyl, iso-hexyl, sec-hexyl, n-heptyl, iso-heptyl, n-octyl, 2-ethyl-hexyl, n-nonyl, n-decyl, n-undecyl, n-dodecyl, n-undecyl, n-dodecyl, n-tridecyl, n-tetradecyl, n-pentadecyl, n-hexadecyl, n-heptadecyl, n-octadecyl, n-nonadecyl, n-icosyl, n-triacontyl, n-tetracontyl, and the like.

The term "halogen" generally refers to fluorine, chlorine, bromine or iodine, and it may be, for example, fluorine or chlorine.

Antibody-drug conjugate

The present invention generally relates to antibody-drug conjugates (ADCs) for uses as defined herein. ADCs are a class of targeted therapeutics that improves both the selectivity and the cytotoxic activity of drugs, such as cancer drugs, by targeting the drugs to specific targets such as cancer cells.

In general, ADCs comprise three main components: (i) an antibody (such as a monoclonal antibody) conjugated to (ii) a linker, which in turn is also conjugated to (iii) a cargo or payload (such as a cytotoxic or chemotherapeutic drug) . The term “cytotoxic” or “chemotherapeutic drug” refers to a drug that reduces or eliminates the viability of a cell. Suitable cytotoxic or chemotherapeutic drugs will be known in the art. Components (ii) and (iii) are together referred to herein as the “payload-linker” or “linker-payload” moiety, and are described in more detail below.

It will be understood by the person skilled in the art that all references in this specification to the term “antibody-drug conjugate” will also include compositions comprising mixtures of antibody-drug conjugates, each which may have different drug-antibody ratios (DARs) . These are described in more detail below with reference to compositions. The drug-antibody ratio (DAR) of the antibody-drug conjugates as described herein may vary. Consequently, the composition may comprise a mixture of antibody-drug conjugates having a number of different DARs, and may therefore have an average DAR which is non-integral. In this specification, the terms “DAR” and “connection number” are synonymous, and the terms “average DAR” and “average connection number” are synonymous.

It will be understood that the term “DAR” or “connection number” when referring to a specific antibody-drug conjugate refers to the number of drug molecules connected (e.g. conjugated) to the antibody, optionally via linkers, in any single antibody-drug conjugate. Thus, the connection number (referred to herein as “n” ) can be considered as the specific number of linker-payload structures conjugated to the antibody in a given antibody-drug conjugate. It will also be understood that the connection number n can affect the safety and therapeutic effectiveness of the antibody-drug conjugate. The number of drug molecules per antibody molecule, also referred to herein as the drug-antibody ratio (DAR) , can be characterized by conventional methods such as UV/visible spectroscopy, mass spectrometry, ELISA assays and HPLC.

The antibody-drug conjugate for use according to the invention has the structure represented by formula (I) :

or a pharmaceutically acceptable salt thereof,

wherein:

Ab is an anti-B7-H3 antibody or a fragment thereof as defined herein;

n is a connection number, and is an integer from 1 to 10;

L is -L_a-L_b-L_c-;

L_a-is

-L_b-is:

-L_c-is –CH₂-;

X is -L¹-CH₂-C (O) -;

m is selected from the group consisting of integers ≥ 0;

In some embodiments, the antibody-drug conjugate has the structure represented by formula (II) :

or a pharmaceutically acceptable salt thereof,

wherein:

Ab is an anti-B7-H3 antibody or a fragment thereof as defined herein; and

n is a connection number, and is an integer from 1 to 10.

In one embodiment of the antibody-drug conjugate for use according to the present invention, the antibody component and the payload-linker component may take the structures shown herein.

In another embodiment, the antibody component and the payload-linker component of the antibody-drug conjugate for use in the present invention may take the form of a tautomer, a mesomer, a racemate, an enantiomer or a diastereoisomer of any of the structures shown.

As used herein, the connection number n is an integer from 1 to 10. In some embodiments, the connection number n may be selected from the group of integers of: 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10.

In some embodiments, the connection number n may be an integer selected from the list of ranges selected from: 1 to 10, 2 to 10, 3 to 9, 4 to 8, or 5 to 7.

In some embodiments, the connection number n may be 1 to 10. In some embodiments, the connection number n may be 2 to 10. In some embodiments, the connection number n may be 3 to 9. In some embodiments, the connection number n may be 4 to 8. In some embodiments, the connection number n may be 5 to 7. In some embodiments, the connection number n may be 5. In some embodiments, the connection number n may be 6.

The antibody component and the linker-payload drug component of the antibody-drug conjugate of the invention are linked (i.e. conjugated) to each other via the linker as defined herein. Such linkers typically have chemically reactive groups at each end. These linkers can form a covalent attachment between two molecules, e.g. the antibody and the drug. Thus, the antibody and the drug are typically be covalently linked via a linker. Suitably, one region of the linker may bind to the antibody and another region of the linker may bind to the drug.

In some embodiments, the antibody-drug conjugate has structural formula (II) :

wherein,

Ab is an anti-B7-H3 antibody or a fragment thereof as defined herein; and

n represents a connection number, and n is selected from the group consisting of integers from 1 to 10, preferably integers from 2 to 8; for example, n is an integer from 4 to 8, for example, n is an integer from 5 to 7, preferably 6.

An antibody-drug conjugate of formula (II) where n is 6 and Ab is an anti-B7-H3 antibody comprising a heavy chain amino acid sequence as set forth in SEQ ID NO: 15 and a light chain amino acid sequence as set forth in SEQ ID NO: 16 is referred to herein as “Compound 1” . It is disclosed as compound WBP301088-X2 in WO2023/236949 and national applications deriving therefrom.

The antibody component and the drug component of the antibody-drug conjugate of the invention are linked (i.e. conjugated) to each other via the linker as defined in the claims.

Such linkers typically have chemically reactive groups at each end. These linkers can form a covalent attachment between two molecules, e.g. the antibody and the drug. Thus, the antibody and the drug may be covalently linked to a linker. Suitably, one region of the linker may bind to the antibody and another region of the linker may bind to the drug.

In some embodiments, the linker may be a cleavable linker. In some embodiments, the linker may be a maleimide tetrapeptide-based cleavable linker.

Composition

The present invention also relates to compositions containing antibody-drug conjugates (ADCs) as defined herein, and mixtures thereof, for uses as defined herein. It will be understood by the person skilled in the art that the composition may comprise multiple antibody-drug conjugates, and that the drug-antibody ratio (as defined herein) of each antibody-drug conjugates may be the same or different.

The composition for use according to the invention is a composition comprising an antibody-drug conjugate of formula (I’) :

wherein:

Ab is an anti-B7-H3 antibody or a fragment thereof as defined herein;

L is -L_a-L_b-L_c-;

L_a-is

-L_b-is:

-L_c-is –CH₂-;

X is -L¹-CH₂-C (O) -;

m is selected from the group consisting of integers ≥ 0;

wherein:

Ab is an anti-B7-H3 antibody or a fragment thereof as defined herein; and

In contrast to the term “connection number” when used alone, it will be understood that the term “average connection number” in the context of a composition containing antibody-drug conjugates refers to the average number of drug molecules connected (e.g. conjugated) to the antibody in all of the ADCs present in the composition. As the skilled person will readily understand, the average connection number (defined as n’ when referred to in the context of a composition containing multiple ADCs) may therefore be an integer or be non-integral.

Thus, the average connection number n’ can be considered as the number of linker-payload moieties (e.g. cytotoxic drugs attached via a linker) per antibody when expressed as an average across the whole of the composition. It will also be understood that the average connection number n’ can affect the safety and therapeutic effectiveness of the antibody-drug conjugate. The number of drug molecules per antibody molecule can be characterized by conventional methods such as UV/visible spectroscopy, mass spectrometry, ELISA assays and HPLC.

In one embodiment, the term “average” may refer to the arithmetic mean, which is calculated as the sum of the connection numbers in all ADCs present in the composition divided by the number of ADCs present. In one embodiment, the term “average” may refer to the median, which is calculated as the middle value separating the greater and lesser halves of the connection numbers in all ADCs present in the composition. In one embodiment, the term “average” may refer to the mode, which is calculated as most frequent value of the connection number in all ADCs present in the composition. In one embodiment, the term “average” may refer to the mid-range, which is calculated as the arithmetic mean of the highest and lowest values of the connection number in all ADCs present in the composition. Preferably, the term “average” refers to the arithmetic mean.

As used herein, the average connection number n’ is an integer or a decimal from 1 to 10. In some embodiments, the average connection number n’ may be selected from the group of integers or decimals of about: 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, 3.4, 3.5, 3.6, 3.7, 3.8, 3.9, 4.0, 4.1, 4.2, 4.3, 4.4, 4.5, 4.6, 4.7, 4.8, 4.9, 5.0, 5.1, 5.2, 5.3, 5.4, 5.5, 5.6, 5.7, 5.8, 5.9, 6.0, 6.1, 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, 7.0, 7.1, 7.2, 7.3, 7.4, 7.5, 7.6, 7.7, 7.8, 7.9, 8.0, 8.1, 8.2, 8.3, 8.4, 8.5, 8.6, 8.7, 8.8, 8.9, 9.0, 9.1, 9.2, 9.3, 9.4, 9.5, 9.6, 9.7, 9.8, 9.9, or 10.0.

In some embodiments, the average connection number n’ may be an integer or a decimal selected from the list of ranges selected from: 1 to 10, 2 to 10, 3 to 9, 4 to 8, or 5 to 7.

In some embodiments, the average connection number n’ is an integer or decimal from about 1 to about 10. In some embodiments, the average connection number n’ is an integer or decimal of about 1, about 2, about 3, about 4, about 5, about 6, about 7, about 8, about 9, or about 10. In some embodiments, the average connection number n’ is an integer or decimal from about 2 to about 10. In some embodiments, the average connection number n’ is an integer or decimal from about 3 to about 9. In some embodiments, the average connection number n’ is an integer or decimal from about 4 to about 8. In some embodiments, the average connection number n’ is an integer or decimal from about 4.5 to about 7.5. In some embodiments, the average connection number n’ is an integer or decimal from about 5 to about 7. In some embodiments, the average connection number n’ is an integer or decimal from about 5.5 to about 6.5. In some embodiments, the average connection number n’ is an integer or decimal from about 5.5 to about 6.0. In some embodiments, the average connection number n’ is an integer or decimal from about 5.5 to about 5.8.

In some embodiments, the average connection number n’ is about 5.0. In some embodiments, the average connection number n’ is about 5.1. In some embodiments, the average connection number n’ is about 5.2. In some embodiments, the average connection number n’ is about 5.3. In some embodiments, the average connection number n’ is about 5.4. In some embodiments, the average connection number n’ is about 5.5. In some embodiments, the average connection number n’ is about 5.6. In some embodiments, the average connection number n’ is about 5.7. In some embodiments, the average connection number n’ is about 5.8. In some embodiments, the average connection number n’ is about 5.9. In some embodiments, the average connection number n’ is about 6.0. In some embodiments, the average connection number n’ is about 6.1. In some embodiments, the average connection number n’ is about 6.2. In some embodiments, the average connection number n’ is about 6.3. In some embodiments, the average connection number n’ is about 6.4. In some embodiments, the average connection number n’ is about 6.5. In some embodiments, the average connection number n’ is about 6.6. In some embodiments, the average connection number n’ is about 6.7. In some embodiments, the average connection number n’ is about 6.8. In some embodiments, the average connection number n’ is about 6.9. In some embodiments, the average connection number n’ is about 7.0.

B7-H3

The antibody-drug conjugate (ADC) for use according to the invention comprises an antibody, or fragment thereof, that specifically binds to B7-H3.

In some embodiments, B7-H3 is considered the antigen of the antibodies as defined herein.

As used herein, the terms “B7-H3” and “CD276” are interchangeable.

B7-H3 exists in two isoforms determined by its extracellular domain (2Ig and 4Ig) . The 2Ig isoform contains a single pair of IgV-like and IgC-like domains, the 4Ig isoforms contains two pairs.

In some embodiments, B7-H3 comprises the amino acid sequence of SEQ ID NO: 17:

In some embodiments, B7-H3 comprises the amino acid sequence of SEQ ID NO: 18:

In some embodiments “specifically binds to” may indicate that the antibody or fragment thereof binds to the antigen, i.e. B7-H3 in preference to other antigens.

Antibodies, or binding fragments thereof, useful in the present invention may bind to the target protein with an affinity that is at least two-fold greater, preferably at least ten times greater, more preferably at least 20-times greater, and most preferably at least 100-times greater than the affinity with non-target proteins.

In embodiments, it will be understood herein that “specifically binds to” refers to the antibody-like binding of the antibody or fragment thereof, which may be via heavy chain complementarity determining regions (HCDRs) 1-3 and light chain complementarity determining regions (LCDRs) 1-3, to the target B7-H3. Thus, in embodiments, it will be understood herein that the term “specifically” does not exclude the binding molecule from having other targets.

Suitable assays and techniques for measuring/quantifying binding activity of the antibody or fragment thereof as defined herein may include, but are not limited to, ELISA, surface plasmon resonance (SPR) , bio-layer interferometry (BLI) , quartz crystal microbalance (QCM) , bioluminescence assays and flow cytometry. Other suitable techniques will be known in the art. For example, it will be understood that EC50 is a measure of the concentration of a binding molecule that induces a specific response that is 50%between the maximum response and the baseline response. As such, EC50 can be used to assess the ability of a binding molecule to bind to a target.

Anti-B7-H3 antibody

In some embodiments, the antibody-drug conjugate comprises an anti-B7H3 antibody or fragment thereof.

The anti-B7H3 antibody or fragment thereof may specifically bind B7H3. The anti-B7H3 antibody may be a monoclonal antibody. In some embodiments, the anti-B7H3 antibody or fragment thereof is a humanized antibody. In some embodiments, the anti-B7H3 antibody or fragment thereof is a monoclonal humanized antibody. The fragment thereof may be any antigen-binding fragment thereof, for example a Fab, a Fab', a F (ab') 2, a Fv, a scFv, a Fab'-SH, an sdAb, or a VHH. In some embodiments, the anti-B7H3 antibody or fragment thereof is a full-length anti-B7H3 antibody.

The anti-B7H3 antibody or fragment thereof may comprise a variable region that specifically binds B7H3. In some embodiments, the anti-B7H3 antibody or fragment thereof comprises a heavy chain variable region and/or a light chain variable domain. In some embodiments, the anti-B7H3 antibody or fragment thereof comprises a heavy chain variable region and a light chain variable domain. In some embodiments, the anti-B7H3 antibody or fragment thereof comprises a constant region, preferably derived from a human antibody, preferably the constant region is selected from the constant region of human IgG1, IgG2, IgG3 or IgG4. In some embodiments, the anti-B7H3 antibody or fragment thereof comprises a heavy chain and/or a light chain. In some embodiments, the anti-B7H3 antibody or fragment thereof comprises a heavy chain and a light chain. In some embodiments, the anti-B7H3 antibody or fragment thereof comprises or consists of two heavy chains and two light chains.

Antibodies described herein include polyclonal and monoclonal antibodies and include IgA such as IgA1 or IgA2, IgG such as IgG1, IgG2, IgG3, or IgG4, IgE, IgM, and IgD antibodies. In various embodiments, the antibody is an IgG1 antibody, more particularly an IgG1, kappa or IgG1, lambda isotype (i.e. IgG1, κ, λ) , an IgG2a antibody (e.g. IgG2a, κ, λ) , an IgG2b antibody (e.g. IgG2b, κ, λ) , an IgG3 antibody (e.g. IgG3, κ, λ) or an IgG4 antibody (e.g. IgG4, κ, λ) . In preferred embodiments the antibody is an IgG1, preferably IgG1, lambda.

The antibody may be of any species (for example human, monkey, camel, llama, goat, sheep, rabbit, mouse, rat, mouse, hamster or chicken) or it may be a hybrid derived from more than one species. It may be naturally occurring or it may be non-naturally occurring (i.e. an isolated antibody) . The antibody may be created by genetic engineering (for example a chimeric antibody, humanised antibody, camelised antibody, intrabody, bispecific antibody) .

“Antibodies” are glycoproteins belonging to the immunoglobulin superfamily. The term "full-length antibody" may refer to an immunoglobulin molecule that binds to a target molecule and contains four peptide chains: two heavy chains and two light chains which are connected to each other through disulfide bonds. Antibodies may comprise several “regions” or “domains” and the terms may be used interchangeably herein. An antibody may recognise an antigen via the fragment antigen-binding (Fab) variable region. The fragment crystallizable region (Fc region) is the tail region of an antibody that may allow antibodies to activate the immune system. The hinge region is a stretch of heavy chains linking the Fab and Fc regions. The heavy chain and light chain may each comprise a variable domain and one or more constant domains. For example, in IgG antibodies, a heavy chain comprises a variable domain (VH) and three constant domains (CH1, CH2, and CH3) and a light chain comprises a variable domain (VL) and one constant domain (CL) . Example antibodies include a human antibody, a humanized antibody, a chimeric antibody, a multispecific antibody, a monoclonal antibody, and a polyclonal antibody.

The term “antibody fragment” may refer to a fragment of an antibody, or a genetically engineered product of one of more fragments of an antibody, which fragment is involved in binding with the target molecule. Examples of antibody fragments include an antigen-binding fragment (Fab) , a Fab', a Fab'-SH, a fragment antibody (F (ab’) 2) , a variable region (Fv) , a single chain variable fragment (scFv) , a single-domain antibody (sdAb) , a nanobody, a VHH, and a camelid antibody. The term “Antigen-binding fragment” or “Fab” refers to a region of an antibody that binds to antigens and is composed of one constant and one variable region of each of the heavy and the light chain. The term “fragment antibody” or “F (ab’) 2” refers to a region of an antibody that remains following digestion of the Fc region while leaving intact some of the hinge region. The term “Fab’ ” refers to a fragment formed by the reduction of a F (ab') 2 fragment. The term “Fab’ -SH” refers to a Fab’ fragment with a free sulfhydryl group. The term “Single chain variable fragment” or “scFv” refers to an engineered antibody consisting of a light chain variable region and a heavy chain variable region which are connected to each other.

“Variable regions” as used herein means a segment of an antibody which contains three CDRs, designated CDR1, CDR2 and CDR3. A “variable region” of an antibody refers to the variable region of the antibody light chain or the variable region of the antibody heavy chain, either alone or in combination. The variable region of the heavy chain may be referred to as “VH. ” The variable region of the light chain may be referred to as “VL. ” Typically, the variable regions of both the heavy and light chains comprise three hypervariable regions, the CDRs, which are located within relatively conserved framework regions (FR) . The CDRs are usually aligned by the framework regions, enabling binding to a specific epitope. In general, from N-terminal to C-terminal, both light and heavy chains variable domains comprise FR1, CDR1, FR2, CDR2, FR3, CDR3, and FR4.

The term “heavy chain” refers to a large protein subunit of an immunoglobulin. Heavy chains can be of any immunoglobulin isotype (for example IgG, IgE, IgM, IgD, IgA or IgY) , subtype (for example lgG1, lgG2, lgG2a, lgG2b, lgG2c, lgG3, lgG4, lgA1 or lgA2) or allotype.

The term “light chain” refers to a small protein subunit of an immunoglobulin. Light chains can be of any type (for example kappa or lambda) , subtype or allotype.

“CDR” or “CDRs” means complementarity determining region (s) in an immunoglobulin variable region. The variable regions of the heavy and light chains each contain three CDRs, designated CDR1, CDR2 and CDR3. The precise boundaries of these CDRs can be defined according to various numbering systems known in the art (see e.g. Dondelinger, M., et al., 2018. Frontiers in immunology, 9, p. 2278) , such as the Kabat numbering system (Kabat et al., Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, Md., 1991; Kabat et al., 1992, Sequences of Proteins of Immunological Interest, DIANE Publishing: 2719) , the Chothia numbering system (Chothia &Lesk (1987) J. Mol. Biol. 196: 901-917; Chothia et al. (1989) Nature 342: 878-883) or the IMGT numbering system (Lefranc et al., Dev. Comparat. Immunol. 27: 55-77, 2003; Ehrenmann F, Kaas Q, Lefranc M P. IMGT/3Dstructure-DB and IMGT/DomainGapAlign: a database and a tool for immunoglobulins or antibodies, T cell receptors, MHC, IgSF and MhcSF [J] . Nucleic acids research, 2009; 38 (suppl_1) : D301-D307) . For a given antibody, those skilled in the art will readily identify the CDRs defined by each numbering system. Also, the correspondence between different numbering systems is well known to those skilled in the art (for example, see Lefranc et al., Dev. Comparat. Immunol. 27: 55-77, 2003) .

“Kabat, ” as used herein, means an immunoglobulin alignment and numbering system pioneered by Elvin a. Kabat ( (1991) Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, Md. ) .

“Chimeric antibody” refers to an antibody in which a portion of the heavy and/or light chain is identical with or homologous to corresponding sequences in an antibody derived from a particular species (e.g., human) or belonging to a particular antibody class or subclass, while the remainder of the chain (s) is identical with or homologous to corresponding sequences in an antibody derived from another species (e.g., mouse) or belonging to another antibody class or subclass, as well as fragments of such antibodies, so long as they exhibit the desired biological activity.

“Human antibody” refers to an antibody that comprises human immunoglobulin protein sequences only. A human antibody may contain murine carbohydrate chains if produced in a mouse, in a mouse cell, or in a hybridoma derived from a mouse cell. Similarly, “mouse antibody” or “rat antibody” refer to an antibody that comprises only mouse or rat immunoglobulin sequences, respectively.

“Humanized antibody” refers to forms of antibodies that contain sequences from non-human (e.g., murine) antibodies as well as human antibodies. Such antibodies contain minimal sequence derived from non-human immunoglobulin. In general, the humanized antibody will comprise substantially all of at least one, and typically two, variable domains, in which all or substantially all of the hypervariable loops correspond to those of a non-human immunoglobulin and all or substantially all of the FR regions are those of a human immunoglobulin sequence.

The term "monoclonal antibody" refers to an antibody obtained from a substantially homogeneous population of antibodies. The individual antibodies composing the population may be identical except for possible naturally occurring mutations, which may be present in minor amounts. Monoclonal antibodies are highly specific and target a single antigenic epitope. In contrast, polyclonal antibody preparations typically include a large number of antibodies which are specific for different epitopes.

The term “multispecific antibody” refers to antibodies which recognise two or more epitopes located on the same or distinct targets. Formats of multispecific antibodies can be divided into two broad categories: IgG-like antibody formats, with an Fc domain, and non-IgG-like antibody formats, without an Fc domain (see e.g. Elshiaty, M., et al., 2021. International journal of molecular sciences, 22 (11) , p. 5632) . Examples of multispecific antibodies include bispecific and trispecific antibodies.

A “single domain antibody” (sdAb) is an antibody composed of a single variable domain (e.g., heavy chain variable region) composed of antibody fragments. Typically, a single domain antibody, domain antibody or nanobody consists of 4 framework regions and 3 complementarity determining regions, the 4 framework regions are respectively FR1-FR4, and the 3 complementarity determining regions are respectively CDR1 -CDR3. In certain embodiments, the single domain antibody of the present application may have a structure of FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4. These antibodies do not require light chain variable regions to bind antigens with high affinity and specificity. Examples of single-domain antibodies include, but are not limited to, VHH fragments, and VNAR fragments. Compared with antibodies composed of heavy chain and light chain, single domain antibodies have high solubility, high stability to heat, pH, protease and other deforming agents, and only need single-chain expression to facilitate large-scale production. As used herein, the term “framework region” or “FR” residues refers to those amino acid residues in an antibody variable region other than the CDR residues as defined above.

Example anti-B7H3 antibody CDRs, variable region sequences, and heavy and light chain sequences are provided below.

In some embodiments, a heavy chain variable region of the anti-B7H3 antibody or fragment thereof comprises: (i) a HCDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 1, or a variant thereof having up to three amino acid substitutions, additions or deletions; (ii) a HCDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 2, or a variant thereof having up to three amino acid substitutions, additions or deletions; and (iii) a HCDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 3, or a variant thereof having up to three amino acid substitutions, additions or deletions, wherein the CDRs are defined according to the Kabat numbering system.

In some embodiments, a light chain variable region of the anti-B7H3 antibody or fragment thereof comprises: (i) a LCDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 4, or a variant thereof having up to three amino acid substitutions, additions or deletions; (ii) a LCDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 5, or a variant thereof having up to three amino acid substitutions, additions or deletions; and (iii) a LCDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 6, or a variant thereof having up to three amino acid substitutions, additions or deletions, wherein the CDRs are defined according to the Kabat numbering system.

In some embodiments, a heavy chain variable region of the anti-B7H3 antibody or fragment thereof comprises: (i) a HCDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 1, or a variant thereof having up to three amino acid substitutions, additions or deletions; (ii) a HCDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 2, or a variant thereof having up to three amino acid substitutions, additions or deletions; and (iii) a HCDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 3, or a variant thereof having up to three amino acid substitutions, additions or deletions; and a light chain variable region of the anti-B7H3 antibody or fragment thereof comprises: (i) a LCDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 4, or a variant thereof having up to three amino acid substitutions, additions or deletions; (ii) a LCDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 5, or a variant thereof having up to three amino acid substitutions, additions or deletions; and (iii) a LCDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 6, or a variant thereof having up to three amino acid substitutions, additions or deletions, wherein the CDRs are defined according to the Kabat numbering system.

In some embodiments, a heavy chain variable region of the anti-B7H3 antibody or fragment thereof comprises: (i) a HCDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 7, or a variant thereof having up to three amino acid substitutions, additions or deletions; (ii) a HCDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 8, or a variant thereof having up to three amino acid substitutions, additions or deletions; and (iii) a HCDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 9, or a variant thereof having up to three amino acid substitutions, additions or deletions, wherein the CDRs are defined according to the IMGT numbering system.

In some embodiments, a light chain variable region of the anti-B7H3 antibody or fragment thereof comprises: (i) a LCDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 10, or a variant thereof having up to three amino acid substitutions, additions or deletions; (ii) a LCDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 11; and (iii) a LCDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 12, or a variant thereof having up to three amino acid substitutions, additions or deletions, wherein the CDRs are defined according to the IMGT numbering system.

In some embodiments, a heavy chain variable region of the anti-B7H3 antibody or fragment thereof comprises: (i) a HCDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 7, or a variant thereof having up to three amino acid substitutions, additions or deletions; (ii) a HCDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 8, or a variant thereof having up to three amino acid substitutions, additions or deletions; and (iii) a HCDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 9, or a variant thereof having up to three amino acid substitutions, additions or deletions; and a light chain variable region of the anti-B7H3 antibody or fragment thereof comprises: (i) a LCDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 10, or a variant thereof having up to three amino acid substitutions, additions or deletions; (ii) a LCDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 11, or a variant thereof having up to three amino acid substitutions, additions or deletions; and (iii) a LCDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 12, or a variant thereof having up to three amino acid substitutions, additions or deletions, wherein the CDRs are defined according to the IMGT numbering system.

In some embodiments, one or more of the CDRs may comprise one, two or three amino acid mutations relative to the recited sequences. In some embodiments, HCDR1 may comprise one, two or three amino acid mutations. In some embodiments, HCDR2 may comprise one, two or three amino acid mutations. In some embodiments, HCDR3 may comprise one, two or three amino acid mutations. In some embodiments, LCDR1 may comprise one, two or three amino acid mutations. In some embodiments, LCDR2 may comprise one, two or three amino acid mutations. In some embodiments, LCDR3 may comprise one, two or three amino acid mutations.

It will be understood that a mutation in any of the CDRs described herein may encompass a deletion of an amino acid, an insertion of an amino acid, or a substitution of an amino acid. It will also be understood that such a mutation may not prevent the anti-B7-H3 antibody or fragment thereof from binding to B7-H3. In other words, an anti-B7-H3 antibody or fragment thereof comprising a mutation in one or more CDRs described herein may suitably maintain the capacity (e.g. affinity) to bind to B7-H3. In some embodiments, the mutation suitably maintains the same capacity (e.g. affinity) to bind to B7-H3 as the parent anti-B7-H3 antibody or fragment thereof. The term “parent binding molecule” in this context refers to the anti-B7-H3 antibody or fragment thereof without the mutation in question.

In some embodiments, the CDR variants have up to two amino acid substitutions, additions or deletions. In some embodiments, the CDR variants have up to one amino acid substitution, addition or deletion. In some embodiments, the CDR variants have up to three amino acid substitutions. In some embodiments, the CDR variants have up to two amino acid substitutions. In some embodiments, the CDR variants have up to one amino acid substitution.

In some embodiments, a heavy chain variable region of the anti-B7H3 antibody or fragment thereof comprises or consists of the amino acid sequence of SEQ ID NO: 13, or an amino acid sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%identity thereto and/or one or more (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10) amino acid additions, deletions and/or substitutions in a framework region thereof. In some embodiments, a heavy chain variable region of the anti-B7H3 antibody or fragment thereof comprises or consists of the amino acid sequence of SEQ ID NO: 13.

In some embodiments, a light chain variable region of the anti-B7H3 antibody or fragment thereof comprises or consists of the amino acid sequence of SEQ ID NO: 14 or an amino acid sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%identity thereto and/or one or more (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10) amino acid additions, deletions and/or substitutions in a framework region thereof. In some embodiments, a light chain variable region of the anti-B7H3 antibody or fragment thereof comprises or consists of the amino acid sequence of SEQ ID NO: 14.

In some embodiments, a heavy chain variable region of the anti-B7H3 antibody or fragment thereof comprises or consists of the amino acid sequence of SEQ ID NO: 13, or an amino acid sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%identity thereto and/or one or more (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10) amino acid additions, deletions and/or substitutions in a framework region thereof; and a light chain variable region of the anti-B7H3 antibody or fragment thereof comprises or consists of the amino acid sequence of SEQ ID NO: 14 or an amino acid sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%identity thereto and/or one or more (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10) amino acid additions, deletions and/or substitutions in a framework region thereof.

In some embodiments, a heavy chain variable region of the anti-B7H3 antibody or fragment thereof comprises or consists of the amino acid sequence of SEQ ID NO: 13, or an amino acid sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%identity thereto and/or one or more (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10) amino acid additions, deletions and/or substitutions in a framework region thereof, wherein the heavy chain variable region comprises: a HCDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 1; a HCDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 2; and a HCDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 3; and a light chain variable region of the anti-B7H3 antibody or fragment thereof comprises or consists of the amino acid sequence of SEQ ID NO: 14 or an amino acid sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%identity thereto and/or one or more (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10) amino acid additions, deletions and/or substitutions in a framework region thereof, wherein the light chain variable region comprises: a LCDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 4; a LCDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 5; and a LCDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 6.

In some embodiments, a heavy chain variable region of the anti-B7H3 antibody or fragment thereof comprises or consists of the amino acid sequence of SEQ ID NO: 13, or an amino acid sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%identity thereto and/or one or more (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10) amino acid additions, deletions and/or substitutions in a framework region thereof, wherein the heavy chain variable region comprises: a HCDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 7; a HCDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 8; and a HCDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 9; and a light chain variable region of the anti-B7H3 antibody or fragment thereof comprises or consists of the amino acid sequence of SEQ ID NO: 14 or an amino acid sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%identity thereto and/or one or more (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10) amino acid additions, deletions and/or substitutions in a framework region thereof, wherein the light chain variable region comprises: a LCDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 10; a LCDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 11; and a LCDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 12.

In some embodiments, a heavy chain variable region of the anti-B7H3 antibody or fragment thereof comprises or consists of the amino acid sequence of SEQ ID NO: 13; and a light chain variable region of the anti-B7H3 antibody or fragment thereof comprises or consists of the amino acid sequence of SEQ ID NO: 14 In some embodiments, a heavy chain variable region of the anti-B7H3 antibody or fragment thereof consists of the amino acid sequence of SEQ ID NO: 13; and a light chain variable region of the anti-B7H3 antibody or fragment thereof consists of the amino acid sequence of SEQ ID NO: 14.

In some embodiments, a heavy chain of the anti-B7H3 antibody or fragment thereof comprises or consists of the amino acid sequence of SEQ ID NO: 15, or an amino acid sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%identity thereto and/or one or more (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10) amino acid additions, deletions and/or substitutions in a framework region thereof. In some embodiments, a heavy chain of the anti-B7H3 antibody or fragment thereof comprises or consists of the amino acid sequence of SEQ ID NO: 15.

In some embodiments, a light chain of the anti-B7H3 antibody or fragment thereof comprises or consists of the amino acid sequence of SEQ ID NO: 16 or an amino acid sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%identity thereto and/or one or more (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10) amino acid additions, deletions and/or substitutions in a framework region thereof. In some embodiments, a light chain of the anti-B7H3 antibody or fragment thereof comprises or consists of the amino acid sequence of the amino acid sequence of SEQ ID NO: 16.

In some embodiments, a heavy chain of the anti-B7H3 antibody or fragment thereof comprises or consists of the amino acid sequence of the amino acid sequence of SEQ ID NO: 15, or an amino acid sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%identity thereto and/or one or more (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10) amino acid additions, deletions and/or substitutions in a framework region thereof; and a light chain of the anti-B7H3 antibody or fragment thereof comprises or consists of the amino acid sequence of SEQ ID NO: 16 or an amino acid sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity thereto and/or one or more (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10) amino acid additions, deletions and/or substitutions in a framework region thereof.

In some embodiments, a heavy chain of the anti-B7H3 antibody or fragment thereof comprises or consists of the amino acid sequence of SEQ ID NO: 15, or an amino acid sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%identity thereto and/or one or more (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10) amino acid additions, deletions and/or substitutions in a framework region thereof, wherein the heavy chain variable region comprises: a HCDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 1; a HCDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 2; and a HCDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 3; and a light chain of the anti-B7H3 antibody or fragment thereof comprises or consists of the amino acid sequence of SEQ ID NO: 16 or an amino acid sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%identity thereto and/or one or more (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10) amino acid additions, deletions and/or substitutions in a framework region thereof, wherein the light chain variable region comprises: a LCDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 4; a LCDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 5; and a LCDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 6.

In some embodiments, a heavy chain of the anti-B7H3 antibody or fragment thereof comprises or consists of the amino acid sequence of SEQ ID NO: 15, or an amino acid sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%identity thereto and/or one or more (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10) amino acid additions, deletions and/or substitutions in a framework region thereof, wherein the heavy chain variable region comprises: a HCDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 7; a HCDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 8; and a HCDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 9; and a light chain of the anti-B7H3 antibody or fragment thereof comprises or consists of the amino acid sequence of SEQ ID NO: 16 or an amino acid sequence having at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%identity thereto and/or one or more (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10) amino acid additions, deletions and/or substitutions in a framework region thereof, wherein the light chain variable region comprises: a LCDR1 comprising or consisting of the amino acid sequence of SEQ ID NO: 10; a LCDR2 comprising or consisting of the amino acid sequence of SEQ ID NO: 11; and a LCDR3 comprising or consisting of the amino acid sequence of SEQ ID NO: 12.

In some embodiments, a heavy chain of the anti-B7H3 antibody or fragment thereof comprises or consists of the amino acid sequence of SEQ ID NO: 15; and a light chain of the anti-B7H3 antibody or fragment thereof comprises or consists of the amino acid sequence of SEQ ID NO: 16. In some embodiments, a heavy chain of the anti-B7H3 antibody or fragment thereof consists of the amino acid sequence of SEQ ID NO: 15; and a light chain of the anti-B7H3 antibody or fragment thereof consists of the amino acid sequence of SEQ ID NO: 16.

In some embodiments, the anti-B7H3 antibody or fragment thereof comprises or consists of two heavy chains comprising or consisting of the amino acid sequence of SEQ ID NO: 15; and two light chain comprising or consisting of the amino acid sequence of SEQ I D NO: 16. In some embodiments, the anti-B7H3 antibody or fragment thereof comprises or consists of two heavy chains consisting of the amino acid sequence of SEQ ID NO: 15; and two light chains consisting of the amino acid sequence of SEQ ID NO: 16.

Modified Fc region

In some embodiments, the anti-B7-H3 antibody or fragment thereof, e.g. via the Fc region, binds to one or more or all of the Fc receptors. The Fc receptors may comprise one or more or all of FcγRI (CD64) , FcγRIIa (CD32A) , FcγRIIb (CD32B) , FcγRIII (CD16) , C1q and FcRn. In some embodiments, the anti-B7-H3 antibody or fragment thereof, e.g. via the Fc region, binds to FcγRI.

It will be understood that the Fc region may interact with Fc receptors presented on the surface of a cell and/or may interact with proteins of the complement system. The Fc receptors may be Fc gamma receptors, e.g. FcγRI. The proteins of the complement system may include C1q.

In other embodiments the Fc region of the anti-B7-H3 antibody or fragment thereof is silenced to reduce, negate or abolish one or more Fc receptor binding and/or functionalities. In some embodiments, the Fc region of the anti-B7-H3 antibody or fragment thereof is modified to negate one or more Fc receptor functionalities. In some embodiments, the Fc region of the anti-B7-H3 antibody or fragment thereof is silenced in respect of one or more or all of FcγRI (CD64) , FcγRIIa (CD32A) , FcγRIIb (CD32B) , FcγRIII (CD16) and C1q functionality.

Thus, in some embodiments, the Fc region of the anti-B7-H3 antibody or fragment thereof is a modified Fc region.

In some embodiments, the Fc region of the anti-B7-H3 antibody or fragment thereof according to the invention may not be capable of binding to immune cells and/or recruiting immune cells.

The binding of the modified Fc region to FcγRI (CD64) , FcγRIIa (CD32A) , FcγRIIb (CD32B) , FcγRIII (CD16) , C1q and FcRn may be reduced compared to a wild-type Fc region.

In some embodiments, the binding of the modified Fc region to FcγRI may be reduced compared to a wild-type Fc region.

By “reduced binding” is meant at least 10%, at least 20%, at least 30%, at least 40%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%or 100%reduced binding compared to a wild-type Fc region.

In an embodiment, the anti-B7-H3 antibody or fragment thereof comprises an Fc region comprising an amino acid substitution at position 234, or an amino acid substitution at position 235, or an amino acid substitution at positions 234 and 235, according to the Eu numbering scheme. The Fc region may comprise further amino acid substitutions, such as at positions 322, 327, 330 and/or 331. The substitution (s) may be relative to a wild-type Fc region.

In some embodiments, the anti-B7-H3 antibody or fragment thereof comprises an Fc region comprising amino acid substitutions at positions 234 and 235, according to the Eu numbering scheme.

In some such embodiments, the Fc region of the anti-B7-H3 antibody or fragment thereof comprises a silencing modification selected from the STR mutation, the LALA mutation, the LALA-ΔA mutation and the LALA-KA mutation as defined herein.

The amino acid substitution (s) may be selected from the group consisting of:

a) L234A, L235A, A327G, A330S, and P331S (LALA-ΔA) ;

b) L234A, L235A, and K322A (LALA-KA) ; and

c) L234A and L235A (LALA) .

In preferred embodiments, the Fc region of the anti-B7-H3 antibody or fragment thereof comprises the amino acid substitutions L234A and L235A (LALA) .

Linker-payload, linker and payload moieties

The moiety of the antibody-drug conjugate of formula (I) above comprises a linker L, which is a chemical structural fragment, which is linked to the antibody at one end and linked to a cytotoxic drug at the other end, or linked to other linkers and then linked to the cytotoxic drug. The direct or indirect linking of a ligand may mean that the group is directly linked to the ligand via a covalent bond, and may also be linked to the ligand via a linker structure.

The linker structure is a structure shown as -L_a-L_b-L_c-as defined herein.

The moiety of the antibody-drug conjugate of formula (I) above contains a drug linked to the antibody (Ab) via linker L. The remaining moieties of the antibody-drug conjugate, i.e. all those structures including R¹ and all structures to the right of it in formula (I) , are termed “the drug moiety” or “the payload moiety” . In use, the bond between L and R¹ is cleaved, and R¹ and all structures to the right of it are released in vivo.

In the compounds used in the invention, X is -L¹-CH₂-C (O) -.

In the compounds used in the invention, L¹ is - (C (R^3a) (R^3b) ) _m-.

In one embodiment, m is not 0. In one embodiment, m is 1.

In one embodiment, each R^3a and each R^3b are not both hydrogen.

In one embodiment, R^3a is a C_1-6 aliphatic group. In one embodiment, R^3a is a C_1-6 alkyl group. In one embodiment, R^3a is a C_1-4 alkyl group. In one embodiment, R^3a is a C_1-3 alkyl group. In one embodiment, R^3a is methyl or ethyl. In one embodiment, R^3a is methyl.

In one embodiment, R^3b is hydrogen.

In one embodiment, X is -L¹-CH₂-C (O) -, L¹ is - (C (R^3a) (R^3b) ) _m-, m is not 0, and each R^3a and each R^3b are not both hydrogen.

In one embodiment, m is 1, and L¹ is -C (R^3a) (R^3b) -.

In one embodiment, R^3a is a C_1-6 aliphatic group, and R^3b is hydrogen.

In one embodiment, R^3a is methyl, and R^3b is hydrogen.

In one embodiment, R¹ is -O-.

In one embodiment, the antibody-drug conjugate comprises the following structure:

In one embodiment, the payload compound released is a compound of formula (III) :

Dose

In some embodiments of the invention, the antibody-drug conjugate as defined herein or pharmaceutically acceptable salt thereof may be administered to a patient at any dose.

In some embodiments, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered at a dose in the range of about 3 mg/kg to about 15 mg/kg. In some embodiments, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered at a dose in the range of about 4 mg/kg to about 14 mg/kg. In some embodiments, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered at a dose in the range of about 5 mg/kg to about 13 mg/kg. In some embodiments, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered at a dose in the range of about 6 mg/kg to about 12 mg/kg. In some embodiments, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered at a dose in the range of about 7 mg/kg to about 11 mg/kg. In some embodiments, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered at a dose in the range of about 8 mg/kg to about 10 mg/kg. In some embodiments, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered at a dose in the range of about 8.5 mg/kg to about 9.5 mg/kg.

In some embodiments, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered at a dose of about 9 mg/kg. In some embodiments, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered at a dose of 9 mg/kg.

In some embodiments, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered at a dose in the range of about 3 mg/kg to about 15 mg/kg. In some embodiments, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered at a dose in the range of about 3 mg/kg to about 12 mg/kg. In some embodiments, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered at a dose in the range of about 4 mg/kg to about 8 mg/kg. In some embodiments, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered at a dose in the range of about 5 mg/kg to about 7 mg/kg. In some embodiments, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered at a dose in the range of about 5.5 mg/kg to about 6.5 mg/kg.

In some embodiments, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered at a dose of about 6 mg/kg. In some embodiments, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered at a dose of 6 mg/kg.

In some embodiments, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered at a dose of about 6 mg/kg. In some embodiments, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered at a dose of about 9 mg/kg. In some embodiments, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered at a dose of about 12 mg/kg. In some embodiments, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered at a dose of about 15 mg/kg.

In some embodiments, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered at a dose of 6 mg/kg. In some embodiments, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered at a dose of 9 mg/kg. In some embodiments, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered at a dose of 12 mg/kg. In some embodiments, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered at a dose of 15 mg/kg.

In some embodiments, the antibody-drug conjugate is preferably the compound of formula (I) , or a pharmaceutically acceptable salt thereof.

In some embodiments, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered once every three weeks.

It will be understood that the dose may be selected based on considerations of age, body weight, disease symptoms, disease progression and/or severity, sex, and/or any other factors which may interfere with the therapeutic effects the antibody-drug conjugate or pharmaceutically acceptable salt thereof according to the invention.

Administration and treatment cycles

In some embodiments, the form of administration of the antibody-drug conjugate or pharmaceutically acceptable salt thereof, or the composition containing it, may be, for example, in a form suitable for oral, parenteral, intraperitoneal, systemic, intravenous (such as intravenous infusion or intravenous drip) , intramuscular, subcutaneous, topical, inhalative, rectal, sublingual, transdermal, or vaginal administration.

In some embodiments, the antibody-drug conjugate or pharmaceutically acceptable salt thereof, or the composition containing it, may be administered intravenously (e.g. by being injected into a subject) .

In some embodiments, the antibody-drug conjugate or pharmaceutically acceptable salt thereof, or the composition containing it, may be administered to a subject once (i.e. as a one-off treatment) . For example, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered to a subject once over a continuous period of hours or days.

In some embodiments, the antibody-drug conjugate or pharmaceutically acceptable salt thereof, or the composition containing it, may be administered to a subject on multiple, separate occasions (e.g. as part of an on-going treatment) . For example, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered to a subject on multiple, separate occasions over a total period of hours, days, weeks, months or years (generally described herein as “treatment cycles” or “exposure cycles” ) .

The term “treatment cycle” takes its normal meaning in the field of oncology to mean a period of time over which an anti-cancer drug is administered, followed by a rest period during which the drug is not administered.

In some embodiments of the invention, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered over a number of treatment cycles (as defined above) . In each treatment cycle, the period over which the antibody-drug conjugate or pharmaceutically acceptable salt thereof is administered is preferably 30 minutes to 3 hours, more preferably 40 minutes to 2 hours, more preferably 60 to 90 minutes, and the rest period is preferably 7 to 35 days, more preferably 14 to 28 days, even more preferably 18 to 23 days, most preferably 21 days.

The antibody-drug conjugate or composition according to the invention is preferably administered to the subject once every 3 weeks (Q3W) .

In some embodiments of the invention, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered in at least 1 treatment cycle. In some embodiments of the invention, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered in at least 2 treatment cycles. In some embodiments of the invention, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered in at least 3 treatment cycles. In some embodiments of the invention, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered in at least 4 treatment cycles. In some embodiments of the invention, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered in at least 5 treatment cycles. In some embodiments of the invention, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered in at least 6 treatment cycles. In some embodiments of the invention, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered in at least 7 treatment cycles. In some embodiments of the invention, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered in at least 8 treatment cycles. In some embodiments of the invention, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered in at least 9 treatment cycles. In some embodiments of the invention, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered in at least 10 treatment cycles. In some embodiments of the invention, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered in at least 11 treatment cycles. In some embodiments of the invention, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered in at least 12 treatment cycles. In some embodiments of the invention, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered in at least 13 treatment cycles. In some embodiments of the invention, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered in at least 14 treatment cycles. In some embodiments of the invention, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered in at least 15 treatment cycles. In some embodiments of the invention, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered in at least 16 treatment cycles. In some embodiments of the invention, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered in at least 17 treatment cycles. In some embodiments of the invention, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered in at least 18 treatment cycles. In some embodiments of the invention, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered in at least 19 treatment cycles. In some embodiments of the invention, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered in at least 20 treatment cycles. In these embodiments, the antibody-drug conjugate is preferably administered once every 3 weeks. In these embodiments, the antibody-drug conjugate is preferably the compound of formula (I) , or a pharmaceutically acceptable salt thereof.

As is known to the skilled person, there is no upper limit on the number of treatment cycles -treatment is continued until disease progression or unacceptable toxicity occurs, or the patient or oncologist decides to discontinue administration of the antibody-drug conjugate.

In some embodiments of the invention, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered in from 1 to 20 treatment cycles. In some embodiments of the invention, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered in from 2 to 20 treatment cycles. In some embodiments of the invention, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered in from 3 to 18 treatment cycles. In some embodiments of the invention, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered in from 4 to 14 treatment cycles. In some embodiments of the invention, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered in from 1 to 15 treatment cycles. In some embodiments of the invention, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered in from 2 to 12 treatment cycles. In some embodiments of the invention, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered in from 3 to 11 treatment cycles. In some embodiments of the invention, the antibody-drug conjugate or pharmaceutically acceptable salt thereof may be administered in 1 to 6 treatment cycles. In these embodiments, the antibody-drug conjugate is preferably administered once every 3 weeks. In these embodiments, the antibody-drug conjugate is preferably the compound of formula (I) , or a pharmaceutically acceptable salt thereof.

Pharmaceutical composition

The present invention also provides a composition comprising the ADC or pharmaceutically acceptable salt thereof as defined herein, for use according to the invention. The composition may be a pharmaceutical composition.

In some embodiments, the pharmaceutical composition according to the invention may comprise one or more carriers and/or excipients, in addition to the antibody-drug conjugate or pharmaceutically acceptable salt thereof according to the invention.

In some embodiments, the compositions described herein may further comprise one or more selected from this list consisting of: an adjuvant, salt, active polypeptide, compound, component and active agent.

Compositions typically should be sterile and stable under the conditions of manufacture and storage. The composition according to the invention may be produced using current good manufacturing practices (CGMP) .

The pharmaceutical composition may be formulated to be suitable for administration to a patient in order to prevent and/or treat disease. Pharmaceutical compositions can be formulated for administration by different routes, for example, for oral, parenteral, topical, inhalative, intravenous, intramuscular, rectal, sublingual, transdermal, subcutaneous, intratumoral application routes, according to their chemical and physical properties. In some embodiments, the pharmaceutical composition may be in a form suitable for intravenous infusion. In some embodiments, the pharmaceutical composition may be administered intravenously.

The pharmaceutical composition may be in the form of a tablet, a coated tablet, powder, granulate, a pellet, a capsule, an effervescent tablet or a transdermal therapeutic system. The pharmaceutical composition may be in the form of a liquid composition, selected from the group consisting of a solution, a syrup, an infusion, an extract, a solution for intravenous application, or a solution for infusion. The pharmaceutical composition may be in the form of a semisolid composition such as an emulsion, a suspension, a cream, a lotion, a gel, a globule, a buccal tablet or a suppository.

The term “carrier” , as used herein, may refer to a diluent, adjuvant, excipient, or vehicle. Such carriers can be sterile liquids, such as saline solutions in water and oils, including those of petroleum, animal, vegetable or synthetic origin, such as peanut oil, soybean oil, mineral oil, sesame oil. A sterile saline solution is a preferred carrier.

Saline solutions and aqueous dextrose and glycerol solutions can also be employed as liquid carriers. Suitable excipients include starch, glucose, lactose, sucrose, gelatin, malt, rice, flour, chalk, silica gel, sodium stearate, glyceryl monostearate, talc, sodium chloride, dried skim milk, glycerol, propylene, glycol, water, ethanol and the like.

The composition, if desired, can also contain minor amounts of wetting or emulsifying agents, or pH buffering agents. The composition of the invention can be formulated as neutral or salt forms. Salts include those formed with free amino groups such as those derived from hydrochloric, phosphoric, acetic, oxalic, tartaric acids, etc., and those formed with free carboxyl groups such as those derived from sodium, potassium, ammonium, calcium, ferric hydroxides, isopropylamine, triethylamine, 2-ethylamino ethanol, histidine, procaine, etc.

In some embodiments, the salt may comprise a metal cation, such as a sodium salt or a potassium salt.

In some embodiments, the composition may comprise an aqueous diluent or solvent. In some embodiments, the aqueous diluent or solvent may be a phosphate buffered saline solution, such as a sterile phosphate buffered saline solution.

In some embodiments, the composition may comprise a sterile aqueous buffer.

Kit

The present invention provides a kit comprising the ADC according to the invention.

The present invention provides a kit comprising the composition according to the invention.

In some embodiments, the kit may optionally comprise instructions for using the kit to target the one or more payload (s) to a cell expressing B7-H3.

Method of treatment

According to the present invention, the antibody-drug conjugate or pharmaceutically acceptable salt thereof, or composition containing it, as defined herein, are for use in treating cancer.

The invention further provides a method of treating cancer in a subject, comprising administering the ADC as defined herein to a subject.

The invention further provides a method of treating cancer in a subject, comprising administering the composition as defined herein to a subject.

The invention further provides for use of the ADC as defined herein in the manufacture of a medicament for the treatment of cancer.

The invention further provides for use of the composition as defined herein in the manufacture of a medicament for the treatment of cancer.

The cancer may express B7-H3. In some embodiments, the expression of B7-H3 is increased compared to the expression of B7-H3 by the same non-cancerous tissue or cells.

An increase refers to an increase by at least 10%, in particular at least 20%, at least 50%, at least 100%, at least 200%, at least 500%, at least 1000%, at least 10000%or even more. In one embodiment, expression is only found in a diseased tissue, while expression in a corresponding healthy tissue is repressed. For example, B7-H3 is expressed in cancer tissue while expression is not detectable in non-cancerous tissue. According to the invention, diseases associated with cells expressing B7-H3 include cancer diseases. Furthermore, according to the invention, cancer diseases preferably are those wherein the cancer cells express B7-H3.

In some embodiments, the cancer is selected from the group comprising non-small cell lung cancer (NSCLC) , small cell lung cancer (SCLC) , castration-resistant prostate cancer (CRPC) , oesophageal squamous cell carcinoma (ESCC) , melanoma, hepatocellular carcinoma (HCC) , cervical cancer, and other solid tumours.

In some embodiments, the cancer and/or solid tumour is selected from lung cancer, breast cancer, glioma, colon adenocarcinoma, melanoma, or prostate cancer.

In some embodiments, the cancer is selected from the group consisting of NSCLC, SCLC, CRPC, ESCC and melanoma.

In some embodiments, the cancer is NSCLC.

In some embodiments, the cancer is SCLC.

In some embodiments, the cancer is CRPC.

In some embodiments, the cancer is ESCC.

In some embodiments, the cancer is melanoma.

In some embodiments, the cancer is advanced and/or metastatic.

In some embodiments, the cancer is unresectable.

In some embodiments, the cancer has relapsed. In some embodiments, the cancer has progressed on or after standard systemic treatment.

In some embodiments, the cancer is refractory. In some embodiments, the cancer is intolerable to standard treatment.

The standard systemic treatment may be chemotherapy, immunotherapy and/or targeted therapy.

In some embodiments, the chemotherapy is platinum-based chemotherapy. In some embodiments, the immunotherapy is an immune checkpoint inhibitor, such as anti-PD-1/PD-L1 monoclonal antibody. In some embodiments the targeted therapy is surgery or radiation.

According to the invention, the term “B7-H3-positive cancer” means a cancer involving cancer cells expressing B7-H3, preferably on the surface of said cancer cells.

“Cell surface” is used in accordance with its normal meaning in the art, and thus includes the outside of the cell which is accessible to binding by proteins and other molecules. For example, a transmembrane protein having one or more extracellular portions is considered as being expressed on the cell surface.

B7-H3 is expressed on the surface of cells if it is located at the surface of said cells and is accessible to binding by B7-H3-specific antibodies added to the cells which have not been disrupted.

According to the invention, the term “disease” refers to any pathological state, including cancer, in particular those forms of cancer described herein. Any reference herein to cancer or particular forms of cancer also includes cancer metastasis thereof. In a preferred embodiment, a disease to be treated according to the present application involves cells expressing B7-H3.

“Diseases associated with cells expressing B7-H3” or similar expressions means according to the invention that B7-H3 is expressed in cells of a diseased tissue or organ.

As used herein, a “cancer disease” or “cancer” includes a disease characterized by aberrantly regulated cellular growth, proliferation, differentiation, adhesion, and/or migration. By “cancer cell” is meant an abnormal cell that grows by a rapid, uncontrolled cellular proliferation and continues to grow after the stimuli that initiated the new growth cease. Preferably, a “cancer disease” is characterized by cells expressing B7-H3 and a cancer cell expresses B7-H3. A cell expressing B7-H3 preferably is a cancer cell, preferably of the cancers described herein.

According to the invention, the term “cancer” also includes cancer metastasis of a primary tumour such as primary prostate cancer. Thus, if reference is made, for example, to prostate cancer, this also includes metastasis of the prostate cancer, for example metastasis to the lung, liver and/or lymph nodes.

By “metastasis” is meant the spread of cancer cells from its original site to another part of the body. The formation of metastasis is a very complex process and depends on detachment of malignant cells from the primary tumour, invasion of the extracellular matrix, penetration of the endothelial basement membranes to enter the body cavity and vessels, and then, after being transported by the blood, infiltration of target organs. Finally, the growth of a new tumour at the target site depends on angiogenesis. Tumour metastasis often occurs even after the removal of the primary tumour because tumour cells or components may remain and develop metastatic potential. In one embodiment, the term “metastasis” according to the invention relates to “distant metastasis” which relates to a metastasis which is remote from the primary tumour and the regional lymph node system. In one embodiment, the term “metastasis” according to the invention relates to lymph node metastasis. One particular form of metastasis which is treatable using the therapy of the invention is metastasis originating from prostate cancer as primary site. In preferred embodiments such prostate cancer metastasis is metastasis into lymph nodes, metastasis into lung and/or metastasis into liver.

A refractory cancer is a malignancy for which a particular treatment is ineffective, which is either initially unresponsive to treatment, or which becomes unresponsive over time.

An unresectable cancer is that which cannot be removed completely through surgery.

An advanced cancer is a cancer which has spread (metastasised) or recurred.

By “treat” is meant to administer a compound or composition or a combination of compounds or compositions to a subject in order to prevent or eliminate a disease, including reducing the size of a tumour or the number of tumours in a subject; arrest or slow a disease in a subject; inhibit or slow the development of a new disease in a subject; decrease the frequency or severity of symptoms and/or recurrences in a subject who currently has or who previously has had a disease; and/or prolong, i.e., increase the lifespan of the subject.

In particular, the term “treatment of a disease” includes curing, shortening the duration, ameliorating, preventing, slowing down or inhibiting progression or worsening, or preventing or delaying the onset of a disease or the symptoms thereof.

In some embodiments, the antibody-drug conjugate or pharmaceutically acceptable salt thereof according to the invention may be used for the purpose of diagnostics.

Subject

In this specification, the terms “subject” and “patient” are synonymous.

The term “patient” means according to the invention a subject for treatment, in particular a diseased subject, including human beings, nonhuman primates or other animals, in particular mammals such as cows, horses, pigs, sheep, goats, dogs, cats or rodents such as mice and rats.

In a particularly preferred embodiment, the subject is a human.

In some embodiments, the subject may suffer from and/or have been diagnosed with one or more cancer (s) . The subject may suffer from a histologically or cytologically confirmed cancer and/or solid tumour. The subject may suffer from and/or have been diagnosed with a pathologically documented cancer and/or solid tumour.

In some embodiments, the subject, which may be a human, is aged ≥18 years.

In some embodiments, the subject has a histologically or cytologically confirmed unresectable advanced/metastatic solid tumour, who has relapsed or progressed on or after standard systemic treatment, or is intolerable to standard treatment, or for whom no standard treatment is available.

In some embodiments, the subject has at least 1 measurable lesion per RECIST v1.1.

In some embodiments, the subject has Eastern Cooperative Oncology Group (ECOG) performance status (PS) of 0-1.

In some embodiments, the subject has received prior treatment for cancer. In some embodiments, the subject has received at least one line of prior treatment in the metastatic setting.

In some embodiments, the subject has received prior chemotherapy. In some embodiments, the subject has received prior immunotherapy. In some embodiments, the subject has received prior targeted therapy.

In some embodiments, the subject has received prior treatment with a platinum-based chemotherapy, such as cisplatin, oxaliplatin, and carboplatin.

In some embodiments, the subject has received prior treatment with new hormonal agents (NHA) , such as abiraterone acetate and enzalutamide.

In some embodiments, the subject has received prior treatment with a taxane, such as paclitaxel, docetaxel or cabazitaxel.

In some embodiments, the subject has received prior treatment with an immune checkpoint inhibitor, such as anti-PD-1/PD-L1 monoclonal antibody.

In some embodiments, the subject has not received prior treatment with a B7-H3 targeted therapy.

In some embodiments, the subject has not received prior treatment with an ADC that has a topoisomerase inhibitor payload.

In some embodiments, the subject has a pathologically documented locally advanced, or metastatic SCLC that is not amenable to curative surgery or radiation, and has relapsed/progressed on or after two cycles of platinum-based chemotherapy in combination with/without anti-PD-1/PD-L1 monoclonal antibody or is intolerant to completion of 2 cycles of platinum-based chemotherapy due to the toxicity.

In some embodiments, the subject has a pathologically documented locally advanced, or metastatic NSCLC which is not amenable to curative surgery or radiation, and has received prior treatment with platinum-based chemotherapy regimen and/or anti-PD-1/PD-L1 antibody-based regimen in the advanced/unresectable, or metastatic setting.

In some embodiments, the subject has a pathologically documented locally advanced or metastatic ESCC which is not amenable to curative surgery or radiation and has received at least one prior therapy for unresectable disease.

In some embodiments, the subject has a pathologically documented metastatic adenocarcinoma of the prostate and has received prior docetaxel and novel hormone therapy.

In some embodiments, the subject has a histologically or cytologically confirmed diagnosis of unresectable Stage III or metastatic melanoma not amenable to local therapy, and has received prior treatment with a PD-1 or PD-L1 inhibitor, or for melanoma with BRAF mutant, has received a prior treatment regimen that included vemurafenib, dabrafenib, or another BRAF gene and/or mitogen-activated protein kinase protein inhibitor.

In some embodiments, the subject has a histologically or cytologically confirmed solid tumour (s) and has progressed or relapsed after at least one prior standard therapeutic regimen.

EXAMPLES

Example 1 –Synthesis of Compounds

Preparation of Intermediate 1:

N- ( (7S, 15S) -7-benzyl-17- ( ( (1 S, 9S) -9-ethyl-5-fluoro-9-hydroxy-4-methyl-10, 13-dioxo-2, 3, 9, 10, 13, 15-hexahydro-1H, 12H-benzo [de] pyrano [3', 4': 6, 7] indolizino [1, 2-b] quinolin-1-yl) amino) -15-methyl-2, 5, 8, 11, 17-pentaoxo-14-oxa-3, 6, 9, 12-tetraazaheptadecyl) -6- (2, 5-dioxo-2, 5-dihydro-1H-pyrrol-1-yl) hexanamide

Step 1.

34a (5 g, 48.0 mmol) and K₂CO₃ (19.9 g, 144.0 mmol) were dissolved in DMF (20 mL) , followed by the dropwise addition of benzyl bromide (12.3 g, 72.0 mmol) . The mixture was reacted at 25 ℃ for 17 h. After the starting material was consumed completely as detected by TLC (PE/EA = 3/1) , the reaction solution was added to water (200 mL) , extracted with EA (250 mL) , separated and washed with saturated NaCl. The organic phase was dried over anhydrous Na₂SO₄, concentrated and purified by column chromatography (PE: EA = 2: 1) to give 34b (8.7 g, yield 93%) as a colourless liquid. MS-ESI: m/z 195.1 [M+H] +.

Step 2.

43c (7.3 g, 19.8 mmol) and TsOH (1.46 g, 8.5 mmol) were dissolved in THF (20 mL) , and the mixture was cooled to 0 ℃ under nitrogen atmosphere, added dropwise with a solution of 34b (7.7 g, 39.6 mmol) in THF (10 mL) and reacted at 0 ℃ for 2 h after the addition was completed. After most of the starting material was consumed as detected by TLC (PE/EA = 2/1) , the reaction solution was poured into water (100 mL) , extracted with DCM (100 mL) , separated and washed with saturated NaCl. The organic phase was dried over anhydrous Na₂SO₄ and purified by column chromatography (PE/EA = 1/1) to give 34d (3.9 g, yield 39%) as a colourless sticky substance. MS-ESI: m/z 503.3 [M+H] +.

Step 3.

Pd/C (1 g, 10 wt. %) was added to a mixed solution of 34d (1.9 g, 3.78 mmol) in EtOH (100 mL) and EA (100 mL) at 0 ℃ under hydrogen atmosphere, and the mixture was reacted at 0 ℃ for 3 h. After the reaction was completed as detected by TLC (PE/EA = 2/1) , the reaction solution was filtered throughand the filter cake was washed with EA/EtOH (1: 1, 100 mL × 3) . The filtrate was concentrated, and dissolved with THF (50 mL × 3) and dried by rotary evaporation, which was repeated three times, to give 34e (1 g, yield 64%) as a grey solid. MS-ESI: m/z 435.2 [M+H] +.

Step 4.

DIEA (303 mg, 2.35 mmol) was added dropwise to a solution of 34e (426 mg, 1.03 mmol) , exatecan methanesulfonate (KI4) (500 mg, 0.94 mmol) and HATU (429 mg, 1.13 mmol) in DMF (20 mL) at 0 ℃ under nitrogen atmosphere, and the mixture was reacted at 0 ℃ for 2 h after the addition was completed. After the reaction was completed as detected by LCMS, the reaction solution was added dropwise to water (300 mL) and stirred. The resulting mixture was left to stand for 5 min and filtered, and the filter cake was dissolved with DCM/MeOH (10: 1, 100 mL) solution, dried by rotary evaporation, mixed with silica gel and purified by column chromatography (EA: MeOH = 30: 1) to give 34f (600 mg, yield 77%) as a yellow solid. MS-ESI: m/z 830.3 [M+H] +.

Step 5.

Diethylamine (5 mL) was added dropwise to a solution of 34f (150 mg, 0.18 mmol) in DCM (5 mL) at 0 ℃ under nitrogen atmosphere, and the mixture was reacted at 0 ℃ for 2 h. After the reaction was completed as detected by LCMS, a petroleum ether solution (100 mL × 6) was added to the reaction solution, and a solid was precipitated. The resulting mixture was left to stand until the solid was adsorbed on the bottom of the flask, and the solution was poured out and dried with an oil pump to give 34g (120 mg, yield 76%) as a white powder, with the product content of 70%as detected by LCMS. MS-ESI: m/z 608.3 [M+H] +.

Step 6.

A solution of HATU (45 mg, 0.118 mmol) in DMF (1 mL) was added to a solution of 34g (60 mg, 0.099 mmol) , 43h (51 mg, 0.108 mmol) and DIEA (32 mg, 0.25 mmol) in DMF (1 mL) at 0 ℃ under nitrogen atmosphere, and the mixture was reacted at 0 ℃ for 2 h. After the starting material was consumed completely as detected by LCMS, the reaction solution was directly purified by reverse-phase column chromatography (eluent: (MeCN/MeOH = 1/1) : H₂O =60%: 40%) to give L-III-30 (14.8 mg, yield 14%) as a yellow solid.

MS-ESI: m/z 1062.4 [M+H] +.

1H NMR (400 MHz, Methanol-d4) δ 7.69 –7.61 (m, 2H) , 7.22 –7.16 (m, 2H) , 7.16 –7.09 (m, 3H) , 6.76 (s, 2H) , 5.70 –5.64 (m, 1H) , 5.60 (d, J = 16.4 Hz, 1H) , 5.40 –5.31 (m, 2H) , 5.26 (d, J = 19.0 Hz, 1H) , 4.65 –4.50 (m, 7H) , 4.25 –4.16 (m, 1H) , 3.87 (d, J = 16.7 Hz, 1H) , 3.83 – 3.76 (m, 3H) , 3.72 (d, J = 17.0 Hz, 2H) , 3.44 (t, J = 7.1 Hz, 2H) , 3.25 –3.17 (m, 2H) , 3.10 –3.02 (m, 1H) , 2.92 –2.83 (m, 1H) , 2.45 –2.39 (m, 5H) , 2.32 –2.20 (m, 5H) , 1.97 –1.89 (m, 2H) , 1.63 –1.50 (m, 4H) , 1.34 –1.20 (m, 6H) , 0.99 (t, J = 7.3 Hz, 3H) .

Synthesis of Compound 1:

A reducing agent and a protective agent were formulated using ultrapure water: a 2 mg/mL aqueous solution of TCEP (tris-2-carboxyethyl-phosphine, manufacturer: Thermo) and a 100 mmol/L aqueous solution of EDTA (disodium ethylenediaminetetraacetate, manufacturer: Sigma) .

Intermediate 1 was dissolved in anhydrous DMA (N, N-dimethylacetamide, manufacturer: Sinopharm) to prepare a 10 mg/mL solution of Intermediate 1 in DMA.

An anti-B7-H3 monoclonal antibody comprising a heavy chain comprising the sequence of SEQ ID NO: 15 and a light chain comprising the sequence of SEQ ID NO: 16 was produced as described in WO2023/236949. Using the ExpiFectamine^TM transient transfection kit, a plasmid encoding the antibody with a final concentration of 1 μg/mL was transiently transfected into 20 mL Expi293F cells, with a cell density of 3.0 × 10⁶ cells/mL and cell viability exceeding 95%. Cell cultures were grown in a humidified horizontal shaker with a rotation speed of 150 rpm. The temperature is maintained at 37℃, while the CO₂ level is maintained at 8%. After 5 days of incubation of the cell culture, the supernatant expressing the antibody was collected, filtered, and purified using a GE MabSelect SuRE protein A column (Cytiva-175438) . Eluted antibodies were dialyzed into PBS via D-Tube Dialyzer Maxi (EMD Millipore 71508, MWCO 3.5 kDa) . Antibody concentration was determined by absorbance at 280 nm on the NanoDrop device. 2 μg of purified antibody samples were run on SDS-PAGE gels (Invitrogen NuPAGE^TM 4%-12%Bis-Tris protein gel) with and without reducing agent. The purity of the purified antibody samples was quantified by HPLC-SEC using a TSKgel G3000SWXL size exclusion chromatography column (Tosoh 008541) . Purified antibodies were stored at -80℃.

20 mg of 11.3 mg/mL an anti-B7-H3 monoclonal antibody comprising a heavy chain comprising the sequence of SEQ ID NO: 15 and a light chain comprising the sequence of SEQ ID NO: 16 was weighed into a 50 mL centrifuge tube and diluted to 5 mg/mL by adding 30 mM His-HAc, pH 5.5 buffer. The 100 mM aqueous solution of EDTA was added in an amount of 5%of the total volume of the reaction mixture, and the resulting mixture was well mixed by shaking. Then the antibody was reduced by adding the 2 mg/mL aqueous solution of TCEP in a TCEP-to-antibody molar ratio of 8: 1, and the resulting mixture was well mixed by shaking and reacted on a cooling thermomixer at 37 ℃ for 2 h. The above solution of Intermediate 1 in DMA was added in a drug-to-antibody final concentration molar ratio of 12: 1, and additional DMA was added in an amount of 10%of the total volume of the reaction mixture. The resulting mixture was well mixed by shaking and reacted on a cooling thermomixer at 4 ℃ for 1 h. Sample preservation buffer exchanges were performed using ultrafiltration tubes (MWCO 30 KD, manufacturer: Millipore) . First, ultrafiltration was performed three times using a 30 mM His-HAc, pH 5.5 buffer containing 10%DMSO, then ultrafiltration was performed six times using a 30 mM DMSO-free His-HAc, pH 5.5 buffer, and finally, filtration was performed through a 0.2 μm PES membrane to remove bacteria to give an antibody-drug conjugate Compound 1 (10.8 mg, at a concentration of 4.933 mg/mL, 54%yield) .

Tests showed that the drug to antibody ratio (DAR) of Compound 1 was 5.68 and the SEC purity was 99.57%.

Synthesis of reference ADC-1 (DS7300)

DS-7300 is composed of a humanized anti-B7-H3 IgG1 monoclonal antibody conjugated to a topoisomerase I inhibitor payload, an exatecan derivative deruxtecan, via a tetrapeptide-based cleavable linker.

Linker-payload (deruxtecan) was dissolved in anhydrous DMA (N, N-dimethylacetamide, manufacturer: Sinopharm) to prepare a 10 mg/mL solution of linker-payload in DMA.

16 mg of 7.4 mg/mL reference monoclonal anti-B7-H3 antibody (the antibody sequence comprised a heavy chain set forth in sequence No. 9 and a light chain set forth in sequence No. 16 in the patent CN104755494B; the antibody was produced by transfection of CHO cells followed by routine antibody expression and purification, with >95%purity) was weighed into a 50 mL centrifuge tube and diluted to 5 mg/mL by adding 30 mM His-HAc, pH 5.5 buffer. The 100 mM aqueous solution of EDTA was added in an amount of 5%of the total volume of the reaction mixture, and the resulting mixture was well mixed by shaking. Then the antibody was reduced by adding the 2 mg/mL aqueous solution of TCEP in a TCEP-to-antibody molar ratio of 2.7: 1, and the resulting mixture was well mixed by shaking and reacted on a cooling thermomixer at 37 ℃ for 2 h. The above solution of Linker-payload in DMA was added in a drug-to-antibody final concentration molar ratio of 9: 1, and additional DMA was added in an amount of 10%of the total volume of the reaction mixture. The resulting mixture was well mixed by shaking and reacted on a cooling thermomixer at 4 ℃ for 1 h. Sample preservation buffer exchanges were performed using ultrafiltration tubes (MWCO 30 KD, manufacturer: Millipore) . First, ultrafiltration was performed three times using a 30 mM His-HAc, pH 5.5 buffer containing 10%DMSO, then ultrafiltration was performed six times using a 30 mM DMSO-free His-HAc, pH 5.5 buffer, and finally, filtration was performed through a 0.2 μm PES membrane to remove bacteria to give an antibody-drug conjugate reference ADC-1 (11 mg, at a concentration of 5.825 mg/mL, 68.75%yield) .

Tests showed that the drug to antibody ratio (DAR) of reference ADC-1 was 4.59 and the SEC purity was 99.64%.

Example 2 –Phase 1/2a study of Compound 1 in all solid tumours

Background:

Compound 1, the synthesis of which is described in Example 1, is an antibody-drug conjugate composed of a humanized anti-B7 homologue (B7-H3, CD276) immunoglobulin G1 (IgG1) monoclonal antibody attached to a DNA topoisomerase I inhibitor via a cleavable linker. Here, we present the initial results of the first-in-human (FIH) study.

Methods:

As of March 2024, the Phase 1/2a, a multi-centre, open-label, multiple-dose, first-in-human study, is ongoing to assess the safety, tolerability, pharmacokinetics, and preliminary anti-tumour activity of Compound 1 in subjects with advanced/metastatic solid tumours. The study consists of two phases. Phase 1 (dose escalation) adopts an accelerated titration at the starting dose level of 3 mg/kg followed with the classic “3+3” design. Five increasing dose levels (i.e., 3 mg/kg, 6 mg/kg, 9 mg/kg, 12 mg/kg and 15 mg/kg) are studied in advanced/metastatic cancer subjects to identify maximum tolerated dose (MTD) and/or recommended phase 2 dose (RP2D) . Phase 2a (dose expansion) was started with advanced/unresectable, or metastatic small cell lung cancer (SCLC) , non-small cell lung cancer (NSCLC) , and castration-resistant prostate cancer (CRPC) subjects, which are being selected as the randomization cohorts for dose optimization. Other five cohorts are initiated at the dose recommended based on cumulative data from Phase 1 and the randomization cohorts to assess safety/tolerability and efficacy in subjects with advanced/unresectable, or metastatic oesophageal squamous cell carcinoma (ESCC) , melanoma, hepatocellular carcinoma (HCC) , cervical cancer (CC) , and other solid tumours.

Compound 1 is administered intravenously once every 3 weeks until disease progression, withdrawal of consent, or unacceptable toxicity. Adverse events (AEs) are coded using MedDRA and graded according to the NCI-CTCAE version 5.0. Tumour responses are assessed according to the Response Evaluation Criteria in Solid Tumours (RECIST) version 1.1.

Results:

As of data cut-off (DCO) of March 2, 2024, 59 subjects (all were in phase 1) with metastatic solid tumours were enrolled and received at least one dose of Compound 1 across five dose levels (one in 3 mg/kg dose level, 26 each in 6 mg/kg and 9 mg/kg dose levels, and 6 in 12 mg/kg dose level) . Of these subjects, the median age was 60.0 (range: 38-78) years, 59.3%(35/59) subjects were previously treated with immunotherapy, and 40.7% (24/59) subjects were previously treated with targeted therapy. There was a median follow-up time of 1.5 (range: 0.3-5.9) months.

Overall clinical efficacy of solid tumours

As of DCO of March 2, 2024, of 32 efficacy-evaluable subjects (included all subjects who received at least one dose of Compound 1, had at least one measurable lesion per RECIST v1.1 at baseline, and post-baseline efficacy assessment or discontinued study treatment prior to the first post-baseline efficacy assessment) , the unconfirmed objective response rate (ORR) per RECIST v1.1 was 34.4% (95%CI: 18.57, 53.19) and the unconfirmed disease control rate (DCR) was 78.1% (95%CI: 60.03, 90.72) . The efficacy summary is presented in Table 1. More than 30%decrease tumour size from baseline were reported in 11 of 31 subjects (one missing) which were mainly at 6.0 mg/kg and 9.0mg/kg dose cohorts (Figure 1) .

Clinical efficacy of SCLC

As of DCO of March 2, 2024, of the 21 enrolled SCLC subjects, 13 were efficacy-evaluable and treated with Compound 1 in doses ranging from 6-12 mg/kg in the dose-escalation (10 subjects in 6 mg/kg, 2 in 9 mg/kg, and 1 in 12 mg/kg dose cohort) . The unconfirmed ORR and DCR were 46.2% (95%CI: 19.22, 74.87) and 92.3% (95%CI: 63.97, 99.81) , respectively (Table 2) . More than 30%decrease in tumour size from baseline were reported in 6 of 13 subjects which all were at 6.0 mg/kg (N=5) and 9.0mg/kg (N=1) dose cohorts (Figure 2) , median TTR was 1.51 (range, 1.22-3.02) months, median DoR was 5.47 (95%CI, 2.69-not evaluable) months, and median PFS was 5.45 (95%CI, 3.88-NA) months (see Table 2) .

Clinical efficacy of CRPC

As of DCO of March 2, 2024, 11 subjects were CRPC and 4 subjects were efficacy-evaluable and treated with Compound 1 in doses ranging from 6-9 mg/kg (four subjects in 6 mg/kg and 1 subject was in 9 mg/kg dose cohort) in the dose-escalation. Two subjects had PRs with ≥30%decrease in tumour size from baseline (Figure 3) , and one PR was reported at the first tumour assessment and remained in response at the DCO. The unconfirmed ORR and the unconfirmed DCR was 50.0% (95%CI: 6.76, 93.24) and 100.0% (95%CI: 39.76, 100.00) , respectively (Table 3) .

Table 1-Overall Clinical Efficacy in Compound 1 Clinical Study (modified Efficacy Analysis Set)

Note: BOR: best overall response; CR: complete response; PR: partial response; SD: stable disease; PD: progressive disease; NE: not evaluable; ORR: objective response rate; DCR: disease control rate.

Table 2 –Overall Clinical Efficacy in Subjects with SCLC in Compound 1 Clinical Study (modified Efficacy Analysis Set)

Table 3 –Overall Clinical Efficacy in Subjects with CRPC in Compound 1 Clinical Study (modified Efficacy Analysis Set)

Conclusion:

Compound 1 demonstrated a manageable safety profile and promising antitumour activity in subjects with advanced/metastatic solid tumours, especially in subjects with SCLC and CRPC.

Example 3 –Dose-range finding studies

Methods:

A Phase 1/2a, a multi-centre, open-label, multiple-dose, first-in-human study, is ongoing to assess the safety, tolerability, pharmacokinetics, and preliminary anti-tumour activity of Compound 1 in subjects with advanced/metastatic solid tumours. The study protocols are described in Example 2 above.

Results:

Justification for selecting 6.0 mg/kg and 9.0 mg/kg, every 3 weeks, as randomization dose levels in SCLC:

As of March 2, 2024, clinical data collected from 59 subjects in the ongoing Compound 1 study supports the selection of randomization dose levels.

● Efficacy: In the dose escalation part of this study, a total of 32 subjects were included in the Efficacy Analysis Set (EAS) at doses ranging from 6 to 12 mg/kg. One subject in 3 mg/kg dose group was excluded from the EAS due to missing tumour assessment data. 11 (34.4%) subjects achieved PRs, while 8 were in 6 mg/kg (n=8) dose group, 2 were in 9 mg/kg (n=2) dose group, and 1 was in 12 mg/kg dose group, with unconfirmed ORRs of 44.4% (8/18) , 18.2% (2/11) and 33.3% (1/3) , respectively. These results suggested that the 6 mg/kg and 12 mg/kg doses exhibited a more favourable efficacy profile compared with the 9 mg/kg dose. However, of the 13 SCLC subjects in the EAS, both 6 and 9 mg/kg dose groups had an unconfirmed ORR of 50.0%.

● Safety: In the dose escalation part of this study, a lower incidence of following TEAEs were observed in 6 mg/kg and 9 mg/kg dose groups compared with 12 mg/kg dose group: any grades of TRAE in 6 mg/kg vs. 9 mg/kg vs. 12 mg/kg: 88.5%vs. 65.4%vs. 83.3%; grade ≥3 TEAEs: 19.2%vs. 38.5%vs. 66.7%; grade ≥3 TRAEs: 11.5%vs. 19.2%vs. 66.7%; serious TEAEs: 7.7%vs. 19.2%vs. 50.0%; and serious TRAEs: 3.8%vs. 7.7%vs. 50.0%: 10.3%) . Therefore, the 6 mg/kg and 9 mg/kg doses show improved safety profiles than the 12 mg/kg dose.

● A good safety profile was identified in the 3 mg/kg dose group with no grade ≥3, serious TEAEs, however the 3 mg/kg dose was not considered for optimization because of the small sample size and the lack of efficacy data.

● To balance the safety and efficacy between the 6 mg/kg and 9 mg/kg Q3W, both were selected for the RP2D optimization in SCLC and are expected to also be tested in other indications such as CRPC.

Example 4 –Preclinical study of Compound 1 in the treatment of female BALB/c Nude mice bearing KYSE-150 human ESCC tumours

To investigate the efficacy of Compound 1 in the treatment of female BALB/c Nude mice bearing KYSE-150 tumours.

Female BALB/c Nude mice (purchased from Beijing Vital River Laboratory Animal Technology Co., LTD. ) were used as test animals. 5 × 10⁶ KYSE-150 tumour cells were inoculated subcutaneously into the right flank of 6-to 8-week-old female BALB/c Nude mice. The growth of tumours in mice was observed. When the volume of the tumours reached about 128 mm³, the tumour-bearing mice were randomized into groups with 5 mice in each of the blank control group and the treatment groups. Intravenous (i. v. ) injection of Compound 1 or reference ADC-1 was administered on the day of grouping (day 0, day 8, day 14) , and a total of 3 injections were administered. Doses administered were 1/2 mg/kg and 3/6 mg/kg. The experiment ended on day 21. Experimental grouping and dosing were as follows. The tumour volume and weight of mice were measured twice a week, and the data were recorded.

● Vehicle (negative control group) : normal saline

● Compound 1 (treatment group) : 1 mg/kg (day 0) ; 2 mg/kg (day 8) ; 2 mg/kg (day 14) ;

● Compound 1 (treatment group) : 3 mg/kg (day 0) ; 6 mg/kg (day 8) ; 6 mg/kg (day 14) ;

● Reference ADC-1 (positive control group) : 1 mg/kg (day 0) ; 2 mg/kg (day 8) ; 2 mg/kg (day 14) ;

● Reference ADC-1 (positive control group) : 3 mg/kg (day 0) ; 6 mg/kg (day 8) ; 6 mg/kg (day 14) ;

All samples were prepared by dilution with normal saline.

At the end of the experiment, mice were euthanized and tumour inhibition rate TGI was calculated as follows:

TGI (%) = [1 - (T_i -T₀) / (V_i -V₀) ] × 100

T_i: mean tumour volumes of the treatment groups and the positive control group on day i of administration;

T₀: mean tumour volumes of the treatment groups and the positive control group on day 0 of administration;

V_i: mean tumour volume of the negative control group on day i of administration;

V₀: mean tumour volume of the negative control group on day 0 of administration.

The experimental results are shown in Figure 4 and Table 4.

Table 4 –Evaluation of anti-tumour efficacy of compound 1 in KYSE-150 cell subcutaneous xenograft tumour-bearing mouse model (calculations based on tumour volumes on day 21 after administration)

Notes: tumour volumes are expressed as means ± standard errors; tumour growth inhibition was reflected by T/C (T/C (%) = T_i /V_i × 100) and TGI (TGI (%) = [1 - (T_i -T₀) / (V_i -V₀) ] × 100) ; p values were calculated from tumour volumes, ns indicates p>0.05. **indicates p<0.01. (p > 0.05 indicates no statistical difference, and p < 0.01 indicates a significant difference) .

The results in Figure 4 and Table 4 indicate that the antibody-drug conjugate Compound 1 of the present application showed significant anti-tumour activity and its anti-tumour effect was significantly better than that of reference ADC-1 at both the dose of 1/2 mg/kg and the dose of 3/6 mg/kg.

Example 5 –Preclinical study of Compound 1 in the treatment of NCG mice bearing DMS 53 human SCLC tumours

To investigate the anti-tumour efficacy of Compound 1 in the NCG mice model subcutaneously transplanted with DMS 53 tumours.

6.4-7.3 weeks female NCG mice (purchased from GemPharmatech Co., Ltd) were used as test animals. 7.5×10⁶ DMS 53 cells were inoculated subcutaneously into the right flank of NCG mice. The growth of tumours in mice was observed. When the volume of the tumours reached about 135.51 mm³, the tumour-bearing mice were randomized into groups with 5 mice in each of the blank control group and the treatment groups. Intravenous (i. v. ) injection of Compound 1 or reference ADC-1 was administered on the day of grouping (day 0, day 7) , and a total of 2 injections were administered. Doses administered were 3 mg/kg and 6 mg/kg. The experiment ended on day 28. Experimental grouping and dosing were as follows. The tumour volume and weight of mice were measured twice a week, and the data were recorded.

● G1: Vehicle (negative control group) : normal saline

● G2: Compound 1 (treatment group) : 3 mg/kg;

● G3: Compound 1 (treatment group) : 6 mg/kg;

● G4: Reference ADC-1 (positive control group) : 6 mg/kg;

All samples were prepared by dilution with normal saline.

The calculation of Tumour Volume Changes TGI_TV:

V_nt:Tumour volume on day t of mouse n;

V_n0: Tumour volume on day 0 of mouse n;

RTV_n:Relative tumour volume on day n of mouse n;

mean RTV_treat: Mean RTV of administration group;

mean RTV_vehicle: Mean RTV of vehicle group;

The experimental results are shown in Figure 5 and Table 5.

Table 5 -Evaluation of anti-tumour efficacy of compound 1 in DMS 53 cell subcutaneous xenograft tumour-bearing mouse model (calculations based on tumour volumes on day 28 after administration)

Notes: Tumour Volumes are expressed as means ± standard errors; p > 0.05 indicates no statistical difference, and p < 0.01 indicates a significant difference.

The results in Figure 5 and Table 5 indicate that the antibody-drug conjugate Compound 1 of the present application showed significant tumour growth inhibition (P<0.05) and its anti-tumour effect was significantly better than that of reference ADC-1.

Example 6. Evaluation of Efficacy of Antibody-Drug Conjugates in Mice Bearing Calu-6 Human Lung Cancer Cell Tumours

To investigate the inhibitory effect of Compound 1 on in vivo tumour formation, xenograft tumours were formed in mice using Calu-6 human lung cancer cells that positively expressed B7H3, and then the in vivo anti-tumour effect of Compound 1 was evaluated.

6-to 8-week-old female BALB/c Nude mice (purchased from Zhejiang Vital River Laboratory Animal Technology Co., Ltd. ) were used as test animals. 10 × 10⁶ Calu-6 lung cancer cells were inoculated subcutaneously into the right dorsa of 6-to 8-week-old female BALB/c Nude mice. The growth of tumours in mice was observed. When the volume of the tumours reached about 139 mm³, the tumour-bearing mice were randomized into groups with 6 mice in each of the blank control group and the treatment groups. Intravenous (i. v. ) injection of Compound 1 or reference ADC-1 was administered on the day of grouping (day 0) , and a total of one injection was administered. Doses administered were 1 mg/kg and 3 mg/kg. The experiment ended on day 28. Experimental grouping and dosing were as follows. The tumour volume and weight of mice were measured twice a week, and the data were recorded.

● Blank control (negative control group) : normal saline

● Compound 1 (treatment group) : 1 mg/kg

● Compound 1 (treatment group) : 3 mg/kg

● Reference ADC-1 (treatment group, positive control group) : 1 mg/kg

● Reference ADC-1 (treatment group, positive control group) : 3 mg/kg

All samples were prepared by dilution with normal saline.

At the end of the experiment, mice were euthanized and tumour inhibition rate TGI was calculated as above.

The experimental results are shown in Figure 6 and Table 6.

Table 6 -Evaluation of anti-tumour efficacy of test antibody-drug conjugates in Calu-6 cell subcutaneous xenograft tumour-bearing mouse model (calculations based on tumour volumes on day 24 after administration)

Notes: tumour volumes are expressed as means ± standard errors; tumour growth inhibition was reflected by T/C (T/C (%) = T_i /V_i × 100) and TGI (TGI (%) = [1 - (T_i -T₀) / (V_i -V₀) ] × 100) ; and p values were calculated from tumour volumes (p > 0.05 indicates no statistical difference, and p < 0.01 indicates a significant difference) .

The results in Figure 6 and Table 6 indicate that the antibody-drug conjugate Compound 1 of the present application showed significant dose-dependent anti-tumour activity after the administration of a single dose and its anti-tumour effect was significantly better than that of reference ADC-1 at both the dose of 1 mg/kg and the dose of 3 mg/kg.

Example 7. Evaluation of Efficacy of Antibody-Drug Conjugates in Mice Bearing A375 Human Melanoma Cell Tumours

To investigate the inhibitory effect of Compound 1 on in vivo tumour formation, xenograft tumours were formed in mice using A375 human melanoma cells that positively expressed B7H3, and then the in vivo anti-tumour effect of Compound 1 was evaluated.

6-to 8-week-old female NOD/SCID mice (purchased from Jiangsu GemPharmatech Co., Ltd. ) were used as test animals. 5 × 10⁶ A375 human melanoma cells were inoculated subcutaneously into the right dorsa of 6-to 8-week-old female NOD/SCID mice. The growth of tumours was observed. When the volume of the tumours reached about 89 mm³, the tumour-bearing mice were randomized into groups with 6 mice in each of the blank control group and the treatment groups. Intravenous (i. v. ) injection of Compound 1 or reference ADC- 1 was administered on the day of grouping (day 0) , and a total of one injection was administered. Doses administered were 1 mg/kg, 3 mg/kg and 10 mg/kg. The experiment ended on day 20. The tumour volume and weight of mice were measured twice a week, and the data were recorded.

● Blank control (negative control group) : normal saline

● Compound 1 (treatment group) : 1 mg/kg

● Compound 1 (treatment group) : 3 mg/kg

● Compound 1 (treatment group) : 10 mg/kg

● Reference ADC-1 (treatment group) : 3 mg/kg

● Reference ADC-1 (treatment group) : 10 mg/kg

All samples were prepared by dilution with normal saline.

At the end of the experiment, mice were euthanized and TGI_TV (relative tumour inhibition rate) was calculated. The TGI_TV (relative tumour inhibition rate) was calculated by the following formula:

wherein, mean RTV_treat: mean RTV of dosing group;

mean RTV_vehicle: mean RTV of vehicle group (in this example, the vehicle group is the group in which only normal saline was administered) ;

The RTV was calculated by the following formula:

wherein, V_nt: tumour volume in n-numbered mouse on day t;

V_n0: tumour volume in n-numbered mouse on day 0;

RTV_n:relative tumour volume in n-numbered mouse on day t.

The experimental results are shown in Figure 7 and Table 7.

Table 7 -Relative tumour inhibition rates (TGI_TV) in different groups

The results in Figure 7 and Table 7 indicate that the antibody-drug conjugate Compound 1 of the present application showed significant dose-dependent anti-tumour activity after the administration of a single dose.

Example 8. Evaluation of Efficacy of Antibody-Drug Conjugates in Mice Bearing PC-3 Human Prostate Cancer Cell Tumours

To investigate the inhibitory effect of Compound 1 on in vivo tumour formation, xenograft tumours were formed in mice using PC-3 human prostate cancer cells that positively expressed B7H3, and then the in vivo anti-tumour effect of Compound 1 was evaluated.

6-to 8-week-old female BALB/c Nude mice (purchased from Zhejiang Vital River Laboratory Animal Technology Co., Ltd. ) were used as test animals. 5 × 10⁶ PC-3 human prostate cancer cells were inoculated subcutaneously into the right sides of the necks or dorsa of 6-to 8-week-old female BALB/c Nude mice. The growth of tumours was observed. When the volume of the tumours reached about 155 mm³, the tumour-bearing mice were randomized into groups with 6 mice in each of the blank control group and the treatment groups. Intravenous (i. v. ) injection of Compound 1 or reference ADC-1 was administered on the day of grouping (day 0) , and a total of one injection was administered. Doses administered were 2 mg/kg and 6 mg/kg. The experiment ended on day 21. Experimental grouping and dosing were as follows. The tumour volume and weight of mice were measured twice a week, and the data were recorded.

● Blank control (negative control group) : normal saline

● Compound 1 (treatment group) : 2 mg/kg

● Compound 1 (treatment group) : 6 mg/kg

● Reference ADC-1 (treatment group, positive control group) : 2 mg/kg

● Reference ADC-1 (treatment group, positive control group) : 6 mg/kg

All samples were prepared by dilution with normal saline.

The experimental results are shown in Figure 8 and Table 8.

Table 8 -Evaluation of anti-tumour efficacy of test antibody-drug conjugates in PC-3 cell subcutaneous xenograft tumour-bearing mouse model (calculations based on tumour volumes on day 21 after administration)

The results in Figure 8 and Table 8 indicate that the antibody-drug conjugate Compound 1 of the present application showed significant dose-dependent anti-tumour activity after the administration of a single dose and its anti-tumour effect was significantly better than that of reference ADC-1 at both the dose of 2 mg/kg and the dose of 6 mg/kg.

Example 9. Evaluation of Efficacy of Antibody-Drug Conjugate in Prostate Cancer Patient-Derived Xenograft (PDX) Model

To investigate the efficacy of Compound 1 in a prostate cancer patient-derived xenograft model, patient-derived prostate cancer PR9586 and PR9587 tumour tissues (obtained from Crown Biotechnology (Zhongshan) Co., Ltd. ) was subcutaneously xenografted into NPG and NOG male mice, and the anti-tumour effect of Compound 1 was evaluated.

1. Test compounds and materials

● Blank control (control group) : normal saline

● Compound 1 (treatment group) : 10 mg/kg

2. Preparation method for test compounds: all samples were prepared by dilution with normal saline.

3. Experimental animals: NPG mice were purchased from Beijing Vitalstar Biotechnology Co., Ltd., and NOG mice were purchased from Beijing Vital River Laboratory Animal Technology Co., Ltd.

4. Procedures:

Patient-derived prostate cancer PR9586 and PR9587 tumour tissues were separately inoculated subcutaneously into NPG or NOG mice. When the tumours grew to about 500–800 mm³, the tumour tissues were collected and the mice were euthanized. The collected tumour tissues were cut into tumour masses of a diameter of 2–3 mm and then inoculated subcutaneously into the right anterior scapula of the NPG or NOG mice. When the mean tumour volume of the tumour mass-inoculated NPG or NOG mice reached about 100–200 mm³, the mice were randomized into groups with 6 mice in each of the blank control group and the treatment group.

Intravenous (i. v. ) injection of Compound 1 was administered on the day of grouping (day 0) , the administration was performed once every two weeks for a total of two injections at a single dose of 10 mg/kg. The experiment ended on day 25 (for mice inoculated with PR9586) or on day 48 (for mice inoculated with PR9587) . After the administration was started, the body weight and tumour size of the mice were measured twice a week. Calculation formula for tumour volume: Tumour volume (mm³) = 1/2 × (a× b²) (where a represents long diameter and b represents short diameter) .

TGI (%) = [1 - (T_i -T₀) / (V_i -V₀) ] × 100

wherein,

T_i: mean tumour volume of the treatment group on day i of administration;

T₀: mean tumour volume of the treatment group on day 0 of administration;

V₀:mean tumour volume of the negative control group on day 0 of administration.

5. Experimental results:

The experimental results are shown in Figure 9A, Figure 9B, Table 9 and Table 10.

Table 9 -Evaluation of anti-tumour efficacy of test antibody-drug conjugate in PR9586 patient-derived xenograft model (calculations based on tumour volumes on day 25 after first administration)

Note: p values were calculated from tumour volumes (p > 0.05 indicates no statistical difference, and p < 0.01 indicates a significant difference) .

Table 10 -Evaluation of anti-tumour efficacy of test antibody-drug conjugate in PR9587 patient-derived xenograft model (calculations based on tumour volumes on day 48 after first administration)

The results indicate that the antibody-drug conjugate Compound 1 of the present application showed significant anti-tumour activity after the administration.

Example 10. Evaluation of Efficacy of Antibody-Drug Conjugate in Prostate Cancer Micro Patient-Derived Xenograft Model

To investigate the efficacy of Compound 1 in a prostate cancer patient-derived xenograft model, the anti-tumour effect of Compound 1 was evaluated in a prostate cancer micro patient-derived xenograft mouse model.

1. Test compounds and materials

● Blank control (control group) : normal saline

● Compound 1 (treatment group) : 10 mg/kg

Collagenase digest:

The components in the table were mixed according to the manufacturer's instructions to prepare a collagenase digest.

3. Experimental animals: CB17 SCID mice, male, aged 6–8 weeks, weighing about 18–22 g, purchased from Beijing Vital River Laboratory Animal Technology Co., Ltd.

4. Procedures:

Tumour tissues of patient-derived prostate cancer transplantation models LD1-2027-410644 and LD1-2034-362055 (obtained from Shanghai LIDE Biotech., Co. Ltd. ) were inoculated subcutaneously into mice. When the tumours grew to 500–800 mm³, the tumour tissues were surgically and aseptically removed from the mice, and non-tumour tissues and necrotic tissues were removed in a biosafety cabinet to ensure the purity of the inoculated tumour tissues. The mice were euthanized. The treated tumour tissues were cut into small tumour masses of 1–3 mm³, and the tumour masses were digested with the collagenase digest at 37 ℃ for 1–2 h. The supernatant was removed by centrifugation at 1200 rpm for 3 min, and the cells were resuspended in 10 mL of PBS containing 1%FBS and counted on a hemocytometer. Murine cells were removed, and the supernatant was removed by centrifugation at 1200 rpm for 3 min. The cells were resuspended in an RPMI1640 cell culture medium and counted on a hemocytometer. The cell density was adjusted.

The cell suspension was placed into capsules. CB17 SCID mice were randomized into groups based on the weight, with 3 mice in each of the blank control group and the treatment group. Each mouse was inoculated subcutaneously with one capsule on each of the left and right sides, with a total of 6 capsules per group. Each of the capsules contained 5000 tumour cells. The day of inoculation was recorded as day 0, and the animals were subjected to a single administration by tail vein injection. The experiment was conducted for 10 days.

At the end of the experiment, the mice were euthanized. The capsules were removed and cell viability assay was performed with CellTiter-Glo, i.e., the capsules were cut into pieces, PBS and CellTiter-Glo^TM reagent with the same volume were added, and the luminescence value was measured. In the optical signal and system, the luminescence value is in direct proportion to the amount of ATP, which is positively correlated with the number of living cells, so the cell viability can be informed by detecting the ATP content.

5. Experimental results:

The experimental results are shown in Figure 10A, Figure 10B and Table 11.

Table 11 -Results of cell viability assay at the endpoint of the experiment

The results in Figure 10A, Figure 10B, and Table 11 show that Compound 1 had significant anti-tumour effect.

Claims

An antibody-drug conjugate of formula (I) :

or a tautomer, a mesomer, a racemate, an enantiomer or a diastereoisomer thereof, or a pharmaceutically acceptable salt thereof,

wherein:

Ab is an anti-B7-H3 antibody or a fragment thereof, wherein the anti-B7-H3 antibody or a fragment thereof comprises heavy chain complementarity determining regions (HCDRs) 1-3 and light chain complementarity determining regions (LCDRs) 1-3, defined:

(a) according to the Kabat numbering scheme, wherein:

HCDR1 comprises or consists of an amino acid sequence according to SEQ ID NO: 1, HCDR2 comprises or consists of an amino acid sequence according to SEQ ID NO: 2, HCDR3 comprises or consists of an amino acid sequence according to SEQ ID NO: 3, LCDR1 comprises or consists of an amino acid sequence according to SEQ ID NO: 4, LCDR2 comprises or consists of an amino acid sequence according to SEQ ID NO: 5, and LCDR3 comprises or consists of an amino acid sequence according to SEQ ID NO: 6; and/or

(b) according to the IMGT numbering scheme, wherein:

HCDR1 comprises or consists of an amino acid sequence according to SEQ ID NO: 7, HCDR2 comprises or consists of an amino acid sequence according to SEQ ID NO: 8, HCDR3 comprises or consists of an amino acid sequence according to SEQ ID NO: 9, LCDR1 comprises or consists of an amino acid sequence according to SEQ ID NO: 10, LCDR2 comprises or consists of an amino acid sequence according to SEQ ID NO: 11, and LCDR3 comprises or consists of an amino acid sequence according to SEQ ID NO: 12;

n is a connection number, and is an integer from 1 to 10;

L is -L_a-L_b-L_c-;

L_a-is

-L_b-is:

-L_c-is –CH₂-;

R¹ is selected from the group consisting of: -O-, - (R²) N-, -P (=O) (R²) -and -S-;

X is -L¹-CH₂-C (O) -;

L¹ is - (C (R^3a) (R^3b) ) _m-, wherein 0 or at least 1 methylene unit of L¹ is independently replaced by -C (O) -, -C (=S) -, -C (=NR^4b) -or -C (=N₂) -;

wherein each R², each R^3a, each R^3b and each R^4b are each independently hydrogen, protium, deuterium, tritium, halogen, -NO₂, -CN, -OR, -SR, -N (R^a) (R^b) , -C (O) R, -CO₂R, -C (O) C (O) R, -C (O) CH₂C (O) R, -S (O) R, -S (O) ₂R, -C (O) N (R^a) (R^b) , -SO₂N (R^a) (R^b) , -OC (O) R, -N (R) SO₂R, or a C_1-6 aliphatic group optionally substituted with R;

wherein each R, each R^a and each R^b are each independently hydrogen, protium, deuterium, tritium, halogen, -NO₂, -CN, -OH, -SH, -NH₂, -C (O) H, -CO₂H, -C (O) C (O) H, -C (O) CH₂C (O) H, -S (O) H, -S (O) ₂H, -C (O) NH₂, -SO₂NH₂, -OC (O) H, -N (H) SO₂H or a C_1-6 aliphatic group;

m is selected from the group consisting of integers ≥ 0;

when R¹ is -O-or -HN-, at least 1 methylene unit of L¹ is independently replaced by -C (O) -, -C (=S) -, -C (=NR^4b) -or -C (=N₂) -, or each R^3a and each R^3b are not both hydrogen;

for use in treating cancer in a subject.
A composition comprising an antibody-drug conjugate of formula (I’) :

or a tautomer, a mesomer, a racemate, an enantiomer or a diastereoisomer thereof, or a pharmaceutically acceptable salt thereof, or a mixture of any thereof,

wherein:

Ab is an anti-B7-H3 antibody or a fragment thereof, wherein the anti-B7-H3 antibody or a fragment thereof comprises heavy chain complementarity determining regions (HCDRs) 1-3 and light chain complementarity determining regions (LCDRs) 1-3, defined:

(a) according to the Kabat numbering scheme, wherein:

HCDR1 comprises or consists of an amino acid sequence according to SEQ ID NO: 1, HCDR2 comprises or consists of an amino acid sequence according to SEQ ID NO: 2, HCDR3 comprises or consists of an amino acid sequence according to SEQ ID NO: 3, LCDR1 comprises or consists of an amino acid sequence according to SEQ ID NO: 4, LCDR2 comprises or consists of an amino acid sequence according to SEQ ID NO: 5, and LCDR3 comprises or consists of an amino acid sequence according to SEQ ID NO: 6; and/or

(b) according to the IMGT numbering scheme, wherein:

HCDR1 comprises or consists of an amino acid sequence according to SEQ ID NO: 7, HCDR2 comprises or consists of an amino acid sequence according to SEQ ID NO: 8, HCDR3 comprises or consists of an amino acid sequence according to SEQ ID NO: 9, LCDR1 comprises or consists of an amino acid sequence according to SEQ ID NO: 10, LCDR2 comprises or consists of an amino acid sequence according to SEQ ID NO: 11, and LCDR3 comprises or consists of an amino acid sequence according to SEQ ID NO: 12;

n’ is an average connection number, and is an integer or a decimal from 1 to 10;

L is -L_a-L_b-L_c-;

L_a-is

-L_b-is:

-L_c-is –CH₂-;

R¹ is selected from the group consisting of: -O-, - (R²) N-, -P (=O) (R²) -and -S-;

X is -L¹-CH₂-C (O) -;

L¹ is - (C (R^3a) (R^3b) ) _m-, wherein 0 or at least 1 methylene unit of L¹ is independently replaced by -C (O) -, -C (=S) -, -C (=NR^4b) -or -C (=N₂) -;

wherein each R², each R^3a, each R^3b and each R^4b are each independently hydrogen, protium, deuterium, tritium, halogen, -NO₂, -CN, -OR, -SR, -N (R^a) (R^b) , -C (O) R, -CO₂R, -C (O) C (O) R, -C (O) CH₂C (O) R, -S (O) R, -S (O) ₂R, -C (O) N (R^a) (R^b) , -SO₂N (R^a) (R^b) , -OC (O) R, -N (R) SO₂R, or a C_1-6 aliphatic group optionally substituted with R;

wherein each R, each R^a and each R^b are each independently hydrogen, protium, deuterium, tritium, halogen, -NO₂, -CN, -OH, -SH, -NH₂, -C (O) H, -CO₂H, -C (O) C (O) H, -C (O) CH₂C (O) H, -S (O) H, -S (O) ₂H, -C (O) NH₂, -SO₂NH₂, -OC (O) H, -N (H) SO₂H or a C_1-6 aliphatic group;

m is selected from the group consisting of integers ≥ 0;

when R¹ is -O-or -HN-, at least 1 methylene unit of L¹ is independently replaced by -C (O) -, -C (=S) -, -C (=NR^4b) -or -C (=N₂) -, or each R^3a and each R^3b are not both hydrogen;

for use in treating cancer in a subject.
The antibody-drug conjugate or tautomer, mesomer, racemate, enantiomer or diastereoisomer thereof, or pharmaceutically acceptable salt thereof for use according to claim 1, or the composition for use according to claim 2, wherein X is -L¹-CH₂-C (O) -, L¹ is - (C (R^3a) (R^3b) ) _m-, m is not 0, and each R^3a and each R^3b are not both hydrogen.
The antibody-drug conjugate or tautomer, mesomer, racemate, enantiomer or diastereoisomer thereof, or pharmaceutically acceptable salt thereof for use according to claim 1 or claim 3, or the composition for use according to claim 2 or claim 3, wherein m is 1, and L¹ is -C (R^3a) (R^3b) -.
The antibody-drug conjugate or tautomer, mesomer, racemate, enantiomer or diastereoisomer thereof, or pharmaceutically acceptable salt thereof for use according to any one of claims 1 or 3-4, or the composition for use according to any one of claims 2-4, wherein R^3a is a C_1-6 aliphatic group, and R^3b is hydrogen.
The antibody-drug conjugate or tautomer, mesomer, racemate, enantiomer or diastereoisomer thereof, or pharmaceutically acceptable salt thereof for use according to any one of claims 1 or 3-5, or the composition for use according to any one of claims 2-5, wherein R^3a is methyl, and R^3b is hydrogen.
The antibody-drug conjugate or tautomer, mesomer, racemate, enantiomer or diastereoisomer thereof, or pharmaceutically acceptable salt thereof for use according to any one of claims 1 or 3-6, or the composition for use according to any one of claims 2-6, wherein R¹ is -O-.
The antibody-drug conjugate or tautomer, mesomer, racemate, enantiomer or diastereoisomer thereof, or pharmaceutically acceptable salt thereof for use according to any one of claims 1 or 3-7, or the composition for use according to any one of claims 2-7, wherein the antibody-drug conjugate comprises the following structure:
The antibody-drug conjugate or tautomer, mesomer, racemate, enantiomer or diastereoisomer thereof, or pharmaceutically acceptable salt thereof for use according to any one of claims 1 or 3-8, wherein the antibody-drug conjugate is an antibody-drug conjugate of formula (II) :

or a pharmaceutically acceptable salt thereof.
The composition for use according to any one of claims 2-8, wherein the antibody-drug conjugate is an antibody-drug conjugate of formula (II’) :

or a pharmaceutically acceptable salt thereof.
The antibody-drug conjugate or tautomer, mesomer, racemate, enantiomer or diastereoisomer thereof, or pharmaceutically acceptable salt thereof for use according to any one of claims 1 or 3-9, wherein n is 3 to 9.
The antibody-drug conjugate or tautomer, mesomer, racemate, enantiomer or diastereoisomer thereof, or pharmaceutically acceptable salt thereof for use according to claim 11, wherein n is 4 to 8.
The antibody-drug conjugate or tautomer, mesomer, racemate, enantiomer or diastereoisomer thereof, or pharmaceutically acceptable salt thereof for use according to claim 12, wherein n is 5 to 7.
The antibody-drug conjugate or tautomer, mesomer, racemate, enantiomer or diastereoisomer thereof, or pharmaceutically acceptable salt thereof for use according to claim 13, wherein n is 6.
The composition for use according to any one of claims 2-8 or 10, wherein n’ is an integer or decimal from 3 to 9.
The composition for use according to claim 15, wherein n’ is an integer or decimal from 4 to 8.
The composition for use according to claim 16, wherein n’ is an integer or decimal from 5 to 7.
The composition for use according to claim 17, wherein n’ is about 6.
The antibody-drug conjugate or tautomer, mesomer, racemate, enantiomer or diastereoisomer thereof, or pharmaceutically acceptable salt thereof for use according to any one of claims 1, 3-9, or 11-14, or the composition for use according to any one of claims 2-8, 10 or 15-18, wherein the anti-B7-H3 antibody or a fragment thereof comprises a heavy chain variable region comprising or consisting of an amino acid sequence according to SEQ ID NO: 13, or a variant having at least 80%sequence identity thereto, and a light chain variable region comprising or consisting of an amino acid sequence according to SEQ ID NO 14, or a variant having at least 80%sequence identity thereto.
The antibody-drug conjugate or tautomer, mesomer, racemate, enantiomer or diastereoisomer thereof, or pharmaceutically acceptable salt thereof for use according to any one of claims 1, 3-9, 11-14 or 19, or the composition for use according to any one of claims 2-8, 10 or 15-19, wherein the anti-B7-H3 antibody comprises a heavy chain comprising or consisting of an amino acid sequence according to SEQ ID NO: 15, or a variant having at least 80%sequence identity thereto, and a light chain comprising or consisting of an amino acid sequence according to SEQ ID NO 16, or a variant having at least 80%sequence identity thereto.
The antibody-drug conjugate or tautomer, mesomer, racemate, enantiomer or diastereoisomer thereof, or pharmaceutically acceptable salt thereof for use according to any one of claims 1, 3-9, 11-14 or 19-20, or the composition for use according to any one of claims 2-8, 10 or 15-20, wherein the cancer is selected from the group consisting of non-small cell lung cancer (NSCLC) , small cell lung cancer (SCLC) , castration-resistant prostate cancer (CRPC) , esophageal squamous cell carcinoma (ESCC) , melanoma, hepatocellular carcinoma (HCC) , cervical cancer, and solid tumour.
The antibody-drug conjugate or tautomer, mesomer, racemate, enantiomer or diastereoisomer thereof, or pharmaceutically acceptable salt thereof, or the composition for use according to claim 21, wherein the cancer is selected from the group consisting of NSCLC, SCLC, CRPC, ESCC and melanoma.
The antibody-drug conjugate or tautomer, mesomer, racemate, enantiomer or diastereoisomer thereof, or pharmaceutically acceptable salt thereof, or the composition for use according to claim 22, wherein the cancer is NSCLC.
The antibody-drug conjugate or tautomer, mesomer, racemate, enantiomer or diastereoisomer thereof, or pharmaceutically acceptable salt thereof, or the composition for use according to claim 22, wherein the cancer is SCLC.
The antibody-drug conjugate or tautomer, mesomer, racemate, enantiomer or diastereoisomer thereof, or pharmaceutically acceptable salt thereof, or the composition for use according to claim 22, wherein the cancer is CRPC.
The antibody-drug conjugate or tautomer, mesomer, racemate, enantiomer or diastereoisomer thereof, or pharmaceutically acceptable salt thereof, or the composition for use according to claim 22, wherein the cancer is ESCC.
The antibody-drug conjugate or tautomer, mesomer, racemate, enantiomer or diastereoisomer thereof, or pharmaceutically acceptable salt thereof, or the composition for use according to claim 22, wherein the cancer is melanoma.
The antibody-drug conjugate or tautomer, mesomer, racemate, enantiomer or diastereoisomer thereof, or pharmaceutically acceptable salt thereof for use according to any one of claims 1, 3-9, 11-14 or 19-27, or the composition for use according to any one of claims 2-8, 10 or 15-27, wherein the cancer is advanced, unresectable, and/or metastatic.
The antibody-drug conjugate or tautomer, mesomer, racemate, enantiomer or diastereoisomer thereof, or pharmaceutically acceptable salt thereof for use according to any one of claims 1, 3-9, 11-14 or 19-28, or the composition for use according to any one of claims 2-8, 10 or 15-28, wherein the cancer has progressed on or after standard systemic treatment.
The antibody-drug conjugate or tautomer, mesomer, racemate, enantiomer or diastereoisomer thereof, or pharmaceutically acceptable salt thereof for use according to any one of claims 1, 3-9, 11-14 or 19-29, or the composition for use according to any one of claims 2-8, 10 or 15-29, wherein the antibody-drug conjugate is administered to the subject at a dose in the range of about 3 mg/kg to about 15 mg/kg.
The antibody-drug conjugate or tautomer, mesomer, racemate, enantiomer or diastereoisomer thereof, or pharmaceutically acceptable salt thereof for use according to any one of claims 1, 3-9, 11-14 or 19-30, or the composition for use according to any one of claims 2-8, 10 or 15-30, wherein the antibody-drug conjugate is administered to the subject at a dose in the range of about 6 mg/kg to about 12 mg/kg.
The antibody-drug conjugate or tautomer, mesomer, racemate, enantiomer or diastereoisomer thereof, or pharmaceutically acceptable salt thereof for use according to any one of claims 1, 3-9, 11-14 or 19-31, or the composition for use according to any one of claims 2-8, 10 or 15-31, wherein the antibody-drug conjugate is administered to the subject at a dose in the range of about 6 mg/kg to about 9 mg/kg.
The antibody-drug conjugate or tautomer, mesomer, racemate, enantiomer or diastereoisomer thereof, or pharmaceutically acceptable salt thereof for use according to any one of claims 1, 3-9, 11-14 or 19-32, or the composition for use according to any one of claims 2-8, 10 or 15-32, wherein the antibody-drug conjugate is administered to the subject at a dose of about 9 mg/kg.
The antibody-drug conjugate or tautomer, mesomer, racemate, enantiomer or diastereoisomer thereof, or pharmaceutically acceptable salt thereof for use according to any one of claims 1, 3-9, 11-14 or 19-32, or the composition for use according to any one of 1, 3-9, 11-14 or 19-32, wherein the antibody-drug conjugate is administered to the subject at a dose of about 6 mg/kg.
The antibody-drug conjugate or tautomer, mesomer, racemate, enantiomer or diastereoisomer thereof, or pharmaceutically acceptable salt thereof for use according to any one of claims 1, 3-9, 11-14 or 19-34, or the composition for use according to any one of 1, 3-9, 11-14 or 19-34, wherein the antibody-drug conjugate or composition is administered to the subject once every three weeks (Q3W) .
The antibody-drug conjugate or tautomer, mesomer, racemate, enantiomer or diastereoisomer thereof, or pharmaceutically acceptable salt thereof for use according to any one of claims 1, 3-9, 11-14 or 19-35, or the composition for use according to any one of 1, 3-9, 11-14 or 19-35, wherein the antibody-drug conjugate or composition is administered to the subject in the form of a pharmaceutical composition, optionally wherein the pharmaceutical composition comprises a sterile aqueous buffer.
The antibody-drug conjugate or tautomer, mesomer, racemate, enantiomer or diastereoisomer thereof, or pharmaceutically acceptable salt thereof for use according to any one of claims 1, 3-9, 11-14 or 19-36, or the composition for use according to any one of 1, 3- 9, 11-14 or 19-36, wherein the antibody-drug conjugate is administered to the subject intravenously.
The antibody-drug conjugate or tautomer, mesomer, racemate, enantiomer or diastereoisomer thereof, or pharmaceutically acceptable salt thereof for use according to any one of claims 1, 3-9, 11-14 or 19-37, or the composition for use according to any one of 1, 3-9, 11-14 or 19-37, wherein the subject is human.
The antibody-drug conjugate or tautomer, mesomer, racemate, enantiomer or diastereoisomer thereof, or pharmaceutically acceptable salt thereof for use according to any one of claims 1, 3-9, 11-14 or 19-38, or the composition for use according to any one of 1, 3-9, 11-14 or 19-38, wherein the subject is aged ≥18 years.
The antibody-drug conjugate or tautomer, mesomer, racemate, enantiomer or diastereoisomer thereof, or pharmaceutically acceptable salt thereof for use according to any one of claims 1, 3-9, 11-14 or 19-39, or the composition for use according to any one of 1, 3-9, 11-14 or 19-39, wherein the subject has received prior treatment for cancer, optionally wherein the prior treatment is chemotherapy, immunotherapy and/or targeted therapy.
A method of treating cancer in a subject comprising administering to the subject an antibody-drug conjugate of formula (I) :

or a tautomer, a mesomer, a racemate, an enantiomer or a diastereoisomer thereof, or a pharmaceutically acceptable salt thereof,

wherein:

Ab is an anti-B7-H3 antibody or a fragment thereof, wherein the anti-B7-H3 antibody or a fragment thereof comprises heavy chain complementarity determining regions (HCDRs) 1-3 and light chain complementarity determining regions (LCDRs) 1-3, defined:

(a) according to the Kabat numbering scheme, wherein:

HCDR1 comprises or consists of an amino acid sequence according to SEQ ID NO: 1, HCDR2 comprises or consists of an amino acid sequence according to SEQ ID NO: 2, HCDR3 comprises or consists of an amino acid sequence according to SEQ ID NO: 3, LCDR1 comprises or consists of an amino acid sequence according to SEQ ID NO: 4, LCDR2 comprises or consists of an amino acid sequence according to SEQ ID NO: 5, and LCDR3 comprises or consists of an amino acid sequence according to SEQ ID NO: 6; and/or

(b) according to the IMGT numbering scheme, wherein:

HCDR1 comprises or consists of an amino acid sequence according to SEQ ID NO: 7, HCDR2 comprises or consists of an amino acid sequence according to SEQ ID NO: 8, HCDR3 comprises or consists of an amino acid sequence according to SEQ ID NO: 9, LCDR1 comprises or consists of an amino acid sequence according to SEQ ID NO: 10, LCDR2 comprises or consists of an amino acid sequence according to SEQ ID NO: 11, and LCDR3 comprises or consists of an amino acid sequence according to SEQ ID NO: 12;

n is a connection number, and is an integer from 1 to 10;

L is -L_a-L_b-L_c-;

L_a-is

-L_b-is:

-L_c-is –CH₂-;

R¹ is selected from the group consisting of: -O-, - (R²) N-, -P (=O) (R²) -and -S-;

X is -L¹-CH₂-C (O) -;

L¹ is - (C (R^3a) (R^3b) ) _m-, wherein 0 or at least 1 methylene unit of L¹ is independently replaced by -C (O) -, -C (=S) -, -C (=NR^4b) -or -C (=N₂) -;

wherein each R², each R^3a, each R^3b and each R^4b are each independently hydrogen, protium, deuterium, tritium, halogen, -NO₂, -CN, -OR, -SR, -N (R^a) (R^b) , -C (O) R, -CO₂R, -C (O) C (O) R, -C (O) CH₂C (O) R, -S (O) R, -S (O) ₂R, -C (O) N (R^a) (R^b) , -SO₂N (R^a) (R^b) , -OC (O) R, -N (R) SO₂R, or a C_1-6 aliphatic group optionally substituted with R;

wherein each R, each R^a and each R^b are each independently hydrogen, protium, deuterium, tritium, halogen, -NO₂, -CN, -OH, -SH, -NH₂, -C (O) H, -CO₂H, -C (O) C (O) H, -C (O) CH₂C (O) H, -S (O) H, -S (O) ₂H, -C (O) NH₂, -SO₂NH₂, -OC (O) H, -N (H) SO₂H or a C_1-6 aliphatic group;

m is selected from the group consisting of integers ≥ 0;

when R¹ is -O-or -HN-, at least 1 methylene unit of L¹ is independently replaced by -C (O) -, -C (=S) -, -C (=NR^4b) -or -C (=N₂) -, or each R^3a and each R^3b are not both hydrogen.
A method of treating cancer in a subject comprising administering to the subject a composition comprising an antibody-drug conjugate of formula (I’) :

or a tautomer, a mesomer, a racemate, an enantiomer or a diastereoisomer thereof, or a pharmaceutically acceptable salt thereof, or a mixture of any thereof,

wherein:

Ab is an anti-B7-H3 antibody or a fragment thereof, wherein the anti-B7-H3 antibody or a fragment thereof comprises heavy chain complementarity determining regions (HCDRs) 1-3 and light chain complementarity determining regions (LCDRs) 1-3, defined:

(a) according to the Kabat numbering scheme, wherein:

HCDR1 comprises or consists of an amino acid sequence according to SEQ ID NO: 1, HCDR2 comprises or consists of an amino acid sequence according to SEQ ID NO: 2, HCDR3 comprises or consists of an amino acid sequence according to SEQ ID NO: 3, LCDR1 comprises or consists of an amino acid sequence according to SEQ ID NO: 4, LCDR2 comprises or consists of an amino acid sequence according to SEQ ID NO: 5, and LCDR3 comprises or consists of an amino acid sequence according to SEQ ID NO: 6; and/or

(b) according to the IMGT numbering scheme, wherein:

HCDR1 comprises or consists of an amino acid sequence according to SEQ ID NO: 7, HCDR2 comprises or consists of an amino acid sequence according to SEQ ID NO: 8, HCDR3 comprises or consists of an amino acid sequence according to SEQ ID NO: 9, LCDR1 comprises or consists of an amino acid sequence according to SEQ ID NO: 10, LCDR2 comprises or consists of an amino acid sequence according to SEQ ID NO: 11, and LCDR3 comprises or consists of an amino acid sequence according to SEQ ID NO: 12;

n’ is an average connection number, and is an integer or a decimal from 1 to 10;

L is -L_a-L_b-L_c-;

L_a-is

-L_b-is:

-L_c-is –CH₂-;

R¹ is selected from the group consisting of: -O-, - (R²) N-, -P (=O) (R²) -and -S-;

X is -L¹-CH₂-C (O) -;

L¹ is - (C (R^3a) (R^3b) ) _m-, wherein 0 or at least 1 methylene unit of L¹ is independently replaced by -C (O) -, -C (=S) -, -C (=NR^4b) -or -C (=N₂) -;

wherein each R², each R^3a, each R^3b and each R^4b are each independently hydrogen, protium, deuterium, tritium, halogen, -NO₂, -CN, -OR, -SR, -N (R^a) (R^b) , -C (O) R, -CO₂R, -C (O) C (O) R, -C (O) CH₂C (O) R, -S (O) R, -S (O) ₂R, -C (O) N (R^a) (R^b) , -SO₂N (R^a) (R^b) , -OC (O) R, -N (R) SO₂R, or a C_1-6 aliphatic group optionally substituted with R;

wherein each R, each R^a and each R^b are each independently hydrogen, protium, deuterium, tritium, halogen, -NO₂, -CN, -OH, -SH, -NH₂, -C (O) H, -CO₂H, -C (O) C (O) H, -C (O) CH₂C (O) H, -S (O) H, -S (O) ₂H, -C (O) NH₂, -SO₂NH₂, -OC (O) H, -N (H) SO₂H or a C_1-6 aliphatic group;

m is selected from the group consisting of integers ≥ 0;

when R¹ is -O-or -HN-, at least 1 methylene unit of L¹ is independently replaced by -C (O) -, -C (=S) -, -C (=NR^4b) -or -C (=N₂) -, or each R^3a and each R^3b are not both hydrogen.
Use of an antibody-drug conjugate of formula (I) :

or a tautomer, a mesomer, a racemate, an enantiomer or a diastereoisomer thereof, or a pharmaceutically acceptable salt thereof,

wherein:

Ab is an anti-B7-H3 antibody or a fragment thereof, wherein the anti-B7-H3 antibody or a fragment thereof comprises heavy chain complementarity determining regions (HCDRs) 1-3 and light chain complementarity determining regions (LCDRs) 1-3, defined:

(a) according to the Kabat numbering scheme, wherein:

HCDR1 comprises or consists of an amino acid sequence according to SEQ ID NO: 1, HCDR2 comprises or consists of an amino acid sequence according to SEQ ID NO: 2, HCDR3 comprises or consists of an amino acid sequence according to SEQ ID NO: 3, LCDR1 comprises or consists of an amino acid sequence according to SEQ ID NO: 4, LCDR2 comprises or consists of an amino acid sequence according to SEQ ID NO: 5, and LCDR3 comprises or consists of an amino acid sequence according to SEQ ID NO: 6; and/or

(b) according to the IMGT numbering scheme, wherein:

HCDR1 comprises or consists of an amino acid sequence according to SEQ ID NO: 7, HCDR2 comprises or consists of an amino acid sequence according to SEQ ID NO: 8, HCDR3 comprises or consists of an amino acid sequence according to SEQ ID NO: 9, LCDR1 comprises or consists of an amino acid sequence according to SEQ ID NO: 10, LCDR2 comprises or consists of an amino acid sequence according to SEQ ID NO: 11, and LCDR3 comprises or consists of an amino acid sequence according to SEQ ID NO: 12;

n is a connection number, and is an integer from 1 to 10;

L is -L_a-L_b-L_c-;

L_a-is

-L_b-is:

-L_c-is –CH₂-;

R¹ is selected from the group consisting of: -O-, - (R²) N-, -P (=O) (R²) -and -S-;

X is -L¹-CH₂-C (O) -;

L¹ is - (C (R^3a) (R^3b) ) _m-, wherein 0 or at least 1 methylene unit of L¹ is independently replaced by -C (O) -, -C (=S) -, -C (=NR^4b) -or -C (=N₂) -;

wherein each R², each R^3a, each R^3b and each R^4b are each independently hydrogen, protium, deuterium, tritium, halogen, -NO₂, -CN, -OR, -SR, -N (R^a) (R^b) , -C (O) R, -CO₂R, -C (O) C (O) R, -C (O) CH₂C (O) R, -S (O) R, -S (O) ₂R, -C (O) N (R^a) (R^b) , -SO₂N (R^a) (R^b) , -OC (O) R, -N (R) SO₂R, or a C_1-6 aliphatic group optionally substituted with R;

wherein each R, each R^a and each R^b are each independently hydrogen, protium, deuterium, tritium, halogen, -NO₂, -CN, -OH, -SH, -NH₂, -C (O) H, -CO₂H, -C (O) C (O) H, -C (O) CH₂C (O) H, -S (O) H, -S (O) ₂H, -C (O) NH₂, -SO₂NH₂, -OC (O) H, -N (H) SO₂H or a C_1-6 aliphatic group;

m is selected from the group consisting of integers ≥ 0;

when R¹ is -O-or -HN-, at least 1 methylene unit of L¹ is independently replaced by -C (O) -, -C (=S) -, -C (=NR^4b) -or -C (=N₂) -, or each R^3a and each R^3b are not both hydrogen;

in the manufacture of a medicament for the treatment of cancer.
Use of a composition comprising an antibody-drug conjugate of formula (I’) :

or a tautomer, a mesomer, a racemate, an enantiomer or a diastereoisomer thereof, or a pharmaceutically acceptable salt thereof, or a mixture of any thereof,

wherein:

Ab is an anti-B7-H3 antibody or a fragment thereof, wherein the anti-B7-H3 antibody or a fragment thereof comprises heavy chain complementarity determining regions (HCDRs) 1-3 and light chain complementarity determining regions (LCDRs) 1-3, defined:

(a) according to the Kabat numbering scheme, wherein:

HCDR1 comprises or consists of an amino acid sequence according to SEQ ID NO: 1, HCDR2 comprises or consists of an amino acid sequence according to SEQ ID NO: 2, HCDR3 comprises or consists of an amino acid sequence according to SEQ ID NO: 3, LCDR1 comprises or consists of an amino acid sequence according to SEQ ID NO: 4, LCDR2 comprises or consists of an amino acid sequence according to SEQ ID NO: 5, and LCDR3 comprises or consists of an amino acid sequence according to SEQ ID NO: 6; and/or

(b) according to the IMGT numbering scheme, wherein:

HCDR1 comprises or consists of an amino acid sequence according to SEQ ID NO: 7, HCDR2 comprises or consists of an amino acid sequence according to SEQ ID NO: 8, HCDR3 comprises or consists of an amino acid sequence according to SEQ ID NO: 9, LCDR1 comprises or consists of an amino acid sequence according to SEQ ID NO: 10, LCDR2 comprises or consists of an amino acid sequence according to SEQ ID NO: 11, and LCDR3 comprises or consists of an amino acid sequence according to SEQ ID NO: 12;

n’ is an average connection number, and is an integer or a decimal from 1 to 10;

L is -L_a-L_b-L_c-;

L_a-is

-L_b-is:

-L_c-is –CH₂-;

R¹ is selected from the group consisting of: -O-, - (R²) N-, -P (=O) (R²) -and -S-;

X is -L¹-CH₂-C (O) -;

L¹ is - (C (R^3a) (R^3b) ) _m-, wherein 0 or at least 1 methylene unit of L¹ is independently replaced by -C (O) -, -C (=S) -, -C (=NR^4b) -or -C (=N₂) -;

wherein each R², each R^3a, each R^3b and each R^4b are each independently hydrogen, protium, deuterium, tritium, halogen, -NO₂, -CN, -OR, -SR, -N (R^a) (R^b) , -C (O) R, -CO₂R, -C (O) C (O) R, -C (O) CH₂C (O) R, -S (O) R, -S (O) ₂R, -C (O) N (R^a) (R^b) , -SO₂N (R^a) (R^b) , -OC (O) R, -N (R) SO₂R, or a C_1-6 aliphatic group optionally substituted with R;

wherein each R, each R^a and each R^b are each independently hydrogen, protium, deuterium, tritium, halogen, -NO₂, -CN, -OH, -SH, -NH₂, -C (O) H, -CO₂H, -C (O) C (O) H, -C (O) CH₂C (O) H, -S (O) H, -S (O) ₂H, -C (O) NH₂, -SO₂NH₂, -OC (O) H, -N (H) SO₂H or a C_1-6 aliphatic group;

m is selected from the group consisting of integers ≥ 0;

when R¹ is -O-or -HN-, at least 1 methylene unit of L¹ is independently replaced by -C (O) -, -C (=S) -, -C (=NR^4b) -or -C (=N₂) -, or each R^3a and each R^3b are not both hydrogen;

in the manufacture of a medicament for the treatment of cancer.