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CN114621186A - Heterocyclic compounds as modulators of RAS signaling pathway - Google Patents

Heterocyclic compounds as modulators of RAS signaling pathway Download PDF

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Publication number
CN114621186A
CN114621186A CN202210510428.5A CN202210510428A CN114621186A CN 114621186 A CN114621186 A CN 114621186A CN 202210510428 A CN202210510428 A CN 202210510428A CN 114621186 A CN114621186 A CN 114621186A
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amino
group
ethyl
phenyl
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唐国志
黄孟炜
刘永福
尹军
陈军利
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Shanghai Weishen Pharmaceutical Co ltd
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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D401/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom
    • C07D401/02—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings
    • C07D401/04—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings directly linked by a ring-member-to-ring-member bond
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D237/00—Heterocyclic compounds containing 1,2-diazine or hydrogenated 1,2-diazine rings
    • C07D237/26—Heterocyclic compounds containing 1,2-diazine or hydrogenated 1,2-diazine rings condensed with carbocyclic rings or ring systems
    • C07D237/30—Phthalazines
    • C07D237/34—Phthalazines with nitrogen atoms directly attached to carbon atoms of the nitrogen-containing ring, e.g. hydrazine radicals
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D401/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom
    • C07D401/02—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings
    • C07D401/12—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings linked by a chain containing hetero atoms as chain links
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D403/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00
    • C07D403/02—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00 containing two hetero rings
    • C07D403/04—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00 containing two hetero rings directly linked by a ring-member-to-ring-member bond
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D403/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00
    • C07D403/02—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00 containing two hetero rings
    • C07D403/12—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00 containing two hetero rings linked by a chain containing hetero atoms as chain links
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D405/00—Heterocyclic compounds containing both one or more hetero rings having oxygen atoms as the only ring hetero atoms, and one or more rings having nitrogen as the only ring hetero atom
    • C07D405/02—Heterocyclic compounds containing both one or more hetero rings having oxygen atoms as the only ring hetero atoms, and one or more rings having nitrogen as the only ring hetero atom containing two hetero rings
    • C07D405/12—Heterocyclic compounds containing both one or more hetero rings having oxygen atoms as the only ring hetero atoms, and one or more rings having nitrogen as the only ring hetero atom containing two hetero rings linked by a chain containing hetero atoms as chain links
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D471/00—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, at least one ring being a six-membered ring with one nitrogen atom, not provided for by groups C07D451/00 - C07D463/00
    • C07D471/02—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, at least one ring being a six-membered ring with one nitrogen atom, not provided for by groups C07D451/00 - C07D463/00 in which the condensed system contains two hetero rings
    • C07D471/04—Ortho-condensed systems
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D493/00—Heterocyclic compounds containing oxygen atoms as the only ring hetero atoms in the condensed system
    • C07D493/02—Heterocyclic compounds containing oxygen atoms as the only ring hetero atoms in the condensed system in which the condensed system contains two hetero rings
    • C07D493/04—Ortho-condensed systems

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Abstract

The invention provides a heterocyclic compound serving as an RAS signal channel regulator, and particularly provides a compound shown as a formula I, a preparation method thereof and application of the compound serving as the RAS signal channel regulator. The compounds may be used for the preparation of pharmaceutical compositions for the treatment or prevention of diseases related to SOS1 or modulated by SOS 1.

Description

Heterocyclic compounds as modulators of RAS signaling pathway
Technical Field
The invention belongs to the field of chemical medicine, and particularly relates to a heterocyclic compound serving as an RAS signal channel regulator, and a preparation method and application thereof.
Background
The RAS family includes KRAS, NRAS and HRAS. RAS proteins tightly regulate cell proliferation, differentiation, senescence, apoptosis, and the like in normal and abnormal cells (including cancer). RAS proteins are activated by upstream signals, including Receptor Tyrosine Kinases (RTKs), and transduce signals into several downstream signaling pathways, such as the mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase (ERK) pathway and the PI3K-AKT-mTOR signaling pathway. Over-activation of RAS signals is often observed in cancer due to mutations or alterations in the RAS gene or other genes in the RAS pathway. Strategies that inhibit the RAS and RAS signaling pathways are useful in the treatment of cancer or other diseases in which RAS is modulated.
RAS family proteins are small gtpases, which are present in cells in either a GDP-bound (inactive) or GTP-bound (active) state. The activity of RAS proteins is regulated by proteins known as Gtpase Activator Proteins (GAPs) or guanine nucleotide exchange factors (GEFs). RAS family proteins have weak intrinsic gtpase activity and slow nucleotide exchange rates. Binding of a GTPase Activating Protein (GAP) such as NF1 increases the GTPase activity of RAS family proteins. Binding of guanine nucleotide exchange factor (GEF) such as SOS1(SonofSevenless1) promotes release of GDP in RAS family proteins, thereby effecting GTP binding. RAS family proteins are active, continuously activating downstream signaling pathways when in the GTP bound state.
The SOS family includes SOS1 and SOS2, which have about 70% sequence identity. SOS1 appears to be much more active than SOS2 due to the rapid degradation of SOS 2. Mouse SOS2 knockouts are feasible, while SOS1 knockouts are embryonic. Published data indicate that SOS1 plays a significant role in mutant KRAS activation and oncogenic signaling in cancer. Consumption of SOS1 levels decreased the proliferation and survival rate of tumor cells carrying KRAS mutations, whereas no effect was observed in KRAS wild-type cell lines. The effect of the deletion of SOS1 could not be rescued by the introduction of SOS1, a catalytic site mutation, demonstrating the important role of SOS1GEF activity in KRAS mutant cancer cells. The SOS1 mutation was found in the Noonan syndrome and several cancers, including lung adenocarcinoma, embryonal rhabdomyosarcoma, testicular supporting cell tumor, and granulomatosis by skin Gtpase Activating Protein (GAP).
Thus, the compounds of the invention that block the interaction between SOS1 and Ras family members can prevent the recycling of KRAS to the active GTP-bound form, and thus can provide benefits for the treatment of a variety of cancers, particularly cancers associated with Ras family members.
Disclosure of Invention
The invention aims to provide a Ras signal channel regulator with a novel structure.
In a first aspect of the invention, there is provided a compound of formula I, or a pharmaceutically acceptable salt thereof,
Figure 320049DEST_PATH_IMAGE001
x is selected from N or CRa；
RaSelected from the group consisting of: hydrogen, halogen, C1-6Alkyl or C1-6An alkoxy group;
l is selected from the group consisting of: - (CH)2)m-or-O-, wherein m is 0 or 1;
R1selected from the group consisting of: c1-6Alkyl radical, C3-6Cycloalkyl, 4-to 8-membered saturated or unsaturated moietyUnsaturated heterocyclic radical, phenyl, 5-6 membered heteroaryl;
and said R is1Optionally substituted by one or more RbSubstituted by substituent, said RbSelected from the group consisting of: hydrogen, halogen, one or more Rb1Substituted C1-3Alkyl radical, C1-3Alkoxy, -C (= O) -Rb2Or one or more Rb3A substituted 5-6 membered heteroaryl, wherein:
Rb1selected from the group consisting of: hydrogen, halogen, amino or carboxyl;
Rb2selected from the group consisting of: hydrogen, C1-6Alkyl radical, C1-6Alkoxy radical, C3-6Cycloalkyl or- (CH)2)q-N(Rb1ARb1B)；
Rb3Selected from hydrogen, halogen or C1-3An alkyl group;
q is 0, 1,2 or 3;
Rb1A、Rb1Bis hydrogen or C1-3An alkyl group;
one or more methylene groups as a ring skeleton on each of the above-mentioned cycloalkyl groups and heterocyclic groups may each independently be replaced by a carbonyl group or S (= O)2Replacement;
each of said heterocyclyl or heteroaryl groups independently containing from 1 to 3 heteroatoms selected from N, O, S, the number of heteroatoms being from 1 to 3;
R2is optionally selected from C1-3Alkyl radical, C1-3Haloalkyl or cyclopropane;
R3can be the same or different and are each independently selected from halogen, amino, C1-3Alkyl or C1-3A haloalkyl group;
n is 1,2 or 3.
In another preferred embodiment, R is3Selected from the group consisting of: amino group, C1-3A haloalkyl group.
In another preferred embodiment, R is3Selected from the group consisting of: amino, trifluoromethyl.
In another preferred embodiment, the compound of formula I has the structure shown in formula II below:
Figure 33928DEST_PATH_IMAGE002
。
in another preferred embodiment, RaSelected from the group consisting of: hydrogen, halogen, C1-4Alkyl or C1-4An alkoxy group.
In another preferred embodiment, RaSelected from the group consisting of: hydrogen, halogen, methyl or methoxy.
In another preferred embodiment, L is selected from the group consisting of: - (CH)2)m-or-O-, wherein m is 0.
In another preferred embodiment, R2Is optionally selected from C1-3An alkyl group.
In another preferred embodiment, R1Selected from the group consisting of: c1-4Alkyl radical, C3-6Cycloalkyl, 5-8 membered saturated or partially unsaturated heterocyclyl; wherein, R is1Optionally substituted by one or more RbSubstituent, said RbSelected from the group consisting of: hydrogen, halogen, one or more Rb1Substituted C1-3Alkyl radical, C1-3Alkoxy, -C (= O) -Rb2Or one or more Rb3A substituted 5-6 membered heteroaryl, wherein:
Rb1selected from the group consisting of: hydrogen, halogen, or carboxyl;
Rb2selected from the group consisting of: hydrogen, C1-4Alkyl radical, C1-4Alkoxy radical, C3-6Cycloalkyl or- (CH)2)q-N(Rb1ARb1B)；
Rb3Selected from hydrogen, halogen or C1-3An alkyl group;
q is 0 or 1;
Rb1A、Rb1Bis hydrogen or C1-3An alkyl group.
In another preferred embodiment, R1Selected from the group consisting of: c1-4Alkyl, or a group selected from:
Figure 548086DEST_PATH_IMAGE003
；
said R1Can be optionally replaced by oneOr a plurality of RbSubstituted by substituent, said RbAs defined above.
In another preferred embodiment, the compound of formula I is selected from the group consisting of:
Figure 848355DEST_PATH_IMAGE004
Figure 459464DEST_PATH_IMAGE005
Figure 16348DEST_PATH_IMAGE006
in a second aspect of the present invention, there is provided a pharmaceutical composition comprising: a compound of formula I according to the first aspect of the invention or a pharmaceutically acceptable salt thereof, together with one or more pharmaceutically acceptable carriers, excipients, adjuvants and/or diluents.
In a third aspect of the present invention there is provided the use of a compound of formula I as described in the first aspect of the invention, or a pharmaceutically acceptable salt thereof, for the preparation of a pharmaceutical composition for the treatment or prevention of a disease associated with SOS1 or modulated by SOS 1.
It is to be understood that within the scope of the present invention, the above-described features of the present invention and those specifically described below (e.g., in the examples) may be combined with each other to form new or preferred embodiments. Not to be reiterated herein, but to the extent of space.
Detailed Description
Through long-term and intensive research, the inventor discovers a small molecule compound with a Ras signal channel regulator, and the compound has a novel structure and has equivalent or more excellent agonistic activity with the similar compound in the prior art. Based on the above findings, the inventors have completed the present invention.
Term(s) for
In the present invention, the halogen is F, Cl, Br or I.
In the present invention, unless otherwise specified, the terms used have the ordinary meanings well known to those skilled in the art.
In the present invention, the term "C1-C6Alkyl "means a straight or branched chain alkyl group having 1 to 6 carbon atoms, including, but not limited to, methyl, ethyl, propyl, isopropyl, butyl, isobutyl, sec-butyl, tert-butyl, pentyl, hexyl, and the like; ethyl, propyl, isopropyl, butyl, isobutyl, sec-butyl and tert-butyl are preferred.
In the present invention, the term "C1-C6Alkoxy "means a straight or branched chain alkoxy group having 1 to 6 carbon atoms, including, but not limited to, methoxy, ethoxy, propoxy, isopropoxy, butoxy, and the like.
In the present invention, the term "C3-C6Cycloalkyl "refers to cyclic alkyl groups having 3 to 6 carbon atoms in the ring and includes, without limitation, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, and the like. The term "C5-C6Cycloalkyl "has similar meaning.
In the present invention, the term "aromatic ring" or "aryl" has the same meaning, preferably "aryl" is "C6-C12Aryl "or" C6-C10Aryl ". The term "C6-C12Aryl "refers to an aromatic ring group having 6 to 12 carbon atoms, such as phenyl, naphthyl, and the like, which does not contain heteroatoms in the ring. The term "C6-C10Aryl "has a similar meaning.
In the present invention, the term "heteroaromatic ring" or "heteroaryl" has the same meaning and refers to a heteroaromatic group containing one to more heteroatoms. The hetero atoms referred to herein include oxygen, sulfur and nitrogen. Such as furyl, thienyl, pyridyl, pyrazolyl, pyrrolyl, N-alkylpyrrolyl, pyrimidinyl, pyrazinyl, imidazolyl, tetrazolyl, and the like. The heteroaryl ring may be fused to an aryl, heterocyclyl, or cycloalkyl ring, wherein the ring joined to the parent structure is a heteroaryl ring. Heteroaryl groups may be optionally substituted or unsubstituted.
In the present invention, the term "C3-6Cycloalkyl "refers to a saturated 3-6 membered cyclic group whose ring backbone structure includes only carbon atoms, such as cyclopentyl, cyclohexyl, and the like.
In the present invention, the term "4-8 membered heterocyclic group" means a saturated or unsaturated (non-aromatic ring including monocyclic, fused, spiro, bridged, etc. forms) 4-8 membered cyclic group containing 1 to 3 hetero atoms selected from oxygen, sulfur and nitrogen in the ring, for example, dioxolane, etc. The term "5-8 membered heterocyclyl" has a similar meaning.
In the present invention, the term "substituted" means that one or more hydrogen atoms on a specified group are replaced with a specified substituent. Particular substituents are those described correspondingly in the foregoing, or as appearing in the examples. Unless otherwise specified, a certain substituted group may have one substituent selected from a specific group at any substitutable site of the group, and the substituents may be the same or different at each position. A cyclic substituent, such as heterocycloalkyl, may be attached to another ring, such as cycloalkyl, to form a spiro bicyclic ring system, e.g., the two rings have a common carbon atom. It will be understood by those skilled in the art that the combinations of substituents contemplated by the present invention are those that are stable or chemically achievable. Such substituents are for example (but not limited to): c1-8Alkyl radical, C2-8Alkenyl radical, C2-8Alkynyl, C3-8Cycloalkyl, 3-to 12-membered heterocyclyl, aryl, heteroaryl, halogen, hydroxy, carboxy (-COOH), C1-8Aldehyde group, C2-10Acyl radical, C2-10Ester group, amino group, alkoxy group, C1-10Sulfonyl, and the like.
Compounds of formula I as modulators of RAS signaling pathway
The invention provides a compound shown as a formula I or a pharmaceutically acceptable salt thereof,
Figure 955485DEST_PATH_IMAGE007
x is selected from N or CRa；
RaSelected from the group consisting of: hydrogen, halogen, C1-6Alkyl or C1-6An alkoxy group;
l is selected from the group consisting of: - (CH)2)m-or-O-, wherein m is 0 or 1;
R1selected from the group consisting of: c1-6Alkyl radical, C3-6Cycloalkyl, 4-8 membered saturated or partially unsaturated heterocyclyl, phenyl, 5-6 membered heteroaryl;
and said R is1Optionally substituted by one or more RbSubstituted by substituent, said RbSelected from the group consisting of: hydrogen, halogen, one or more Rb1Substituted C1-3Alkyl radical, C1-3Alkoxy, -C (= O) -Rb2Or one or more Rb3A substituted 5-6 membered heteroaryl, wherein:
Rb1selected from the group consisting of: hydrogen, halogen, amino or carboxyl;
Rb2selected from the group consisting of: hydrogen, C1-6Alkyl radical, C1-6Alkoxy radical, C3-6Cycloalkyl or- (CH)2)q-N(Rb1ARb1B)；
Rb3Selected from hydrogen, halogen or C1-3An alkyl group;
q is 0, 1,2 or 3;
Rb1A、Rb1Bis hydrogen or C1-3An alkyl group;
one or more methylene groups as a ring skeleton on each of the above-mentioned cycloalkyl groups and heterocyclic groups may each independently be replaced by a carbonyl group or S (= O)2Replacement;
each of said heterocyclyl or heteroaryl groups independently contains from 1 to 3 heteroatoms selected from N, O, S, the number of heteroatoms being from 1 to 3;
R2is optionally selected from C1-3Alkyl radical, C1-3Haloalkyl or cyclopropane;
R3can be the same or different and are each independently selected from halogen, amino, C1-3Alkyl or C1-3A haloalkyl group;
n is 1,2 or 3.
Process for the preparation of compounds of formula I
The invention also provides a preparation method of the compound shown in the formula I, and specifically, the compound is prepared by adopting the following scheme 1:
scheme 1:
Figure 888806DEST_PATH_IMAGE008
in the process 1, an initial compound II-1 is subjected to esterification reaction and carbonyl insertion reaction to obtain a compound III; carrying out acid-base condensation reaction on the compound III to obtain a compound IV; carrying out deprotection-ring closure reaction on the compound IV to obtain a compound V; the compound V is substituted by chlorine to obtain a compound VI; carrying out Buchwald-Hartwig Cross Coupling Reaction or substitution Reaction on the compound VI to obtain a compound VII; carrying out dealkylation and substitution reaction on the compound VII to obtain a compound IX; and carrying out one-step or multi-step substitution reaction or Suzuki Coupling on the compound IX, and then carrying out reduction reaction to obtain the compound I, or directly obtaining the compound I from the compound IX through one-step or multi-step substitution reaction or Suzuki Coupling.
Q may be chlorine, bromine or OTf.
Pharmaceutical composition containing active ingredient
Since the compound of the present invention has an excellent RAS signaling pathway-regulating effect, the compound of the present invention and various crystalline forms, pharmaceutically acceptable inorganic or organic salts, hydrates or solvates thereof, and a pharmaceutical composition containing the compound of the present invention as a main active ingredient are useful for the prevention and/or treatment (stabilization, alleviation or cure) of diseases or disorders associated with abnormal interaction between SOS1 and RAS family members, particularly RAS family member-associated cancers.
The pharmaceutical compositions of the present invention comprise a safe and effective amount of a compound of the present invention in combination with a pharmaceutically acceptable excipient or carrier. Wherein "safe and effective amount" means: the amount of the compound is sufficient to significantly improve the condition without causing serious side effects. Typically, the pharmaceutical composition contains 0.01-99.99% by weight of the compound/agent of the present invention, more preferably 0.1-99.9% of the compound/agent of the present invention. Preferably, said "dose" is a capsule or tablet.
"pharmaceutically acceptable carrier" refers to: one or more compatible solid or liquid fillers or gel substances which are suitable for human use and must be of sufficient purity and sufficiently low toxicity. By "compatible" is meant herein that the components of the composition are capable of intermixing with and between the compounds of the present invention without significantly diminishing the pharmaceutical effectiveness of the compounds. Examples of pharmaceutically acceptable carrier moieties are cellulose and its derivatives (e.g., sodium carboxymethylcellulose, sodium ethylcellulose, cellulose acetate, etc.), gelatin, talc, solid lubricants (e.g., stearic acid, magnesium stearate), calcium sulfate, vegetable oils (e.g., soybean oil, sesame oil, peanut oil, olive oil, etc.), polyols (e.g., propylene glycol, glycerin, mannitol, sorbitol, etc.), emulsifiers (e.g., tween), wetting agents (e.g., sodium lauryl sulfate), colorants, flavors, stabilizers, antioxidants, preservatives, pyrogen-free water, and the like.
The mode of administration of the compounds or pharmaceutical compositions of the present invention is not particularly limited, and representative modes of administration include (but are not limited to): oral, parenteral (intravenous, intramuscular or subcutaneous).
Solid dosage forms for oral administration include capsules, tablets, pills, powders and granules. In these solid dosage forms, the active compound is mixed with at least one conventional inert excipient (or carrier), such as sodium citrate or dicalcium phosphate, or with the following ingredients: (a) fillers or extenders, for example, starch, lactose, sucrose, glucose, mannitol and silicic acid; (b) binders, for example, hydroxymethylcellulose, alginates, gelatin, polyvinylpyrrolidone, sucrose and acacia; (c) humectants, for example, glycerol; (d) disintegrating agents, for example, agar-agar, calcium carbonate, potato or tapioca starch, alginic acid, certain complex silicates, and sodium carbonate; (e) slow solvents, such as paraffin; (f) absorption accelerators, e.g., quaternary ammonium compounds; (g) wetting agents, such as cetyl alcohol and glycerol monostearate; (h) adsorbents, for example, kaolin; and (i) lubricants, for example, talc, calcium stearate, magnesium stearate, solid polyethylene glycols, sodium lauryl sulfate, or mixtures thereof. In capsules, tablets and pills, the dosage forms may also comprise buffering agents.
Solid dosage forms such as tablets, dragees, capsules, pills, and granules can be prepared using coatings and shells such as enteric coatings and other materials well known in the art. They may contain opacifying agents and the release of the active compound or compounds in such compositions may be delayed in release in a certain part of the digestive tract. Examples of embedding components which can be used are polymeric substances and wax-like substances. If desired, the active compound may also be in microencapsulated form with one or more of the above-mentioned excipients.
Liquid dosage forms for oral administration include pharmaceutically acceptable emulsions, solutions, suspensions, syrups or tinctures. In addition to the active compounds, the liquid dosage forms may contain inert diluents commonly employed in the art such as, for example, water or other solvents, solubilizing agents and emulsifiers such as ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, propylene glycol, 1, 3-butylene glycol, dimethylformamide and oils, in particular, cottonseed, groundnut, corn germ, olive, castor and sesame oils or mixtures of such materials and the like.
In addition to these inert diluents, the compositions can also contain adjuvants such as wetting agents, emulsifying and suspending agents, sweetening, flavoring, and perfuming agents.
Suspensions, in addition to the active compounds, may contain suspending agents as, for example, ethoxylated isostearyl alcohols, polyoxyethylene sorbitol and sorbitan esters, microcrystalline cellulose, aluminum methoxide and agar, or mixtures of these substances, and the like.
Compositions for parenteral injection may comprise physiologically acceptable sterile aqueous or anhydrous solutions, dispersions, suspensions or emulsions, and sterile powders for reconstitution into sterile injectable solutions or dispersions. Suitable aqueous and nonaqueous carriers, diluents, solvents or vehicles include water, ethanol, polyols and suitable mixtures thereof.
The compounds of the present invention may be administered alone or in combination with other pharmaceutically acceptable therapeutic agents.
When administered in combination, the pharmaceutical composition further comprises one or more (2, 3,4, or more) other pharmaceutically acceptable compounds. One or more (2, 3,4, or more) of the other pharmaceutically acceptable compounds can be used simultaneously, separately or sequentially with a compound of the invention for the prevention and/or treatment of Ras family member-associated cancer.
When using pharmaceutical compositions, a safe and effective amount of a compound of the present invention is administered to a mammal (e.g., a human) in need of treatment at a dosage that is pharmaceutically acceptable for effective administration. Of course, the particular dosage will depend upon such factors as the route of administration, the health of the patient, and the like, and is within the skill of the skilled practitioner.
The invention will be further illustrated with reference to the following specific examples. It should be understood that these examples are for illustrative purposes only and are not intended to limit the scope of the present invention. Experimental procedures without specific conditions noted in the following examples, generally according to conventional conditions, or according to conditions recommended by the manufacturer. Unless otherwise indicated, percentages and parts are by weight.
In the following examples, the structures of the compounds were determined by Nuclear Magnetic Resonance (NMR) or/and Mass Spectrometry (MS). NMR shift (. delta.) of 10-6 The units in (ppm) are given. NMR was measured by Bruker AVANCE-400 NMR spectrometer using deuterated dimethyl sulfoxide (DMSO-d)6) Deuterated chloroform (CDCl)3) Deuterated methanol (CD)3OD), internal standard Tetramethylsilane (TMS).
SHIMADZU LC system (chromatographic column: Xselect CSH ® CSHTMPrep-C18, 19 x 150mm, liquid handler LH-40, pump LC-20AP, detector SPD-20A, system controller CBM-20A, solvent system: acetonitrile and 0.05% aqueous trifluoroacetic acid).
LC/MS spectra of the compounds were obtained using LC/MS (Agilent Technologies 1200 Series). LC/MS conditions were as follows (run time 10 min):
acid conditions: a: 0.05% trifluoroacetic acid in water; b: 0.05% trifluoroacetic acid in acetonitrile;
alkaline conditions: a: 0.05% NH3•H2An aqueous solution of O; b:acetonitrile
Neutral conditions are as follows: a: 10 mM NH4An aqueous solution of OAC; b: acetonitrile
In the following examples, the intermediates and final compounds were purified using silica gel column chromatography, or Xselect CSH, unless otherwise specifiedTM Prep-C18(5μm，OBDTM19 x 150mm) chromatography column or using xbridge (tm) Prep Phenyl (5 μm, OBD)TM30 x 100mm) was purified by preparative HPLC on a reverse phase chromatography column.
Silica gel column chromatography generally uses 200-300 mesh silica gel of wakame as a carrier.
The CombiFlash rapid preparation instrument uses CombiFlash Rf200 (TELEDYNE ISCO).
The silica gel plate used in the Thin Layer Chromatography (TLC) detection product adopts a cigarette platform yellow sea HSGF254 or Qingdao GF254 silica gel plate, the specification of the silica gel plate used in the thin layer chromatography detection product is 0.15 mm-0.2 mm, and the specification of the thin layer chromatography separation and purification product is 0.4 mm-0.5 mm.
Known starting materials of the present invention can be synthesized by or according to methods known in the art, or can be purchased from companies such as ABCR GmbH & Co.KG, Acros Organics, Aldrich Chemical Company, Shao Yuan Chemical technology (Accela ChemBio Inc), Darri Chemicals, and the like.
Abbreviation: AcCl: acetyl chloride; BBr3Boron tribromide; BCl3Boron trichloride; BINAP: 1,1 '-binaphthyl-2, 2' -bis-diphenylphosphine; boc2O: tert-butyloxycarbonyl carbonate; conc. HCl: concentrated hydrochloric acid, Cs2CO3: cesium carbonate; DCM: dichloromethane; DCE: 1, 2-dichloroethane; the DIAD: diisopropyl azodicarboxylate; 1, 4-Dioxane: 1, 4-dioxane; DIEA: n, N-diisopropylethylamine; DMAP: 4-dimethylaminopyridine; DMF: dimethylformamide; DMSO, DMSO: dimethyl sulfoxide; EtOH: ethanol; HATU 2- (7-azobenzotriazol) -N, N, N, N-tetramethyluronium hexafluorophosphate; acetic acid; h2: hydrogen gas; i is2: iodine; k2CO3: potassium carbonate; KMnO4Potassium permanganate; k3PO4: potassium phosphate; LiAlH4: lithium aluminum hydride; LiOH: lithium hydroxide; MeOH: first of allAn alcohol; MTBE methyl tert-butyl ether; NaBH4Sodium borohydride; NaH: sodium hydride; NaHCO 23: sodium bicarbonate; pd (dppf) Cl2: [1,1' -bis (diphenylphosphino) ferrocene]Palladium dichloride; pd (OAc)2: palladium acetate; Pd/C: palladium on carbon; pd2(dba)3: tris (dibenzylideneacetone) dipalladium; pd (PPh)3)4: palladium tetrakistriphenylphosphine; POCl3: phosphorus oxychloride; PPh3: triphenylphosphine;t-BuOK: potassium tert-butoxide;t-BuONa: sodium tert-butoxide; TEA: triethylamine; TFA: trifluoroacetic acid; THF: tetrahydrofuran; TLC: thin layer chromatography; tf (f)2O is trifluoromethanesulfonic anhydride; ti (OEt)4Tetraethyl titanate; TsCl is p-toluenesulfonyl chloride; ts is p-toluenesulfonyl; XantPhos: 4, 5-bis (diphenylphosphino) -9, 9-dimethylxanthene; XPhos Pd g 3 (2-dicyclohexylphosphine-2, 4, 6-triisopropyl-1, 1-biphenyl) [2- (2-amido-1, 1-biphenyl ]]Palladium methane sulfonate.
Preparation example
Intermediate A1 (R) -2-methyl-4- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) -1-oxo-1, 2-dihydrophthalazin-6-yl trifluoromethanesulfonate
Figure 88843DEST_PATH_IMAGE009
Step 1 preparation of isopropyl 2-bromo-4-methoxybenzoate (intermediate A1-1)
2-bromo-4-methoxybenzoic acid (5.0 g, 21.70 mmol) was dissolved in DMF (40 ml). 2-iodopropane (4.4 g, 26.10 mmol) and K were added slowly with stirring at 0 deg.C2CO3 (4.5 g, 32.50 mmol) and after addition the reaction mixture was stirred at 60 ℃ for an additional 12 h, then quenched by addition of ice water and extracted by dilution with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous dry sodium sulfate, then filtered and concentrated to give the crude product, which was purified by silica gel column chromatography to give isopropyl 2-bromo-4-methoxybenzoate as a yellow oil (5.1 g, 86.4%). MS: 273.1&275.1 (M+H)+。
Step 2 preparation of 2- (Isopropoxycarbonyl) -5-methoxybenzoic acid (intermediate A1-2)
Isopropyl 2-bromo-4-methoxybenzoate (5.5 g, 20.40 mmol), oxalic acid (3.67 g, 40.8 mmol), Pd (OAc)2(0.458 g, 2.040 mmol) and XantPhos (1.181 g, 2.040 mmol) were dissolved in DMF (20 mL). Ac was added slowly with stirring at 0 deg.C2O (4.17 g, 40.8 mmol) and DIEA (5.27 g, 40.8 mmol) were added and after the addition the reaction mixture was stirred at 100 ℃ for 2 hours, then quenched by addition of ice water and extracted by dilution with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous dry sodium sulfate, then filtered and concentrated to give the crude product, which was purified by silica gel column chromatography to give 2- (isopropoxycarbonyl) -5-methoxybenzoic acid (2.2 g, 45.8%) as a yellow oil. MS: 239.1 (M + H)+。
Step 3 preparation of tert-butyl 2- (2- (isopropyloxycarbonyl) -5-methoxybenzoyl) -1-methylhydrazine-1-carboxylate (intermediate A1-3)
2- (Isopropoxycarbonyl) -5-methoxybenzoic acid (2.0 g, 8.40 mmol) and tert-butyl 1-methylhydrazine-1-carboxylate (1.84 g, 12.60 mmol) were dissolved in DMF (30 mL). HATU (4.80 g, 12.60 mmol) and DIEA (2.17 g, 16.80 mmol) were added slowly with stirring at 0 ℃ and after addition the reaction mixture was stirred for 2 hours at 20 ℃ before quenching with ice water and diluting the extract with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous dry sodium sulfate, then filtered and concentrated to give the crude product, which was purified by silica gel column chromatography to give tert-butyl 2- (2- (isopropoxycarbonyl) -5-methoxybenzoyl) -1-methylhydrazine-1-carboxylate (2.82 g, 91.2%) as a yellow oil. MS: 366.2 (M + H)+。
Step 4 preparation of 6-methoxy-2-methyl-2, 3-dihydrophthalazine-1, 4-dione (intermediate A1-4)
A solution of tert-butyl 2- (2- (isopropyloxycarbonyl) -5-methoxybenzoyl) -1-methylhydrazine-1-carboxylate (2.82 g, 7.65 mmol) in TFA (20 ml) was stirred at 70 ℃ for 12 hours and then concentrated to give the crude product, which was purified by silica gel column chromatography to give 6-methoxy-2-methyl-2, 3-dihydrophthalazine as a white solid-1, 4-dione (1.4 g, 88.6%). MS: 207.1 (M + H)+。
Step 5 preparation of 4-chloro-6-methoxy-2-methylphthalazin-1 (2H) -one (intermediate A1-5)
The mixture of 6-methoxy-2-methyl-2, 3-dihydrophthalazine-1, 4-dione (500 mg, 2.42 mmol) in POCl3The solution (5 ml) was stirred at 110 ℃ for 1 hour and then concentrated to give the crude product, which was dissolved in ethyl acetate and quenched with ice water, the organic phase was washed with saturated brine, dried over anhydrous dry sodium sulfate and then filtered, concentrated to give the crude product, which was purified by silica gel column chromatography to give 4-chloro-6-methoxy-2-methylphthalazin-1 (2H) -one (250 mg, 45.9%) as a white solid. MS: 225.1 (M + H)+。
Step 6 preparation of (R) -6-methoxy-2-methyl-4- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) phthalazin-1 (2H) -one (intermediate A1-6)
4-chloro-6-methoxy-2-methylphthalazin-1 (2H) -one (500 mg, 2.226 mmol) and (R) -1- (3-nitro-5- (trifluoromethyl) phenyl) ethan-1-amine (intermediate B1, 625 mg, 2.67 mmol) were dissolved in 1,4-dioxane (10 mL). Sodium tert-butoxide (642 mg, 6.68 mmol) and XPhos Pd 3 (186 mg, 0.22 mmol) were added slowly with stirring, and after addition was complete, the reaction mixture was stirred at 80 ℃ for an additional 2 hours, then quenched by addition of ice water and extracted by dilution with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous dry sodium sulfate, then filtered and concentrated to give the crude product, which was purified by silica gel column chromatography to give (R) -6-methoxy-2-methyl-4- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) phthalazin-1 (2H) -one (650 mg, 69.1%) as a yellow solid. MS: 423.1 (M + H)+。
Step 7 preparation of (R) -6-hydroxy-2-methyl-4- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) phthalazin-1 (2H) -one (intermediate A1-7)
(R) -6-methoxy-2-methyl-4- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) phthalazin-1 (2H) -one (150 mg, 0.355 mmol) was dissolved in DCM (2 mL). Slowly add 1.0M boron tribromide dichloromethane solution (1) with 0 ℃ stirringMl), after the addition was complete, the reaction mixture was stirred at 20 ℃ for a further 12 hours, then quenched by addition of ice water and extracted by dilution with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous dry sodium sulfate, then filtered and concentrated to give the crude product, which was purified by silica gel column chromatography to give (R) -6-hydroxy-2-methyl-4- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) phthalazin-1 (2H) -one (120 mg, 83%) as a yellow solid. MS: 409.1 (M + H)+。
Step 8 preparation of (R) -2-methyl-4- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) -1-oxo-1, 2-dihydrophthalazin-6-yl trifluoromethanesulfonate (intermediate A1)
(R) -6-hydroxy-2-methyl-4- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) phthalazin-1 (2H) -one (1.0 g, 2.449 mmol) was dissolved in DCM (15 ml). TEA (0.743 g, 7.35 mmol) and triflic anhydride (1.382 g, 4.90 mmol) were added slowly with stirring at 0 deg.C, after which the reaction mixture was stirred at 20 deg.C for an additional 12 hours, then quenched by addition of ice water and extracted by dilution with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous dry sodium sulfate, then filtered and concentrated to give the crude product, which was purified by silica gel column chromatography to give (R) -2-methyl-4- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) -1-oxo-1, 2-dihydrophthalazin-6-yl trifluoromethanesulfonate as a yellow solid (700 mg, 52.9%). MS: 541.1 (M + H)+。
Intermediate A2 (R) -7-methoxy-2-methyl-4- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) -1-oxy-1, 2-dihydrophthalazin-6-yl trifluoromethanesulfonate
Figure 754311DEST_PATH_IMAGE010
Step 1 preparation of 2-bromo-4-isopropoxy-5-methoxybenzaldehyde (intermediate A2-1)
2-bromo-4-hydroxy-5-methoxybenzaldehyde (63.5 g, 220 mmol) was dissolved in DMF (200 ml). Slowly adding K under stirring2CO3(30.4 g, 220 mm)Moles) and 2-iodopropane (112 g, 659 mmol), after addition the reaction mixture was stirred at 50 ℃ for a further 12 hours, then quenched by addition of ice water and extracted by dilution with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous dry sodium sulfate, then filtered and concentrated to give 2-bromo-4-isopropoxy-5-methoxybenzaldehyde as an orange oil (60 g, 82%). MS: 273.2 (M + H)+。
Step 2 preparation of 2-bromo-4-isopropoxy-5-methoxybenzoic acid (intermediate A2-2)
2-bromo-4-isopropoxy-5-methoxybenzaldehyde (60 g, 220 mmol) was dissolved in DMF (200 ml). Sodium hydroxide (17.57 g, 439 mmol) was added slowly with stirring, followed by potassium permanganate (69.4 g, 439 mmol) in DMF (300 ml), after completion of the reaction, the reaction was quenched by addition of ice water and extracted by dilution with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous dry sodium sulfate, then filtered and concentrated to give 2-bromo-4-isopropoxy-5-methoxybenzoic acid (63.5 g, 100%) as a white solid. MS: 289.1 (M + H)+。
Step 3 preparation of 6-isopropoxy-7-methoxy-2-methyl-2, 3-dihydrophthalazine-1, 4-dione (intermediate A2-6)
Synthesis of intermediate A2-6 referring to intermediate A1-4, intermediate A2-6 was prepared by using (2-bromo-4-isopropoxy-5-methoxybenzoic acid (intermediate A2-2) in place of 2-bromo-4-methoxybenzoic acid to afford intermediate A2-6. MS: 265.4 (M + H)+。
Step 4 preparation of 4-chloro-6-isopropoxy-7-methoxy-2-methylphthalazin-1 (2H) -one (intermediate A2-7)
6-Isopropoxy-7-methoxy-2-methyl-2, 3-dihydrophthalazine-1, 4-dione (6.7 g, 25.4 mmol) and phosphorus pentachloride (7.92 g, 38.0 mmol) in POCl3(30 ml) the solution was stirred at 110 ℃ for 1 hour and then concentrated to give the crude product, which was dissolved in ethyl acetate and quenched with ice water, the organic phase was washed with saturated sodium bicarbonate solution, saturated brine, dried over anhydrous dry sodium sulfate, then filtered, concentrated to give the crude product, which was purified by silica gel column chromatography to give 4-chloro-6-isopropoxy-7-methoxy-2-methyl-solid as a white solidPhthalazin-1 (2H) -one (4.3 g, 60%). MS: 283.1 (M + H)+。
Step 5 preparation of (R) -6-isopropoxy-7-methoxy-2-methyl-4- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) phthalazin-1 (2H) -one (intermediate A2-8)
Synthesis of intermediate A2-8 with reference to intermediate A1-6, intermediate A2-8 was prepared by using intermediate A2-7 instead of intermediate A1-5. MS: 481.2 (M + H)+。
Step 6 preparation of (R) -6-hydroxy-7-methoxy-2-methyl-4- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) phthalazin-1 (2H) -one (intermediate A2-9)
(R) -6-isopropoxy-7-methoxy-2-methyl-4- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) phthalazin-1 (2H) -one (120 mg, 0.250 mmol) was dissolved in DCM (5 mL). Boron trichloride (344 mg, 0.500 mmol) was added slowly with stirring at-78 deg.C, and after the addition was complete, the reaction mixture was stirred at 20 deg.C for an additional 5 hours, then quenched by addition of an ice-water solution of ammonium chloride and extracted by dilution with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous dry sodium sulfate, then filtered and concentrated to give (R) -6-hydroxy-7-methoxy-2-methyl-4- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) phthalazin-1 (2H) -one (110 mg) as a yellow solid. MS: 439.2 (M + H)+。
Step 7 preparation of (R) -7-methoxy-2-methyl-4- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) -1-oxo-1, 2-dihydrophthalazin-6-yl trifluoromethanesulfonate (intermediate A2)
Synthesis of intermediate a2 referring to intermediate a1, intermediate a2 was prepared by using intermediate a2-9 instead of intermediate a 1-7. MS 570.0 (M + H)+。
Intermediate A3 (R) -3-methyl-1- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) -4-oxo-3, 4-dihydropyridine [3,4-d ] pyridazine-7-trifluoromethane sulfonate
Figure 243061DEST_PATH_IMAGE011
Step 1 preparation of methyl 5-bromo-2-methoxyisonicotinate (intermediate A3-1)
5-bromo-2-methoxyisonicotinal (21.6 g, 100 mmol) was dissolved in MeOH (100 mL). A solution of iodine (33.0 g, 130 mmol) in methanol (75 ml) and potassium hydroxide (14.59 g, 260 mmol) in methanol (75 ml) were added slowly with stirring at 0 deg.C, and after addition, the reaction mixture was stirred for an additional 2 hours at 20 deg.C, then NaHSO was added3The reaction was quenched with aqueous solution, stirred, filtered and the solid dried to give methyl 5-bromo-2-methoxyisonicotinate (21 g, 85%) as a white solid. MS: 247.1& 249.1 (M+H)+。
Step 2 preparation of methyl 5- (2- (tert-Butoxycarbonyl) -1-methylhydrazine-1-carbonyl) -2-methoxyisonicotinate (intermediate A3-3)
Synthesis of intermediate A3-3 with reference to intermediate A1-3, intermediate A3-3 was prepared by using intermediate A3-1 in place of intermediate A1-1 and tert-butyl 2-methylhydrazine-1-carboxylate in place of tert-butyl 1-methylhydrazine-1-carboxylate. MS: 340.1 (M + H)+。
Step 3 preparation of 7-methoxy-3-methyl-2, 3-dihydropyrido [3,4-d ] pyridazine-1, 4-dione (intermediate A3-4)
A solution of methyl 5- (2- (tert-butoxycarbonyl) -1-methylhydrazine-1-carbonyl) -2-methoxyisonicotinate (11 g, 32.4 mmol) in TFA (24 ml) was reacted at 20 ℃ with stirring for 12 hours. Then concentrating the reaction solution and evaporating to dryness to obtain a white solid 7-methoxy-3-methyl-2, 3-dihydropyrido [3,4-d ]]Pyridazine-1, 4-dione (4.6 g, 68.5%). MS: 208.1 (M + H)+。
Step 4 preparation of 1-chloro-7-methoxy-3-methylpyridin [3,4-d ] pyridazin-4 (3H) -one (intermediate A3-5)
Reacting 7-methoxy-3-methyl-2, 3-dihydropyrido [3,4-d ]]Pyridazine-1, 4-dione (4.6 g, 22.20 mmol), DIEA (4.30 g, 33.3 mmol) and phosphorus pentachloride (6.93 g, 33.3 mmol) in POCl3(30 ml) solution is stirred and reacted for 1 hour at 100 ℃, then the reaction liquid is cooled, yellow solid is separated out, the filter cake is washed by dichloromethane and dried to obtain yellow solid 1-chloro-7-methoxy-3-methylpyridine [3,4-d ]]Pyridazin-4 (3H) -one (2.8 g, 55.9%). MS: 226.1 (M + H)+。
Step 5 preparation of (R) -7-methoxy-3-methyl-1- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) pyrido [3,4-d ] pyridazin-4 (3H) -one (intermediate A3-6)
Sodium tert-butoxide (10.5 mmol), 1-chloro-7-methoxy-3-methylpyridine [3,4-d]Pyridazin-4 (3H) -one (1.6 g, 7.09 mmol), (R) -1- (3-nitro-5- (trifluoromethyl) phenyl) ethan-1-amine (1.919 g, 7.09 mmol) and a solution of methanesulfonic acid (2-dicyclohexylphosphine-2 ", 4", 6 "-triisopropyl-1, 1" -biphenyl) (2 "-methylamino-1, 1" -biphenyl-2-yl) palladium (II) (0.600 g, 0.709 mmol) in toluene (50 ml) were reacted at 90 ℃ with stirring for 12 hours, then quenched by addition of ice water and extracted by dilution with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous dry sodium sulfate, then filtered and concentrated to give a crude product, which was purified by silica gel column chromatography to give (R) -7-methoxy-3-methyl-1- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) pyrido [3, 4-d-as a yellow solid]Pyridazin-4 (3H) -one (1.0 g, 33.3%). MS: 424.1 (M + H)+。
Step 6 preparation of (R) -7-hydroxy-3-methyl-1- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) pyrido [3,4-d ] pyridazin-4 (3H) -one (intermediate A3-7)
Reacting (R) -7-methoxy-3-methyl-1- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) pyrido [3,4-d]Pyridazin-4 (3H) -one (940 mg, 2.220 mmol) and NaI (998 mg, 6.66 mmol) were dissolved in acetonitrile (45 ml), TMSCl (724 mg, 6.66 mmol) was slowly added with stirring at 20 degrees, and then the reaction solution was stirred at 80 degrees for 3 hours. The reaction solution was then quenched with ice water and extracted with ethyl acetate, and the organic phase was washed with saturated sodium bicarbonate solution, saturated brine, dried over anhydrous dry sodium sulfate, then filtered and concentrated to give (R) -7-hydroxy-3-methyl-1- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) pyrido [3,4-d as a pale brown solid]Pyridazin-4 (3H) -one (740 mg, 79%). MS 410.1 (M + H)+。
Step 7 preparation of (R) -3-methyl-1- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) -4-oxo-3, 4-dihydropyridine [3,4-d ] pyridazin-7-yl trifluoromethanesulfonate (intermediate A3)
Reacting (R) -7-hydroxy-3-methyl-1- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) pyrido [3,4-d]Pyridazin-4 (3H) -one (740 mg, 1.808 mmol) and pyridine (286 mg, 3.62 mmol) were dissolved in DCM (45 mL) and Tf was added slowly with stirring at 20 deg.C2O (1020 mg, 3.62 mmol), and then the reaction solution was stirred at 20 ℃ for 2 hours. Then quenching the reaction solution with ice water, extracting with ethyl acetate, washing the organic phase with saturated brine, drying over anhydrous dry sodium sulfate, filtering, concentrating to obtain a crude product, and purifying the crude product by silica gel column chromatography to obtain yellow solid (R) -3-methyl-1- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) -4-oxo-3, 4-dihydropyridine [3, 4-d-]Pyridazin-7-yl trifluoromethanesulfonate (310 mg, 31.7%). MS: 542.1 (M + H)+。
Intermediate A4 (R) -7-fluoro-2-methyl-4- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) -1-oxo-1, 2-dihydrophthalazin-6-yl trifluoromethanesulfonate
Figure 511231DEST_PATH_IMAGE012
Step 1 preparation of 2-bromo-5-fluoro-4-methoxybenzoic acid (intermediate A4-1)
2-bromo-4, 5-difluorobenzoic acid (20 g, 84 mmol) was dissolved in DMF (150 ml), MeONa (6.84 g, 127 mmol) was added slowly with stirring at 20 degrees, and the reaction solution was then reacted for 12 hours with stirring at 90 degrees. The reaction was then quenched with ice water and extracted with ethyl acetate, the organic phase was washed with saturated brine, dried over anhydrous dry sodium sulfate, then filtered and concentrated to give the crude product which was purified by silica gel column chromatography to give 2-bromo-5-fluoro-4-methoxybenzoic acid (20.0 g, 95%) as a white solid. MS 248.1& 250.1 (M+H)+。
Step 2 preparation of (R) -7-fluoro-2-methyl-4- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) -1-oxo-1, 2-dihydrophthalazin-6-yl trifluoromethanesulfonate (intermediate A4)
Synthesis of intermediate A4 reference intermediate A1 was made by using intermediate A4-1 generationThe intermediate A4 was obtained instead of 2-bromo-4-methoxybenzoic acid. MS: 559.2 (M + H)+。
Intermediate A5 (R) -2, 7-dimethyl-4- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) -1-oxo-1, 2-dihydrophthalazin-6-yl trifluoromethanesulfonate
Figure 706720DEST_PATH_IMAGE013
Synthesis of intermediate a5 referring to intermediate a4, intermediate a5 was prepared by using 2-bromo-4-fluoro-5-methylbenzoic acid instead of 2-bromo-4, 5-difluorobenzoic acid. MS: 555.2 (M + H)+。
Intermediate B1 (R) -1- (3-nitro-5- (trifluoromethyl) phenyl) ethan-1-amine
Figure 870985DEST_PATH_IMAGE014
Step 1 preparation of 1- (3-Nitro-5- (trifluoromethyl) phenyl) ethan-1-one (intermediate B1-1)
1-bromo-3-nitro-5- (trifluoromethyl) benzene (50.0 g, 185.18 mmol) and tributyl (1-ethoxyvinyl) stannane (80.26 g, 222.22 mmol) were dissolved in 1,4-dioxane (500 ml). Pd (PPh) was added slowly with stirring3)2Cl2 (4.07 g, 9.26 mmol) and TEA (37.48 g, 370.36 ml), after addition, the reaction mixture was stirred at 80 ℃ for an additional 12 hours, then quenched by addition of ice water and extracted by dilution with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous dry sodium sulfate, then filtered, and concentrated to give a crude product, which was dissolved in 2N hydrochloric acid (200 ml), and the reaction mixture was reacted with stirring at 20 ℃ for 2 hours, and extracted with dilution with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous dry sodium sulfate, then filtered and concentrated to give the crude product, which was purified by silica gel column chromatography to give 1- (3-nitro-5- (trifluoromethyl) phenyl) ethan-1-one (28.0 g, 64.86%) as a yellow oil.
Step 2 preparation of (R, Z) -2-methyl-N- (1- (3-nitro-5- (trifluoromethyl) phenyl) ethylene) propane-2-sulfinylamine (intermediate B1-2)
1- (3-nitro-5- (trifluoromethyl) phenyl) ethan-1-one (28.0 g, 120.10 mmol) was dissolved in THF (300 mL). (R) -2-methylpropane-2-sulfinylamine (21.83 g, 180.14 mmol) and Ti (OEt) were added slowly with stirring4(41.09 g, 180.14 mmol), and after addition, the reaction mixture was stirred at 60 ℃ for 2 hours, then quenched by addition of ice water and extracted by dilution with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous dry sodium sulfate, then filtered and concentrated to give the crude product, which was purified by silica gel column chromatography to give (R, Z) -2-methyl-N- (1- (3-nitro-5- (trifluoromethyl) phenyl) ethylene) propane-2-sulfinylamine (25.5 g, 63.1%) as a yellow solid. MS: 337.1 (M + H)+。
Step 3 preparation of (R) -2-methyl-N- ((R) -1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) propane-2-sulfinylamine (intermediate B1-3)
(R, Z) -2-methyl-N- (1- (3-nitro-5- (trifluoromethyl) phenyl) ethylene) propane-2-sulfinylamine (25.5 g, 75.82 mmol) was dissolved in THF (250 mL) and water (5 mL). Slowly add NaBH under-78 ℃ stirring4(3.73 g, 98.56 mmol), after addition, the reaction mixture was stirred at-78 ℃ for 2 hours, then quenched by addition of ice water and extracted by dilution with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous dry sodium sulfate, then filtered and concentrated to give the crude product, which was purified by silica gel column chromatography to give (R) -2-methyl-N- ((R) -1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) propane-2-sulfinylamine as a yellow solid (20.1 g, 78.3%). MS: 339.1 (M + H)+。
Step 4 preparation of (R) -1- (3-nitro-5- (trifluoromethyl) phenyl) ethan-1-amine (intermediate B1)
(R) -2-methyl-N- ((R) -1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) propane-2-sulfinylamine (20.1 g, 62.4 mmol) was dissolved in 4N hydrogen chloride 1,4-dioxane solution (20 ml). The reaction mixture was stirred at 20 ℃ for 1 hour and then concentrated to give the crude product which was washed with MTBE to give (R) -1- (3-nitro-5- (trifluoromethyl) methyl tert-butyl ether as a white solid) Phenyl) ethan-1-amine (12.5 g, 89.86%). MS 235.1 (M + H)+。
Intermediate B2 (R) -tetrahydrofuran-3-yl 4-tosylate
Figure 909349DEST_PATH_IMAGE015
(R) -tetrahydrofuran-3-ol (5.0 g, 56.7 mmol) was dissolved in DCM (60 mL). TsCl (12.98 g, 68.1 mmol), TEA (8.61 g, 85 mmol) and DMAP (0.693 g, 5.67 mmol) were added slowly with stirring at 0 deg.c, after which the reaction mixture was stirred at 20 deg.c for an additional 12 hours, then quenched by addition of ice water and extracted by dilution with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous dry sodium sulfate, then filtered and concentrated to give the crude product, which was purified by silica gel column chromatography to give (R) -tetrahydrofuran-3-yl 4-tosylate (8.5 g, 61.8%) as a colorless oil.
Intermediate B3 tert-butyl (R) -3- (p-toluenesulfonyloxy) pyrrolidine-1-carboxylate
Figure 918893DEST_PATH_IMAGE016
Synthesis of intermediate B3 referring to intermediate B2, intermediate B3 was prepared by using (R) -3-hydroxypyrrolidine-1-carboxylic acid tert-butyl ester instead of (R) -tetrahydrofuran-3-ol. MS: 342.2 (M + H)+。
Intermediate B4: 4- (p-toluenesulfonyloxy) piperidine-1-carboxylic acid tert-butyl ester
Figure 467424DEST_PATH_IMAGE017
Synthesis of intermediate B4 referring to intermediate B2, intermediate B4 was prepared by using tert-butyl 4-hydroxypiperidine-1-carboxylate instead of (R) -tetrahydrofuran-3-ol. MS: 356.2 (M + H)+。
Intermediate B5: (3S,3aS,6R,6aR) -6-methoxy hexahydro-furan [3,2-b ] furan-3-yl 4-methylbenzenesulfonate
Figure 599328DEST_PATH_IMAGE018
Step 1: preparation of (3R,3aS,6S,6aS) -6- (tert-butyldiphenylsiloxy) hexahydrofuro [3,2-B ] furan-3-yl nitrate (intermediate B5-1)
Reacting (3R,3aS,6S,6aR) -6-hydroxy hexahydrofuran [3,2-b ]]Furan-3-yl nitrate (10 g, 52.3 mmol) was dissolved in DCM (200 ml). T-butyldiphenylchlorosilane (21.57 g, 78 mmol) and imidazole (7.12 g, 105 mmol) were added slowly with stirring at 0 deg.c, and after addition was complete, the reaction mixture was stirred at 20 deg.c for an additional 2 hours. The reaction was then quenched by addition of ice water and extracted by dilution with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous dry sodium sulfate, then filtered and concentrated to give a crude product, which was purified by silica gel column chromatography to give (3R,3aS,6S,6aS) -6- (tert-butyldiphenylsiloxy) hexahydrofuro [3,2-b ] aS a colorless oil]Furan-3-yl nitrate (18 g, 80%). MS: 452.2 (M + Na)+。
Step 2 preparation of (3R,3aR,6S,6aS) -6- ((tert-butyldiphenylsilyl) oxy) hexahydrofuran [3,2-B ] furan-3-ol (intermediate B5-2)
Reacting (3R,3aS,6S,6aS) -6- (tert-butyl diphenyl siloxy) hexahydrofuran [3,2-b]Furan-3-yl nitrate (18 g, 41.9 mmol) was dissolved in methanol (100 ml) and ethyl acetate (100 ml). Palladium on carbon (5 g) was added with stirring at 20 ℃ and after completion of the addition, the reaction mixture was reacted with stirring under hydrogen at 20 ℃ for 12 hours, followed by filtration and concentration to give a crude product, which was purified by pre-HPLC to give (3R,3aR,6S,6aS) -6- ((tert-butyldiphenylsilyl) oxy) hexahydrofuran [3,2-b ] aS a colorless oil]Furan-3-ol (15.0 g, 93%). MS: 407.2 (M + Na)+。
Step 3 tert-butyl (((3S,3aS,6R,6aR) -6-methoxyhexahydrofuro [3,2-B ] furan-3-yl) oxy) diphenylsilane (intermediate B5-3)
Reacting (3R,3aS,6S,6aR) -6- ((tert-butyl diphenyl silicon) oxy) hexahydrofuran [3,2-b]Furan-3-ol (4.0 g, 10.40 mmol) was dissolved in DMF (30 ml). Slowly adding the mixture under stirring at 0 DEG CNaH (0.300 g, 12.48 mmol) was added and, after the addition was complete, the reaction mixture was stirred at 0 ℃ for an additional 0.5 h, then iodomethane (2.215 g, 15.60 mmol) was added to the reaction and stirring continued at room temperature for an additional 2 h. The reaction was then quenched by addition of ice water and extracted by dilution with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous dry sodium sulfate, then filtered and concentrated to give a crude product, which was purified by silica gel column chromatography to give t-butyl (((3S,3aS,6R,6aR) -6-methoxyhexahydrofuro [3,2-b ] -a colorless oil]Furan-3-yl) oxy) diphenylsilane (2.8 g, 67.5%). MS: 421.2 (M + Na)+。
Step 4 preparation of (3S,3aR,6R,6aR) -6-methoxyhexahydrofuro [3,2-B ] furan-3-ol (intermediate B5-4)
Tert-butyl (((3S,3aS,6R,6aR) -6-methoxy hexahydrofuran [3, 2-b)]Furan-3-yl) oxy) diphenylsilane (2.8 g, 7.03 mmol) was dissolved in THF (20 ml). A solution of 1M TBAF in THF (10.5 mL) was added slowly with stirring at 20 ℃ and after addition was complete, the reaction mixture was stirred for an additional 1 hour at 20 ℃. The reaction was then quenched by addition of ice water and extracted by dilution with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous dry sodium sulfate, then filtered and concentrated to give a crude product, which was purified by silica gel column chromatography to give (3S,3aR,6R,6aR) -6-methoxyhexahydrofuran [3,2-b ] as a colorless oil]Furan-3-ol (800 mg, 71.1%). MS: 161.2 (M + H)+。
Step 5 preparation of (3S,3aS,6R,6aR) -6-methoxyhexahydrofuro [3,2-B ] furan-3-yl 4-methylbenzenesulfonate (intermediate B5)
Reacting (3S,3aR,6R,6aR) -6-methoxy hexahydrofuran [3,2-b]Furan-3-ol (300 mg, 1.873 mmol) was dissolved in DCM (10 ml). P-methanesulfonyl chloride (536 mg, 2.81 mmol), TEA (379 mg, 3.75 mmol) and DMAP (22.88 mg, 0.187 mmol) were added slowly with stirring at 0 deg.c, and after addition was complete, the reaction mixture was stirred at 20 deg.c for an additional 12 hours. The reaction mixture was then quenched with ice water and extracted by dilution with ethyl acetate. Washing the organic phase with saturated saline, drying with anhydrous dry sodium sulfate, filtering, concentrating to obtain crude product, and purifying by silica gel column chromatography to obtainTo a white solid (3S,3aS,6R,6aR) -6-methoxy-hexahydrofuran [3,2-b ]]Furan-3-yl 4-methylbenzenesulfonate (400 mg, 67.9%). MS: 315.1 (M + H)+。
Examples
Compound I-1: 4- (((R) -1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -2-methyl-6- (((S) -tetrahydrofuran-3-yl) oxy) phthalazin-1 (2H) -one
Figure 62670DEST_PATH_IMAGE019
Step 1 preparation of 2-methyl-4- (((R) -1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) -6- (((S) -tetrahydrofuran-3-yl) oxy) phthalazin-1 (2H) -one (intermediate I-1-1)
(R) -6-hydroxy-2-methyl-4- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) phthalazin-1 (2H) -one (intermediate A1-7, 120 mg, 0.294 mmol) and (R) -tetrahydrofuran-3-yl 4-tosylate (intermediate B2,107 mg, 0.441 mmol) were dissolved in DMF (2 mL). Slowly add Cs under agitation2CO3(144 mg, 0.441 mmol), after the addition was complete, the reaction mixture was stirred at 80 ℃ for an additional 1 hour, then quenched by addition of ice water and extracted by dilution with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous dry sodium sulfate, then filtered and concentrated to give the crude product, which was purified by silica gel column chromatography to give 2-methyl-4- (((R) -1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) -6- (((S) -tetrahydrofuran-3-yl) oxy) phthalazin-1 (2H) -one (100 mg, 671.1%) as a yellow solid. MS: 479.2 (M + H)+。
Step 2 preparation of 4- (((R) -1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -2-methyl-6- (((S) -tetrahydrofuran-3-yl) oxy) phthalazin-1 (2H) -one (Compound I-1)
2-methyl-4- (((R) -1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) -6- (((S) -tetrahydrofuran-3-yl) oxy) phthalazin-1 (2H) -one (100 mg, 0.209 mmol) was dissolved in methanol (2 mL). Palladium on carbon (30 mg) was added under stirring at 20 ℃ and, after the addition was completed, the reaction mixture was reacted under stirring at 20 ℃ for 2 hours, thenPost-filtration and concentration gave the crude product which was purified by pre-HPLC to give 4- (((R) -1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -2-methyl-6- (((S) -tetrahydrofuran-3-yl) oxy) phthalazin-1 (2H) -one as a white solid (65 mg, 69.3%). MS: 449.2 (M + H)+。1H NMR (400 MHz, DMSO-d 6 ) δ 8.13 (d, J = 8.8 Hz, 1H), 7.72 (s, 1H), 7.42-7.35 (m, 1H), 6.93-6.80 (m, 3H), 6.69 (s, 1H), 5.32-5.26 (m, 1H), 4.92-4.87 (m, 1H), 3.99-3.95 (m, 1H), 3.91-3.76 (m, 5H), 3.41 (s, 3H), 2.35-2.28 (mz, 1H), 2.04-1.95 (m, 1H), 1.49 (d, J = 7.0 Hz, 3H)。
Compound I-2: (R) -4- ((1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -6-methoxy-2-methylphthalazin-1 (2H) -one
Figure 813588DEST_PATH_IMAGE020
Synthesis of compound I-2 referring to compound I-1, compound I-2 was prepared as a white solid by using (R) -6-methoxy-2-methyl-4- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) phthalazin-1 (2H) -one (intermediate a1-6) instead of 2-methyl-4- (((R) -1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) -6- (((S) -tetrahydrofuran-3-yl) oxy) phthalazin-1 (2H) -one (intermediate I-1-1).
Compound I-2 (35 mg), MS: 393.2 (M + H)+。1H NMR (400 MHz, DMSO-d 6) δ 8.12 (d, J = 8.7 Hz, 1H), 7.72 (s, 1H), 7.36 (dd, J = 8.9, 2.2 Hz, 1H), 6.97 (d, J= 7.0 Hz, 1H), 6.88 (d, J = 9.9 Hz, 2H), 6.69 (s, 1H), 4.91 (t, J = 6.9 Hz, 1H), 3.95 (s, 3H), 3.41 (s, 3H), 1.50 (d, J = 6.9 Hz, 3H)。
Compound I-3: 6- (((S) -1-acetylpyrrolidin-3-yl) oxy) -4- (((R) -1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -2-methylphthalazin-1 (2H) -one
Figure 45987DEST_PATH_IMAGE021
Step 1 preparation of tert-butyl (S) -3- ((2-methyl-4- ((R) -1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) -1-oxo-1, 2-dihydrophthalocyanin-6-yl) oxy) pyrrolidine-1-carboxylate (intermediate I-3-1)
Synthesis of intermediate I-3-1 reference was made to intermediate I-1-1 by using (R) -3- (p-toluenesulfonyloxy) pyrrolidine-1-carboxylic acid tert-butyl ester (intermediate B3) in place of (R) -tetrahydrofuran-3-yl 4-tosylate (intermediate B2). MS: 578.2 (M + H)+。
Step 2 preparation of 2-methyl-4- (((R) -1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) -6- (((S) -pyrrolidin-3-yl) oxy) phthalazin-1 (2H) -one (intermediate I-3-2)
A reaction mixture of (S) -tert-butyl 3- ((2-methyl-4- ((R) -1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) -1-oxo-1, 2-dihydrophthalocyanin-6-yl) oxy) pyrrolidine-1-carboxylate (150 mg, 0.260 mmol) in TFA (2 ml) was stirred at 20 ℃ for 1 hour, then concentrated to give 2-methyl-4- (((R) -1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) -6- (((S) -pyrrolidin-3-yl) oxy) phthalazin-1 (2H) -one (125 mg) as a yellow oil. MS: 478.2 (M + H)+。
Step 3 preparation of 6- (((S) -1-acetylpyrrolidin-3-yl) oxy) -2-methyl-4- (((R) -1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) phthalazin-1 (2H) -one (intermediate I-3-3)
2-methyl-4- (((R) -1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) -6- (((S) -pyrrolidin-3-yl) oxy) phthalazin-1 (2H) -one (125 mg) was dissolved in DCM (2 mL). Acetyl chloride (1.5 mg, 0.019 mmol) and TEA (3 mg, 0.030 mmol) were added slowly with stirring at 0 deg.c, and after the addition was complete, the reaction mixture was stirred at 20 deg.c for an additional 1 hour, then quenched by addition of ice water and extracted by dilution with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous dry sodium sulfate, then filtered and concentrated to give the crude product, which was purified by silica gel column chromatography to give 6- (((S) -1-acetylpyrrolidin-3-yl) oxy) -2-methyl-4- (((R) -1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) phthalazin-1 (2H) -one (65 mg, 48.2%) as a yellow solid. MS (Mass Spectrometry): 520.2 (M+H)+。
Step 4 preparation of 6- (((S) -1-acetylpyrrolidin-3-yl) oxy) -4- (((R) -1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -2-methylphthalazin-1 (2H) -one (Compound I-3)
6- (((S) -1-acetylpyrrolidin-3-yl) oxy) -2-methyl-4- (((R) -1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) phthalazin-1 (2H) -one (90 mg, 0.173 mmol) was dissolved in methanol (2 mL). Palladium on carbon (45 mg) was added with stirring at 20 degrees, and after addition, the reaction mixture was stirred under hydrogen at 20 degrees for 2 hours, followed by filtration and concentration to give a crude product, which was purified by pre-HPLC to give 6- (((S) -1-acetylpyrrolidin-3-yl) oxy) -4- (((R) -1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -2-methylphthalazin-1 (2H) -one (75 mg, 88%) as a white solid. MS: 490.2 (M + H)+。1H NMR (400 MHz, DMSO-d 6) δ 8.14 (dd, J = 8.7, 2.4 Hz, 1H), 7.76 (d, J = 10.5 Hz, 1H), 7.43 (t, J = 9.1 Hz, 1H), 6.98-6.90 (m, 3H), 6.75 (s, 1H), 5.38-5.25 (m, 1H), 4.92-4.85 (m, 1H), 3.70-3.53 (m, 3H), 3.41 (s, 3H), 3.38-3.32 (m, 1H), 2.35-2.10 (m, 2H), 1.96 (d, J= 16.3 Hz, 3H), 1.50 (d, J = 6.9 Hz, 3H)。
Compound I-4: (R) -6- ((1-acetylpiperidin-4-yl) oxy) -4- ((1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -2-methylphthalazin-1 (2H) -one
Figure 348792DEST_PATH_IMAGE022
Synthesis of Compound I-4 with reference to Compound I-3, Compound I-4 was prepared as a white solid by using tert-butyl 4- (p-toluenesulfonyloxy) piperidine-1-carboxylate (intermediate B4) in place of tert-butyl (R) -3- (p-toluenesulfonyloxy) pyrrolidine-1-carboxylate (intermediate B3).
Compound I-4 (20 mg), MS: 504.2 (M + H)+。1H NMR (400 MHz, DMSO-d 6) δ 8.13 (d, J = 8.8 Hz, 1H), 7.77 (s, 1H), 7.47 (d, J = 9.0 Hz, 1H), 6.95-6.85(m, 3H), 6.70 (s, 1H), 4.93-4.90 (m, 2H), 3.84-3.64 (m, 2H), 3.44-3.33 (m, 5H), 2.07-2.03 (m, 1H), 2.02 (s, 3H), 1.96-1.91 (m, 1H), 1.72-1.68 (m, 1H), 1.62-1.58 (m, 1H), 1.49 (d, J = 7.0 Hz, 3H)。
Compound I-5: (R) -4- ((1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -6- ((1- (dimethylglycine) piperidin-4-yl) oxy) -2-methylphthalazin-1 (2H) -one
Figure 971534DEST_PATH_IMAGE023
Synthesis of Compound I-5 with reference to Compound I-4, Compound I-5 was prepared as a white solid by using dimethylglycinoyl chloride instead of acetyl chloride.
Compound I-5 (12 mg), MS: 547.3 (M + H)+。1H NMR (400 MHz, DMSO-d 6) δ 9.49 (s, 1H), 8.15 (d, J = 8.8 Hz, 1H), 7.78 (s, 1H), 7.48 (d, J = 9.0 Hz, 1H), 6.89 (m, 3H), 6.68 (s, 1H), 4.91 (dd, J = 14.8, 8.2 Hz, 2H), 4.30 (s, 2H), 3.86-3.82 (m, 1H), 3.55-3.30 (m, 7H), 2.81 (s, 6H), 2.10-1.95 (m, 2H), 1.77-1.65 (m, 2H), 1.49 (d, J = 7.0 Hz, 3H)。
Compound I-6: (R) -2- (4- ((4- ((1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -2-methyl-1-oxo-1, 2-dihydrophthalazin-6-yl) oxy) piperidin-1-yl) acetic acid
Figure 588460DEST_PATH_IMAGE024
Synthesis of Compound I-6 with reference to Compound I-4, Compound I-6 was prepared as a white solid by using 2-bromoacetic acid instead of acetyl chloride.
Compound I-6 (32 mg), MS: 550.2 (M + H)+。1H NMR (400 MHz, DMSO-d 6) δ 8.16 (d, J = 8.9 Hz, 1H), 7.76 (s, 1H), 7.48-7.45 (m, 1H), 6.92-6.85 (m, 3H), 6.68 (s, 1H), 4.98-4.86 (m, 2H), 4.20 (s, 2H), 3.45-3.32 (m, 7H), 2.30-2.20 (m, 4H), 1.49 (d, J = 6.9 Hz, 3H)。
Compound I-7: (R) -4- ((1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -2-methyl-6- (1-methyl-2-oxo-1, 2-dihydropyridin-4-yl) phthalazin-1 (2H) -one
Figure 737682DEST_PATH_IMAGE026
Step 1 preparation of (R) -2-methyl-6- (1-methyl-2-oxo-1, 2-dihydropyridin-4-yl) -4- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) phthalazin-1 (2H) -one (intermediate I-7-1)
(R) -2-methyl-4- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) -1-oxo-1, 2-dihydrophthalazin-6-yl trifluoromethanesulfonate (intermediate A1, 100 mg, 0.185 mmol) and 1-methyl-2-oxo-1, 2-dihydropyridine-4-boronic acid pinacol ester (65.3 mg, 0.278 mmol) were dissolved in 1,4-Dioxane (2 ml) and H2O (0.4 ml). PdCl was added slowly with stirring2(dppf) (13.54 mg, 0.019 mmol) and K2CO3(77 mg, 0.555 mmol), after addition was complete, the reaction mixture was stirred at 80 ℃ for 1 hour, then quenched by addition of ice water and extracted by dilution with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous dry sodium sulfate, then filtered and concentrated to give the crude product, which was purified by silica gel column chromatography to give (R) -2-methyl-6- (1-methyl-2-oxo-1, 2-dihydropyridin-4-yl) -4- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) phthalazin-1 (2H) -one (55 mg, 59.5%) as a yellow solid. MS 500.2 (M + H)+。
Step 2 preparation of (R) -4- ((1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -2-methyl-6- (1-methyl-2-oxo-1, 2-dihydropyridin-4-yl) phthalazin-1 (2H) -one (Compound I-7)
Synthesis of Compound I-7 reference compound I-1 was prepared by using (R) -2-methyl-6- (1-methyl-2-oxo-1, 2-dihydropyridin-4-yl) -4- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) phthalazin-1 (2H) -one (intermediate I-7-1) instead of 2-methyl-4- (((R) -1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) -6- (((S) -tetrahydrofuran-3-yl) oxy) phthalazin-1 (2H) -one (intermediate I-1-1) to give Compound I-7 as a white solid.
Compound I-7 (40 mg), MS: 470.2 (M + H)+。1H NMR (400 MHz, DMSO-d 6) δ 8.63 (s, 1H), 8.27 (d, J = 8.5 Hz, 1H), 8.17-8.13 (m, 1H), 7.91-7.86 (m, 1H), 7.29-7.26 (m, 1H), 7.05 (s, 1H), 6.88 (s, 2H), 6.79-6.75 (m, 1H), 6.69 (s, 1H), 4.95 (t, J = 7.0 Hz, 1H), 3.49 (s, 3H), 3.46 (s, 3H), 1.53 (d, J = 7.0 Hz, 3H)。
Compound I-8: (R) -4- ((1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -2-methyl-6- (6-oxo-1, 6-dihydropyridin-3-yl) phthalazin-1 (2H) -one
Figure 86755DEST_PATH_IMAGE027
Synthesis of Compound I-8 with reference to Compound I-7, Compound I-8 was prepared as a white solid by using 6-hydroxypyridine-3-boronic acid pinacol ester in place of 1-methyl-2-oxo-1, 2-dihydropyridine-4-boronic acid pinacol ester.
Compound I-8 (25 mg), MS: 456.2 (M + H)+。1H NMR (400 MHz, DMSO-d 6) δ 8.41 (s, 1H), 8.21-8.16 (m, 1H), 8.10-8.00 (m, 4H), 7.16-7.11 (m, 1H), 6.96-6.91 (m, 3H), 6.74 (s, 1H), 6.54 (d, J = 9.7 Hz, 1H), 4.97-4.94 (m, 1H), 3.44 (s, 3H), 1.53 (d, J = 6.9 Hz, 3H)。
Compound I-9: 4- (((R) -1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -2-methyl-6- (1-methyl-2-oxopiperidin-4-yl) phthalazin-1 (2H) -one
Figure 259110DEST_PATH_IMAGE028
Synthesis of Compound I-9 reference compound I-1 was prepared by using (R) -4- ((1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -2-methyl-6- (1-methyl-2-oxo-1, 2-dihydropyridin-4-yl) phthalazin-1 (2H) -one (Compound I-7) instead of 2-methyl-4- (((R) -1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) -6- (((S) -tetrahydrofuran-3-yl) oxy) phthalazin-1 (2H) -one (intermediate I-1-1) to give Compound I-9 as a white solid.
Compound I-9 (20 mg), MS: 474.2 (M + H)+。1H NMR (400 MHz, DMSO-d 6) δ 8.19 (s, 1H), 8.15 (d, J = 8.2 Hz, 1H), 7.74 (d, J = 8.3 Hz, 1H), 7.02 (s, 1H), 6.93 (d, J = 13.2 Hz, 2H), 6.74 (s, 1H), 4.97-4.93 (m, 1H), 3.48-3.42 (m, 4H), 3.38-3.35 (m, 1H), 3.28-3.25 (m, 1H), 2.88 (s, 3H), 2.59-2.55 (m, 2H), 2.11-1.98 (m, 2H), 1.51 (d, J = 6.9 Hz, 3H)。
Compound I-10: (R) -2- (4- (4- ((1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -2-methyl-1-oxo-1, 2-dihydrophthalazin-6-yl) piperidin-1-yl) acetic acid
Figure 476465DEST_PATH_IMAGE029
Step 1 preparation of (R) -4- (2-methyl-4- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) -1-oxo-1, 2-dihydrophthalazin-6-yl) -3, 6-dihydropyridine-1 (2H) -carboxylic acid tert-butyl ester (intermediate I-10-1)
(R) -2-methyl-4- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) -1-oxo-1, 2-dihydrophthalazin-6-yl trifluoromethanesulfonate (intermediate A1, 400 mg, 0.74 mmol) and N-tert-butoxycarbonyl-1, 2,5, 6-tetrahydropyridine-4-boronic acid pinacol ester (343 mg, 1.110 mmol) were dissolved in 1,4-Dioxane (5 ml) and H2O (1 ml). PdCl is added slowly with stirring2(dppf) (54.2 mg, 0.074 mmol) and K2CO3 (307 mg, 2.221 mmol), after addition, the reaction mixture is stirred at 80 ℃ for a further 1h, then quenched by addition of ice-water and extracted by dilution with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous dry sodium sulfate, then filtered and concentrated to give a crude product, which was purified by silica gel column chromatography to give (R) -4- (2-methyl-4- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) as a yellow solid-1-oxo-1, 2-dihydrophthalazin-6-yl) -3, 6-dihydropyridine-1 (2H) -carboxylic acid tert-butyl ester (300 mg, 70.7%). MS: 574.2 (M + H)+。
Step 2 preparation of (R) -2-methyl-4- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) -6- (1,2,3, 6-tetrahydropyridin-4-yl) phthalazin-1 (2H) -one (intermediate I-10-2)
The reaction mixture of (R) -4- (2-methyl-4- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) -1-oxo-1, 2-dihydrophthalazin-6-yl) -3, 6-dihydropyridine-1 (2H) -carboxylic acid tert-butyl ester (300 mg, 0.523 mmol) in TFA (2 ml) was stirred at 20 ℃ for 1 hour and then concentrated to give (R) -2-methyl-4- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) -6- (1,2,3, 6-tetrahydropyridin-4-yl) phthalazin-1 (2H) -one (230 mg) as a yellow solid, 93%). MS: 474.2 (M + H)+。
Step 3 preparation of ethyl (R) -2- (4- (2-methyl-4- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) -1-oxo-1, 2-dihydrophthalazin-6-yl) -3, 6-dihydropyridin-1 (2H) -yl) acetate (intermediate I-10-3)
(R) -2-methyl-4- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) -6- (1,2,3, 6-tetrahydropyridin-4-yl) phthalazin-1 (2H) -one (100 mg, 0.211 mmol) and TEA (42.7 mg, 0.422 mmol) were dissolved in DCM (2 ml). Ethyl 2-bromoacetate (52.9 mg, 0.317 mmol) was added slowly with stirring at 0 deg.c, after which the reaction mixture was stirred at 20 deg.c for an additional 1 hour, then quenched by addition of ice water and extracted with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous dry sodium sulfate, then filtered and concentrated to give the crude product, which was purified by silica gel column chromatography to give ethyl (R) -2- (4- (2-methyl-4- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) -1-oxo-1, 2-dihydrophthalazin-6-yl) -3, 6-dihydropyridin-1 (2H) -yl) acetate as a yellow solid (85 mg, 71.9%). MS 560.2 (M + H)+。
Step 4 preparation of Ethyl 2- (4- (4- ((1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -2-methyl-1-oxo-1, 2-dihydrophthalazin-6-yl) piperidin-1-yl) acetate (intermediate I-10-4)
Ethyl (R) -2- (4- (2-methyl-4- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) -1-oxo-1, 2-dihydrophthalazin-6-yl) -3, 6-dihydropyridin-1 (2H) -yl) acetate (80 mg, 0.151 mmol) was dissolved in methanol (2 ml). Palladium on carbon (40 mg) was added with stirring at 20 degrees and, after addition was complete, the reaction mixture was stirred under hydrogen at 20 degrees for 2 hours, then filtered and concentrated to give the crude product which was purified by pre-HPLC to give ethyl (R) -2- (4- (4- ((1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -2-methyl-1-oxo-1, 2-dihydrophthalazin-6-yl) piperidin-1-yl) acetate (80 mg) as a yellow oil. MS: 532.2 (M + H)+。
Step 5 preparation of (R) -2- (4- (4- ((1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -2-methyl-1-oxo-1, 2-dihydrophthalazin-6-yl) piperidin-1-yl) acetic acid (Compound I-10)
Ethyl (R) -2- (4- (4- ((1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -2-methyl-1-oxo-1, 2-dihydrophthalazin-6-yl) piperidin-1-yl) acetate (80 mg) was dissolved in methanol (1 ml) and water (1 ml). Lithium hydroxide (30.2 mg, 0.75 mmol) was added slowly with stirring at 0 deg.c, after which the reaction mixture was stirred at 20 deg.c for an additional 1 hour, then quenched by addition of ice water and extracted by dilution with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous dry sodium sulfate, then filtered and concentrated to give the crude product, which was purified by pre-HPLC to give (R) -2- (4- (4- ((1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -2-methyl-1-oxo-1, 2-dihydrophthalazin-6-yl) piperidin-1-yl) acetic acid as a white solid (23 mg, 28.3%). MS 504.2 (M + H)+, 1H NMR (400 MHz, DMSO-d 6) δ 8.23 (s, 1H), 8.13 (d, J = 8.1 Hz, 1H), 7.70 (d, J = 8.2 Hz, 1H), 7.11 (d, J = 7.2 Hz, 1H), 6.86 (d, J = 9.2 Hz, 2H), 6.66 (s, 1H), 5.48 (s, 2H), 4.95-4.86 (m, 1H), 3.44 (s, 3H), 3.28 (s, 2H), 3.26-3.22 (m, 2H), 2.82-2.75 (m, 1H), 2.62-2.53 (m, 2H), 2.05-1.96 (m, 2H), 1.88-1.82 (m, 2H), 1.50 (d, J= 7.0 Hz, 3H)。
Compound I-11: (R) -6- (1-acetylpiperidin-4-yl) -4- ((1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -2-methylphthalazin-1 (2H) -one
Figure 683455DEST_PATH_IMAGE030
Synthesis of Compound I-11 referring to Compound I-10, Compound I-11 was prepared as a white solid by using acetyl chloride instead of ethyl 2-bromoacetate.
Compound I-11(24 mg), MS: 488.2 (M + H)+。1H NMR (400 MHz, DMSO-d 6) δ 8.15-8.11 (m, 2H), 7.70 (d, J = 8.1 Hz, 1H), 7.05-7.00 (m, 1H), 6.85 (d, J = 6.7 Hz, 2H), 6.66 (s, 1H), 5.48 (s, 2H), 4.86-4.83 (m, 1H), 4.62-4.58 (m, 1H), 3.99-3.96 (m, 1H), 3.43 (s, 3H), 3.20-3.12 (M, 1H), 2.99-2.90 (m, 1H), 2.64-2.55 (m, 1H), 2.04 (s, 3H), 1.90-1.82 (m, 2H), 1.73-1.61 (m, 2H), 1.50 (d, J= 6.9 Hz, 3H)。
Compound I-12: (R) -4- ((1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -6- (1- (dimethylglycine) piperidin-4-yl) -2-methylphthalazin-1 (2H) -one
Figure 967544DEST_PATH_IMAGE031
Synthesis of Compound I-12 with reference to Compound I-11, Compound I-12 was prepared as a white solid by using dimethylglycinoyl chloride instead of acetyl chloride.
Compound I-12(26 mg), MS: 531.2 (M + H)+。1H NMR (400 MHz, DMSO-d 6) δ 8.16-8.10 (m, 2H), 7.70 (d, J = 8.3 Hz, 1H), 7.05-7.01 (m, 1H), 6.85 (d, J = 7.5 Hz, 2H), 6.66 (s, 1H), 5.48 (s, 2H), 4.90 (t, J = 7.0 Hz, 1H), 4.60-4.54 (m, 1H), 4.24-4.18 (m, 1H), 3.43 (s, 3H), 3.10 (s, 3H), 2.99-2.92 (m, 1H), 2.68-2.60 (m, 1H), 2.19 (s, 6H), 1.91-1.84 (m, 2H), 1.70-1.61 (m, 2H), 1.50 (d, J= 7.0 Hz, 3H)。
Compound I-13: (R) -4- ((1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -2-methyl-6- (1-methyl-6-oxo-1, 6-dihydropyridin-3-yl) phthalazin-1 (2H) -one
Figure 627195DEST_PATH_IMAGE032
Synthesis of Compound I-13 with reference to Compound I-7, Compound I-13 was prepared as a white solid by using 1-methyl-2-oxo-1, 2-dihydropyridine-5-boronic acid pinacol ester in place of 1-methyl-2-oxo-1, 2-dihydropyridine-4-boronic acid pinacol ester.
Compound I-13(17 mg), MS: 456.2 (M + H)+。1H NMR (400 MHz, DMSO-d 6) δ 8.39 (d, J = 9.2 Hz, 2H), 8.21 (d, J = 8.5 Hz, 1H), 8.11-8.01 (m, 2H), 7.11-7.07 (m, 1H), 6.93-6.88 (m, 2H), 6.71 (s, 1H), 6.63-6.58 (m, 1H), 4.95-4.92 (m, 1H), 3.56 (s, 3H), 3.44 (s, 3H), 1.53 (d, J = 6.9 Hz, 3H)。
Compound I-14: (R) -4- ((1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -6-isopropoxy-7-methoxy-2-methylphthalazin-1 (2H) -one
Figure 382662DEST_PATH_IMAGE033
Synthesis of Compound I-14 referring to Compound I-1, Compound I-14 was prepared as a white solid by using (R) -6-isopropoxy-7-methoxy-2-methyl-4- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) phthalazin-1 (2H) -one (intermediate A2-8) in place of 2-methyl-4- (((R) -1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) -6- (((S) -tetrahydrofuran-3-yl) oxy) phthalazin-1 (2H) -one (intermediate I-1-1).
Compound I-14(25 mg), MS: 451.1(M + H)+。1H NMR (400 MHz, DMSO-d 6 ) δ 7.70 (s, 1H), 7.57 (s, 1H), 6.94 (s, 1H), 6.90 (s, 2H), 6.74 (s, 1H), 4.95 (p, J = 6.1 Hz, 4H), 3.88 (s, 3H), 3.42 (s, 3H), 1.51 (d, J = 7.0 Hz, 3H), 1.35 (t, J= 5.7 Hz, 6H)。
Compound I-15: (R) -4- ((1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -6, 7-dimethoxy-2-methylphthalazin-1 (2H) -one
Figure 116263DEST_PATH_IMAGE034
Step 1 preparation of (R) -6, 7-dimethoxy-2-methyl-4- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) phthalazin-1 (2H) -one (intermediate I-15-1)
(R) -6-hydroxy-7-methoxy-2-methyl-4- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) phthalazin-1 (2H) -one (intermediate a2-9, 60 mg, 0.137 mmol) was dissolved in acetonitrile (5 ml). Slowly add K with 0 ℃ stirring2CO3(56.7 mg, 0.411 mmol) and methyl iodide (58.3 mg, 0.411 mmol), and after the addition was complete, the reaction mixture was stirred at 20 ℃ for an additional 12 hours, then quenched by the addition of ice water and extracted by dilution with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous dry sodium sulfate, then filtered and concentrated to give (R) -6, 7-dimethoxy-2-methyl-4- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) phthalazin-1 (2H) -one (62 mg, 100%) as a yellow solid. MS: 453.1 (M + H)+。
Step 2 preparation of (R) -4- ((1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -6, 7-dimethoxy-2-methylphthalazin-1 (2H) -one (Compound I-15)
Synthesis of Compound I-15 with reference to Compound I-1, Compound I-15 was prepared as a white solid by using ((R) -6, 7-dimethoxy-2-methyl-4- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) phthalazin-1 (2H) -one (intermediate I-15-1) in place of 2-methyl-4- (((R) -1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) -6- (((S) -tetrahydrofuran-3-yl) oxy) phthalazin-1 (2H) -one (intermediate I-1-1).
Compound I-15(15.68 mg), MS: 423.0 (M + H)+。1H NMR (400 MHz, DMSO-d 6 ) δ 7.68 (s, 1H), 7.56 (d, J = 1.9 Hz, 1H), 6.95 (d, J = 13.8 Hz, 3H), 6.76 (s, 1H), 4.93 (d, J = 7.3 Hz, 2H), 3.97 (s, 3H), 3.89 (d, J = 1.9 Hz, 3H), 3.42 (d, J = 1.9 Hz, 3H), 1.51 (d, J = 6.9 Hz, 3H)。
Compound I-16: (R) -6- (1-acetylpiperidin-4-yl) -4- ((1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -7-methoxy-2-methylphthalazin-1 (2H) -one
Figure 135034DEST_PATH_IMAGE035
Synthesis of Compound I-16 referring to Compound I-11, Compound I-16 was prepared as a white solid by using (R) -7-methoxy-2-methyl-4- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) -1-oxo-1, 2-dihydrophthalazin-6-yl trifluoromethanesulfonate (intermediate A2) in place of (R) -2-methyl-4- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) -1-oxo-1, 2-dihydrophthalazin-6-yl trifluoromethanesulfonate (intermediate A1).
Compound I-16 (17 mg), MS: 518.2 (M + H)+。1H NMR (400 MHz, DMSO-d 6 ) δ 8.04 (s, 1H), 7.60 (d, J = 1.8 Hz, 1H), 7.02 – 6.84 (m, 3H), 6.72 (s, 1H), 4.90 (s, 1H), 4.60 (d, J = 13.1 Hz, 1H), 3.94 (d, J = 1.8 Hz, 5H), 3.42 (s, 3H), 3.16 (t, J = 12.9 Hz, 3H), 2.03 (s, 3H), 1.86 – 1.58 (m, 5H), 1.49 (d, J = 7.0 Hz, 3H)。
Compound I-17 4- (((R) -1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -7-methoxy-6- (((3R, 3aR,6R,6aR) -6-methoxy hexahydrofuro [3,2-b ] furan-3-yl) oxy) -2-methylphthalazin-1 (2H) -one
Figure 344299DEST_PATH_IMAGE036
Synthesis of Compound I-17 with reference to Compound I-1, Compound I-17 was prepared as a white solid by using intermediate A2-9 in place of intermediate A1-7 and intermediate B5 in place of intermediate B2.
Compound I-17 (13 mg), MS: 551.2 (M + H)+。1H NMR (400 MHz, DMSO-d 6 ) δ 7.70 (s, 1H), 7.57 (s, 1H), 6.85-6.76 (m, 3H), 6.67 (s, 1H), 5.16-5.12 (m, 1H), 4.96-4.91 (m, 2H), 4.68-4.63 (m, 1H), 4.18-4.13 (m, 1H), 3.95-3.91 (m, 1H), 3.89 (s, 3H), 3.86-3.79 (m, 2H), 3.53-3.50 (m, 2H), 3.42 (s, 3H), 3.35 (s, 3H), 1.50 (d, J = 6.8 Hz, 3H)。
Compound I-18 (R) -4- ((1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -7-methoxy-2-methyl-6- (1-methyl-6-oxo-1, 6-dihydropyridin-3-yl) phthalazin-1 (2H) -one
Figure 778822DEST_PATH_IMAGE037
Synthesis of Compound I-18 with reference to Compound I-13, Compound I-18 was prepared as a white solid by using intermediate A2 in place of intermediate A1.
Compound I-18 (13 mg), MS: 500.2 (M + H)+。1H NMR (400 MHz, DMSO-d 6 ) δ 8.23 (s, 1H), 8.05 (s, 1H), 7.72 (d, J = 9.2 Hz, 1H), 7.67 (s, 1H), 7.17 -7.10 (m, 2H), 7.05 (s, 1H), 6.93 (s, 1H), 6.50 (d, J = 9.4 Hz, 1H), 4.94 (d, J = 7.4 Hz, 2H), 3.93 (s, 3H), 3.53 (s, 3H), 3.44 (s, 3H), 1.50 (d, J = 7.0 Hz, 3H)。
Compound I-19 (R) -4- ((1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -7-methoxy-2-methyl-6- (1-methyl-2-oxo-1, 2-dihydropyridin-4-yl) phthalazin-1 (2H) -one
Figure 694826DEST_PATH_IMAGE038
Synthesis of Compound I-19 with reference to Compound I-7, Compound I-19 was prepared as a white solid by using intermediate A2 in place of intermediate A1.
Compound I-19 (7 mg), MS: 500.2 (M + H)+。1H NMR (400 MHz, DMSO-d 6 ) δ 8.31 (s, 1H), 7.75 (d, J = 7.0 Hz, 1H), 7.71 (s, 1H), 7.08 (d, J = 6.8 Hz, 1H), 6.91 (d, J = 10.0 Hz, 2H), 6.72 (s, 1H), 6.63 (s, 1H), 6.45 (d, J = 6.9 Hz, 1H), 4.90 (s, 1H), 3.93 (s, 3H), 3.47 (s, 3H), 3.45 (s, 3H), 1.47 (d, J = 6.9 Hz, 3H)。
Compound I-20 (R) -1- ((1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -7-methoxy-3-methylpyridine [3,4-d ] pyridazin-4 (3H) -one
Figure 681236DEST_PATH_IMAGE039
Synthesis of Compound I-20 with reference to Compound I-2, Compound I-20 was prepared as a white solid by substituting intermediate A3-6 for intermediate A1-6.
Compound I-20 (17 mg), MS: 394.2 (M + H)+。
Compound I-21 (R) -1- ((1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -3-methyl-7- (1-methyl-2-oxo-1, 2-dihydropyridin-4-yl) pyridine [3,4-d ] pyridazin-4 (3H) -one
Figure 315480DEST_PATH_IMAGE040
Synthesis of Compound I-21 with reference to Compound I-7, Compound I-21 was prepared as a white solid by using intermediate A3 in place of intermediate A1.
Compound I-21 (27 mg), MS: 471.0 (M + H)+。1H NMR (400 MHz, DMSO-d 6 ) δ 9.44 (s, 1H), 8.80 (s, 1H), 7.90 (d, J = 7.1 Hz, 1H), 7.50 – 7.34 (m, 2H), 7.12 – 6.95 (m, 3H), 6.82 (s, 1H), 4.94 (t, J = 6.8 Hz, 1H), 3.50 (s, 3H), 3.45 (s, 3H), 1.54 (d, J = 6.9 Hz, 3H)。
Compound I-22 (R) -6- (1-acetyl-1, 2,3, 6-tetrahydropyridin-4-yl) -4- ((1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -7-methoxy-2-methylphthalazin-1 (2H) -one
Figure 553694DEST_PATH_IMAGE041
(R) -6- (1-acetyl-1, 2,3, 6-tetrahydropyridin-4-yl) -7-methoxy-2-methyl-4- ((1- (3-nitro-5- (trifluoromethyl) phenyl) ethyl) amino) phthalazin-1 (2H) -one (50 mg, 0.092 mmol) was dissolved in ethanol (5 ml) and water (0.5 ml). Iron powder (25.3 mg, 0.453 mmol) and NH were added with 20 degrees stirring4Cl (40.4 mg, 0.755 mmol), and after the addition was complete, the reaction mixture was stirred at 80 ℃ for 3 hours, then quenched by addition of ice water, filtered, and extracted by dilution with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous dry sodium sulfate, then filtered and concentrated to give the crude product, which was purified by pre-HPLC to give (R) -6- (1-acetyl-1, 2,3, 6-tetrahydropyridin-4-yl) -4- ((1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -7-methoxy-2-methylphthalazin-1 (2H) -one (20 mg) as a white solid. MS: 516.1 (M + H)+, 1H NMR (400 MHz, DMSO-d 6 ) δ 8.07 (s, 1H), 7.60 (d, J = 1.7 Hz, 1H), 7.00 (s, 1H), 6.91 (d, J = 13.7 Hz, 2H), 6.72 (s, 1H), 5.96 (s, 1H), 4.89 (d, J = 7.6 Hz, 1H), 3.91 (d, J = 1.8 Hz, 3H), 3.62 (s, 4H), 3.43 (d, J = 1.7 Hz, 3H), 2.08 – 2.03 (m, 3H), 1.47 (d, J = 6.9 Hz, 3H)。
Compound I-23 (R) -1- ((1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -3-methyl-7- (1-methyl-6-oxo-1, 6-dihydropyridin-3-yl) pyridine [3,4-d ] pyridazin-4 (3H) -one
Figure 386521DEST_PATH_IMAGE042
Synthesis of Compound I-23 with reference to Compound I-13, Compound I-23 was prepared as a white solid by using intermediate A3 in place of intermediate A1.
Compound I-23 (13 mg), MS: 471.1 (M + H)+。1H NMR (400 MHz, DMSO-d 6 ) δ 9.32 (s, 1H), 8.71 (d, J = 2.6 Hz, 1H), 8.45 (s, 1H), 8.29 (dd, J = 9.5, 2.6 Hz, 1H), 7.24 (d, J = 6.9 Hz, 1H), 6.91 (d, J = 6.9 Hz, 2H), 6.73 (s, 1H), 6.62 (d, J = 9.6 Hz, 1H), 4.97 – 4.88 (m, 1H), 3.58 (s, 3H), 3.44 (s, 3H), 1.54 (d, J = 7.0 Hz, 3H)。
Compound I-24 (R) -7- (1-acetyl-1, 2,3, 6-tetrahydropyridin-4-yl) -1- ((1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -3-methylpyridine [3,4-d ] pyridazin-4 (3H) -one
Figure 481516DEST_PATH_IMAGE043
Synthesis of Compound I-24 with reference to Compound I-22, Compound I-24 was prepared as a white solid by using intermediate A3 in place of intermediate A2.
Compound I-24 (33 mg), MS: 487.1 (M + H)+。1H NMR (400 MHz, DMSO-d 6 ) δ 9.32 (s, 1H), 8.27 (d, J = 10.5 Hz, 1H), 7.33 (d, J = 7.0 Hz, 1H), 7.02 (s, 1H), 6.88 (d, J = 7.1 Hz, 2H), 6.71 (s, 1H), 4.91 (t, J = 6.9 Hz, 1H), 4.28 (s, 1H), 4.23 (s, 1H), 3.43 (s, 3H), 2.70 (d, J = 40.4 Hz, 2H), 2.07 (d, J = 15.5 Hz, 3H), 1.51 (d, J = 6.9 Hz, 3H)。
Compound I-25 (R) -4- ((1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -7-fluoro-2-methyl-6- (1-methyl-6-oxo-1, 6-dihydropyridin-3-yl) phthalazin-1 (2H) -one
Figure 39274DEST_PATH_IMAGE044
Synthesis of Compound I-25 with reference to Compound I-13, Compound I-25 was prepared as a white solid by using intermediate A4 in place of intermediate A1.
Compound I-25 (8 mg), MS: 488.2 (M + H)+。1H NMR (400 MHz, DMSO-d 6 ) δ 8.43 (d, J = 7.1 Hz, 1H), 8.18 (d, J = 2.6 Hz, 1H), 7.93 (d, J = 10.8 Hz, 1H), 7.81 (d, J = 9.5 Hz, 1H), 7.11 (d, J = 7.0 Hz, 1H), 6.89 (d, J = 7.7 Hz, 2H), 6.71 (s, 1H), 6.59 (d, J = 9.4 Hz, 1H), 4.95-4.87 (m, 1H), 3.55 (s, 3H), 3.45 (s, 3H), 1.50 (d, J = 7.0 Hz, 3H)。
Compound I-26 (R) -6- (1-acetyl-1, 2,3, 6-tetrahydropyridin-4-yl) -4- ((1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -7-fluoro-2-methylphthalazin-1 (2H) -one
Figure 143496DEST_PATH_IMAGE045
Synthesis of Compound I-26 with reference to Compound I-24, Compound I-26 was prepared as a white solid by using intermediate A4 in place of intermediate A3.
Compound I-26 (7.3 mg), MS: 504.2 (M + H)+, 1H NMR (400 MHz, DMSO-d 6 ) δ 8.30 (d, J = 6.8 Hz, 1H), 7.84 (d, J = 10.9 Hz, 1H), 7.13 (d, J = 7.0 Hz, 1H), 6.89 (d, J = 8.8 Hz, 2H), 6.71 (s, 1H), 6.21 (s, 1H), 4.93-4.88 (m, 1H), 4.22-4.15 (m, 2H), 3.69-3.64 (m, 2H), 3.43 (s, 3H), 2.66-2.52 (m, 2H), 2.07 (d, J = 13.3 Hz, 3H), 1.49 (d, J = 6.9 Hz, 3H)。
Compound I-27 (R) -6- (1-acetyl-2, 5-dihydro-1H-pyrrol-3-yl) -4- (1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -7-fluoro-2-methylphthalazin-1 (2H) -one
Figure 830830DEST_PATH_IMAGE046
Synthesis of compound I-27 referring to compound I-26, a white solid compound I-27 was prepared by using tert-butyl 3- (4,4,5, 5-tetramethyl-1, 3, 2-dioxaborolan-2-yl) -2, 5-dihydro-1H-pyrrole-1-carboxylate instead of N-tert-butoxycarbonyl-1, 2,5, 6-tetrahydropyridine-4-boronic acid pinacol ester.
Compound I-27 (17.3 mg), MS: 490.2 (M + H)+, 1H NMR (400 MHz, DMSO-d 6 ) δ 8.24 (d, J = 6.8 Hz, 1H), 7.90 (d, J = 11.7 Hz, 1H), 7.34 (d, J = 7.1 Hz, 1H), 6.90 (d, J = 6.8 Hz, 2H), 6.73 (d, J = 10.0 Hz, 2H), 4.97-4.90 (m, 1H), 4.68-4.62 (m, 2H), 4.56-4.52 (m, 2H), 2.06 (d, J = 18.9 Hz, 3H), 1.52 (d, J = 7.1 Hz, 3H)。
Compound I-28 (R) -6- (1-acetylpiperidin-4-yl) -4- ((1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -7-fluoro-2-methylphthalazin-1 (2H) -one
Figure 362305DEST_PATH_IMAGE047
Synthesis of Compound I-28 with reference to Compound I-16, Compound I-28 was prepared as a white solid by using intermediate I-26-3 in place of intermediate I-16-3.
Compound I-28 (6.8 mg), MS: 506.2 (M + H)+。1H NMR (400 MHz, DMSO-d 6 ) δ 8.24 (d, J = 6.5 Hz, 1H), 7.82 (d, J = 10.5 Hz, 1H), 7.11 (s, 1H), 6.88 (d, J = 8.9 Hz, 2H), 6.71 (s, 1H), 4.93-4.86 (m, 1H), 4.63-4.58 (m, 1H), 4.03-3.98 (m, 1H), 3.42 (s, 3H), 3.26-3.16 (m, 2H), 2.70-2.60 (m, 1H), 2.04 (s, 3H), 1.89-1.66 (m, 4H), 1.50 (d, J = 7.0 Hz, 3H)。
Compound I-29: 6- (1-acetylpyrrolidin-3-yl) -4- (((R) -1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -7-fluoro-2-methylphthalazin-1 (2H) -one
Figure 377666DEST_PATH_IMAGE048
Synthesis of Compound I-29 referring to Compound I-16, Compound I-29 was prepared as a white solid by using intermediate I-27-3 in place of intermediate I-16-3.
Compound I-29 (18 mg), MS: 492.2 (M + H)+。1H NMR (400 MHz, DMSO-d 6 ) δ 8.38-8.29 (m, 1H), 7.87-7.82 (m, 1H), 7.18-7.10 (m, 1H), 6.88-6.85 (m, 2H), 6.69 (s, 1H), 4.93-4.90 (m, 1H), 3.98-3.91 (m, 2H), 3.56-3.50 (m, 2H), 3.44 (s, 3H), 2.43- 2.10 (m, 3H), 1.98 (d, J = 5.8 Hz, 3H), 1.50 (d, J = 6.9 Hz, 3H)。
Compound I-30 (R) -4- ((1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -7-fluoro-2-methyl-6- (1-methyl-2-oxo-1, 2-dihydropyridin-4-yl) phthalazin-1 (2H) -one
Figure 347896DEST_PATH_IMAGE049
Synthesis of Compound I-30 referring to Compound I-7, Compound I-30 was prepared as a white solid by using intermediate A4 in place of intermediate A1.
Compound I-30 (17 mg), MS: 488.0 (M + H)+。1H NMR (400 MHz, DMSO-d 6 ) δ 8.55 (d, J = 6.8 Hz, 1H), 7.96 (d, J = 10.6 Hz, 1H), 7.86 (d, J = 7.0 Hz, 1H), 7.24 (d, J = 6.8 Hz, 1H), 6.91 (d, J = 8.5 Hz, 2H), 6.77- 6.72 (m, 2H), 6.53 (d, J = 7.0 Hz, 1H), 4.96 – 4.87 (m, 2H), 3.45 (s, 3H), 1.49 (d, J = 7.0 Hz, 3H)。
Compound I-31 (R) -7- (1-acetylpiperidin-4-yl) -1- ((1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -3-methylpyridine [3,4-d ] pyridazin-4 (3H) -one
Synthesis of Compound I-31 referring to Compound I-16, Compound I-31 was prepared as a white solid by using intermediate I-24-3 in place of intermediate I-16-3.
Compound I-31 (21 mg), MS: 489.1 (M + H)+。1H NMR (400 MHz, DMSO-d 6 ) δ 9.31 (s, 1H), 8.09 (s, 1H), 7.24 (d, J = 7.0 Hz, 1H), 6.89 (d, J = 11.5 Hz, 2H), 6.72 (s, 1H), 4.88 (t, J = 6.9 Hz, 1H), 4.56 (d, J = 13.0 Hz, 1H), 3.96 (d, J = 13.6 Hz, 1H), 3.43 (s, 3H), 3.16 (m, 2H), 2.66 (t, J = 12.7 Hz, 1H), 2.04 (s, 6H), 1.78 – 1.60 (m, 2H), 1.50 (d, J = 6.9 Hz, 4H)。
Compound I-32- (1-acetylpyrrolidin-3-yl) -4- ((R) -1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -2, 7-dimethylphthalazin-1 (2H) -one
Figure 92998DEST_PATH_IMAGE050
Synthesis of Compound I-32 with reference to Compound I-29, Compound I-32 was prepared as a white solid by using intermediate A5 in place of intermediate A4.
Compound I-32 (19.3 mg), MS: 488.2 (M + H)+,1H NMR (400 MHz, DMSO-d 6 ) δ 8.22-8.14 (m, 1H), 8.00 (d, J = 4.2 Hz, 1H), 7.10-6.93 (m, 1H), 6.87 (d, J = 9.4 Hz, 2H), 6.69 (s, 1H), 4.95-4.87 (m, 1H), 3.98-3.85 (m, 1H), 3.78-3.72 (m, 1H), 3.64-3.55 (m, 1H), 3.45-3.31 (m, 5H), 2.52 (d, J = 8.9 Hz, 3H), 2.37-2.13 (m, 2H), 2.00-1.96 (m, 3H), 1.50 (dd, J = 7.3, 2.7 Hz, 3H).
Compound I-33 (R) -6- (1-acetyl-1, 2,3, 6-tetrahydropyridin-4-yl) -4- ((1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -2, 7-dimethylphthalazin-1 (2H) -one
Figure 733058DEST_PATH_IMAGE052
Synthesis of Compound I-33 referring to Compound I-26, Compound I-33 was prepared as a white solid by using intermediate A5 in place of intermediate A4.
Compound I-33 (7.9 mg), MS: 500.2 (M + H)+, 1H NMR (400 MHz, DMSO-d 6 ) δ 8.02 (s, 2H), 7.02 (s, 1H), 6.92 (d, J = 16.0 Hz, 2H), 6.73 (s, 1H), 5.74 (s, 1H), 4.89 (s, 1H), 4.16-4.11 (m, 2 H), 3.72 - 3.66 (m, 2H), 3.42 (s, 3H), 2.53 (s, 1H), 2.45 (s, 1H), 2.41 (s, 3H), 2.09 – 2.05 (m, 4H), 1.47 (d, J = 7.0 Hz, 3H).
Compound I-34 (R) -6- (1-acetylpiperidin-4-yl) -4- ((1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -2, 7-dimethylphthalazin-1 (2H) -one
Figure 563610DEST_PATH_IMAGE053
Synthesis of Compound I-34 referring to Compound I-31, Compound I-34 was prepared as a white solid by substituting intermediate I-33-3 for intermediate I-24-3.
Compound I-34 (9 mg), MS: 502.2 (M + H)+, 1H NMR (400 MHz, DMSO-d 6 ) δ 8.03 (d, J = 2.5 Hz, 1H), 7.98 (s, 1H), 7.06 (d, J = 7.7 Hz, 1H), 6.89 (d, J = 11.9 Hz, 2H), 6.71 (s, 1H), 4.95 – 4.84 (m, 1H), 4.62 (d, J = 13.1 Hz, 1H), 3.98 (d, J = 13.7 Hz, 2H), 3.40 (s, 3H), 3.23 – 3.12 (m, 2H), 2.70 – 2.59 (m, 1H), 2.52 (d, J = 10.8 Hz, 3H), 2.07 (s, 1H), 2.05 (s, 3H), 1.74 (s, 3H), 1.50 (d, J = 7.1 Hz, 3H).
Compound I-35 (R) -1- ((1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -7- (1, 1-dioxothiomorpholine) -3-methylpyridine [3,4-d ] pyridazin-4 (3H) -one
Figure 71952DEST_PATH_IMAGE054
Step 1: preparation of intermediate I-35-1
Intermediate A3 (705 mg, 1.304 mmol), 1, 1-dioxo-4-thiomorpholine (353 mg, 2.61 mmol) and K2CO3(541 mg, 3.91 mmol) of DMF (5 mL) the reaction mixture was stirred at 100 ℃ for an additional 11 hours, then quenched by addition of ice water and extracted by dilution with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous dry sodium sulfate, then filtered and concentrated to give the crude product, which was purified by silica gel column chromatography to give intermediate I-35-1 (460 mg, 67.1%) as a yellow solid. MS: 527.2 (M + H)+。
Step 2: preparation of (R) -1- ((1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -7- (1, 1-dioxothiomorpholine) -3-methylpyridine [3,4-d ] pyridazin-4 (3H) -one (Compound I-35)
Synthesis of Compound I-35 referring to Compound I-26, Compound I-35 was prepared as a white solid by substituting intermediate I-35-1 for intermediate I-26-3.
Compound I-35 (17.9 mg), MS: 497.2 (M + H)+, 1H NMR (400 MHz, DMSO-d 6 ) δ 8.96 (s,1H),7.44 (s, 1H), 7.02 (d, J = 7.0 Hz, 1H), 6.88 (d, J = 8.1 Hz, 2H), 6.73 (s, 1H), 4.90 (t, J = 6.9 Hz, 1H), 4.24 (s, 2H), 3.36-3.33 (m, 4H), 3.25-3.22 (m, 5H), 3.18 (s, 1H), 1.50 (d, J = 6.9 Hz, 3H).
Compound I-36 (R) -1- ((1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -3-methyl-7-morpholinopyridin [3,4-d ] pyridazin-4 (3H) -one
Figure 343665DEST_PATH_IMAGE055
Synthesis of Compound I-36 reference compound I-35 was prepared by using morpholine instead of 1, 1-dioxo-4-thiomorpholine to give compound I-36 as a white solid. MS: 497.2 (M + H)+。
Compound I-37 (R) -1- ((1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -3-methyl-7- (4- (1-methyl-1H-pyrazol-4-yl) piperazin-1-yl) pyrido [3,4-d ] pyridazin-4 (3H) -one
Figure 216943DEST_PATH_IMAGE056
Synthesis of Compound I-37 referring to Compound I-35, Compound I-37 was prepared as a white solid by using 1- (1-methyl-1H-pyrazol-4-yl) piperazine instead of 1, 1-dioxo-4-thiomorpholine.
Compound I-37 (11.1 mg), MS: 528.2 (M + H)+, 1H NMR (400 MHz, DMSO-d 6 ) δ 8.93 (s,1H), 7.38 (s, 1H), 7.33 (s, 1H), 7.26 (s, 1H), 7.09 – 7.00 (m, 1H), 6.89 (d, J = 9.2 Hz, 2H), 6.74 (s, 1H), 4.94 – 4.81 (m, 2H), 3.85 (t, J = 4.8 Hz, 4H), 3.73 (d, J = 1.4 Hz, 3H), 3.34 (s, 3H), 3.01 (t, J = 5.0 Hz, 4H), 1.50 (d, J = 6.9 Hz, 3H).
Compound I-38 (R) -1- ((1- (3-amino-5- (trifluoromethyl) phenyl) ethyl) amino) -7- (4- (cyclopropanecarbonyl) piperazin-1-yl) -3-methylpyridin [3,4-d ] pyridazin-4 (3H) -one
Figure 597108DEST_PATH_IMAGE057
Synthesis of Compound I-38 with reference to Compound I-35, Compound I-38 was prepared as a white solid by using cyclopropyl (piperazin-1-yl) methanone instead of 1, 1-dioxo-4-thiomorpholine.
Compound I-38 (19 mg), MS: 516.2 (M + H)+, 1H NMR (400 MHz, DMSO-d 6 ) δ 8.92 (s,1H)，7.37 (s, 1H), 7.35 – 7.32 (m, 1H), 7.25 – 7.20 (m, 2H), 7.11 – 7.10 (m, 1H), 4.93 (d, J = 7.2 Hz, 2H), 3.84 (s, 4H), 3.74 - 3.64(m, 4H), 3.33 (s, 3H), 2.06 - 2.01 (m, 1H), 1.54 (d, J = 7.0 Hz, 3H), 0.78 – 0.71 (m, 4H).
Evaluation of drug efficacy at cellular level
This experiment was conducted to verify the proliferation inhibitory effect of the compound of the present invention on NCI-H358 human non-small cell lung cancer cells activated by RAS signaling pathway.
Main experimental materials and instruments:
RPMI 1640 Cell culture medium + L-Glutamine (Invitrogen), fetal bovine serum (Corning), dimethyl sulfoxide DMSO (Sigma), penicillin streptomycin (Gibico), Du's phosphate buffer DPBS (Corning), pancreatin TryPLETM Express (Gibco), Collagen Type I Rat Tail (Corning), 96-well U-shaped substrate culture plate (ThermoFisher), Cell titer Glo (Promega), TC-20 Cell counter (Bio-rad), multifunctional microplate reader Bio Synergy NEO2 (Bio-tek).
Human non-small cell lung carcinoma cells NCI-H358 (ATCC) were cultured in RPMI 1640 medium of 10% fetal bovine serum, 100U penicillin and 100. mu.g/mL streptomycin.
The experimental steps are as follows: compounds were diluted in DMSO as a semilog dilution (i.e., 3.16 fold dilution), with 7 concentration gradients (giving an initial concentration of compound of 6mM) and DMSO control wells. Then, the diluted compound was diluted 100-fold with the culture medium, and 80. mu.L of the diluted compound was added to a 96-well U-plate.
Then, NCI-H358 cells in the logarithmic growth phase were digested with pancreatin and subjected to cell counting using a Bio-Rad TC20 cell counter, the cells were diluted with a culture medium containing Collagen I (1:125 dilution), 80. mu.L of the cell suspension was inoculated into the above 96-well plate containing 80. mu.L of the diluted compound so that the maximum concentration of the compound was 30. mu.M and the cell density was about 2400 cells per wellAnd (4) cells. Cells were spheronized by centrifugation at 250g for 5 minutes. At 37 ℃ 5% CO2Culturing in an incubator for 7 days. The cell culture plate was removed and allowed to equilibrate at room temperature for 30 minutes. Add 80. mu.l of Celltiter Glo reagent to each well. Cells were lysed thoroughly by shaking on a shaker at 350rpm for 20 minutes. Measuring chemiluminescence values under different cell numbers by using an enzyme-labeling instrument, calculating the cytotoxicity percentage of a compound to be measured according to DMSO control and culture medium control, fitting (fit) half inhibition concentration IC (integrated circuit) by using GraphPad Prism according to a four-parameter method by using the concentration of the compound as a horizontal coordinate and the cytotoxicity percentage (cytoxicity%) of the compound as a vertical coordinate50。
Figure DEST_PATH_IMAGE058
All documents referred to herein are incorporated by reference into this application as if each were individually incorporated by reference. Furthermore, it should be understood that various changes and modifications of the present invention can be made by those skilled in the art after reading the above teachings of the present invention, and these equivalents also fall within the scope of the present invention as defined by the appended claims.

Claims (10)

1. A compound shown as a formula I or a pharmaceutically acceptable salt thereof,
Figure 919823DEST_PATH_IMAGE001
x is selected from N or CRa；
RaSelected from the group consisting of: hydrogen, halogen, C1-6Alkyl or C1-6An alkoxy group;
l is selected from the group consisting of: - (CH)2)m-or-O-, wherein m is 0 or 1;
R1selected from the group consisting of: c1-6Alkyl radical, C3-6Cycloalkyl, 4-8 membered saturated or partially unsaturated heterocyclyl, phenyl, 5-6 membered heteroaryl;
and saidR1Optionally substituted by one or more RbSubstituted by substituent, said RbSelected from the group consisting of: hydrogen, halogen, one or more Rb1Substituted C1-3Alkyl radical, C1-3Alkoxy, -C (= O) -Rb2Or one or more Rb3A substituted 5-6 membered heteroaryl, wherein:
Rb1selected from the group consisting of: hydrogen, halogen, amino or carboxyl;
Rb2selected from the group consisting of: hydrogen, C1-6Alkyl radical, C1-6Alkoxy radical, C3-6Cycloalkyl or- (CH)2)q-N(Rb1ARb1B)；
Rb3Selected from hydrogen, halogen or C1-3An alkyl group;
q is 0, 1,2 or 3;
Rb1A、Rb1Bis hydrogen or C1-3An alkyl group;
one or more methylene groups as a ring skeleton on each of the above-mentioned cycloalkyl groups and heterocyclic groups may each independently be substituted by a carbonyl group or S (= O)2Replacement;
each of said heterocyclyl or heteroaryl groups independently containing from 1 to 3 heteroatoms selected from N, O, S, the number of heteroatoms being from 1 to 3;
R2is optionally selected from C1-3Alkyl radical, C1-3Haloalkyl or cyclopropane;
R3can be the same or different and are each independently selected from halogen, amino, C1-3Alkyl or C1-3A haloalkyl group;
n is 1,2 or 3.
2. The compound of formula I of claim 1, or a pharmaceutically acceptable salt thereof, wherein the compound of formula I has the structure shown in formula II:
Figure 200763DEST_PATH_IMAGE002
。
3. the compound of formula I of claim 1, or a pharmaceutically acceptable salt thereof, wherein R isaSelected from the group consisting of: hydrogen, halogen, C1-4Alkyl or C1-4An alkoxy group.
4. A compound of formula I according to claim 1, or a pharmaceutically acceptable salt thereof, wherein L is selected from the group consisting of: - (CH)2)m-or-O-, wherein m is 0.
5. The compound of formula I of claim 1, or a pharmaceutically acceptable salt thereof, wherein R is2Is optionally selected from C1-3An alkyl group.
6. The compound of formula I of claim 1, or a pharmaceutically acceptable salt thereof, wherein R is1Selected from the group consisting of: c1-4Alkyl radical, C3-6Cycloalkyl, 5-8 membered saturated or partially unsaturated heterocyclyl; wherein, R is1Optionally substituted by one or more RbSubstituted by substituent, said RbSelected from the group consisting of: hydrogen, halogen, one or more Rb1Substituted C1-3Alkyl radical, C1-3Alkoxy, -C (= O) -Rb2Or one or more Rb3A substituted 5-6 membered heteroaryl, wherein:
Rb1selected from the group consisting of: hydrogen, halogen, or carboxyl;
Rb2selected from the group consisting of: hydrogen, C1-4Alkyl radical, C1-4Alkoxy radical, C3-6Cycloalkyl or- (CH)2)q-N(Rb1ARb1B)；
Rb3Selected from hydrogen, halogen or C1-3An alkyl group;
q is 0 or 1;
Rb1A、Rb1Bis hydrogen or C1-3An alkyl group.
7. The compound of formula I of claim 6, or a pharmaceutically acceptable salt thereof, wherein R is1Selected from the group consisting of: c1-4Alkyl, or a group selected from:
Figure 905414DEST_PATH_IMAGE003
said R1Optionally substituted by one or more RbSubstituent, said RbAs defined in claim 6.
8. The compound of formula I of claim 1, or a pharmaceutically acceptable salt thereof, selected from the group consisting of:
Figure 496188DEST_PATH_IMAGE004
Figure 401827DEST_PATH_IMAGE005
Figure 294697DEST_PATH_IMAGE006
。
9. a pharmaceutical composition, comprising: a compound of formula I according to claim 1 or a pharmaceutically acceptable salt thereof, together with one or more pharmaceutically acceptable carriers, excipients, adjuvants and/or diluents.
10. The use of a compound of formula I as claimed in claim 1, or a pharmaceutically acceptable salt thereof, for the preparation of a pharmaceutical composition for the treatment or prevention of a disease associated with SOS1 or modulated by SOS 1.
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Application publication date: 20220614