WO2024107096A1 - Treprostinil formulations - Google Patents

Abstract

This disclosure provides treprostinil compositions and its use thereof.

Description

TREPROSTINIL FORMULATIONS

FIELD OF THE DISCLOSURE

[0001] The present disclosure relates to formulation precursors (pre-formulations) for the in situ generation of compositions for the controlled release of treprostinil, and methods of treatment with such formulations. In particular, the disclosure relates to pre-formulations of amphiphilic components and treprostinil or a salt thereof, which undergo phase transition upon exposure to aqueous fluids, such as body fluids, thereby forming a controlled release composition.

BACKGROUND

[0002] Many bioactive agents including pharmaceuticals, nutrients, vitamins and so forth have a “functional window.” That is to say that there is a range of concentrations over which these agents can be observed to provide some biological effect. Where the concentration in the appropriate part of the body (e.g., locally or as demonstrated by serum concentration) falls below a certain level, no beneficial effect can be attributed to the agent. Similarly, there is generally an upper concentration level above which no further benefit is derived by increasing the concentration. In some cases increasing the concentration above a particular level results in undesirable or even dangerous effects.

[0003] Some bioactive agents have a long biological half-life and/or a wide functional window and thus may be administered occasionally, maintaining a functional biological concentration over a substantial period of time (e.g., 6 hours to several days). In other cases the rate of clearance is high and/or the functional window is narrow and thus to maintain a biological concentration within this window regular (or even continuous) doses of a small amount are required. This can be particularly difficult where non-oral routes of administration (e.g., parenteral administration) are desirable or necessary, since self-administration may be difficult and thus cause inconvenience and/or poor compliance. In such cases it would be advantageous for a single administration to provide active agent at a therapeutic level over the whole period during which activity is needed.

[0004] One particular class of active agents having a high rate of clearance and short half-life are prostacyclin and its analogues. Prostacyclin is an endogenous member of the eicosanoid family and is involved in several processes including platelet activation, vasodilation, and blood pressure regulation. Prostacyclin is also known as epoprostenol when referring to synthetically derived material, and the terms are used interchangeably herein.

[0005] Epoprostenol was approved for the treatment of pulmonary arterial hypertension (PAH) by the FDA in 1995. PAH is potentially fatal condition characterized by a mean pulmonary artery pressure (mPAP) of >25 mmHg, with normal pulmonary artery wedge pressure (PAWP) (<15 mmHg). However, as epoprostenol itself has an in vivo half-life of less than one minute, it requires continual administration, typically through a central venous catheter. Epoprostenol sodium for intravenous therapy is marketed as Flolan® (GlaxoSmithKline). Since 2008 a room-temperature stable formulation of epoprostenol (Veletri®, Actelion Pharmaceuticals) has also been available. An estimated 100,000 to 200,000 individuals are believed to be affected by PAH worldwide.

[0006] Several prostacyclin analogues with longer half-lives are known, including treprostinil. Treprostinil was approved by the FDA in 2002 and has a plasma half-life of 2.9 to 4.6 hours. Despite the longer half-life compared with epoprostenol, continual i.v. infusion or regular s.c. administration of treprostinil is still generally necessary. IV therapy requires surgical insertion of a central venous catheter, carries the risk of infection and thrombosis and is naturally uncomfortable for the patient. Epoprostenol can also be administered through inhalation or oral routes. However, these routes provide a lower cumulative dose of epoprostenol than the IV route. They may thus not be suitable for all patients.

[0007] Remodulin® (United Therapeutics Corporation) is a formulation of treprostinil designed for IV or continuous s.c. injection. Continuous s.c. injection is achieved by means of a microinfusion pump. Although this addresses some of the issues associated with bulky pump equipment, it is still not ideal and furthermore it is recommended that patients have immediate access to a backup infusion pump.

[0008] Although regular s.c. administration somewhat addresses the disadvantages of i.v. or continuous s.c. administration, oral or inhalation routes, administration-site pain is a significant obstacle in the majority of patients (experienced by 85% of patients) and is responsible for almost all withdrawals from treprostinil due to adverse events (a total of 23% of the long-term study population). This has hereto been managed, to the extent possible, by appropriate site selection. Site pain peaks in the first few days after a site change, and use of a single site for 4 weeks or more can be helpful and safe in some cases. [0009] There is an evident need for a preparation of treprostinil which is stable to storage, which can be administered without the need for continuous administration though a central venous catheter or by continuous s.c. administration, which is not susceptible to the risk of mechanical failure, and/or which can be administered less frequently whilst causing less site pain than existing s.c. formulations. The present disclosure addresses some or all of these deficiencies.

[00010] Patients undergoing treatment for PAH typically require a therapeutic dose to be maintained for a considerable period and typically require ongoing treatment for many months or years. Thus a depot system allowing loading and controlled release of a larger dose over a longer period would offer a considerable advantage over conventional delivery systems.

[00011] In this regard, polymer delivery systems containing treprostinil have been developed, such as TransCon Treprostinil (Ascendis Pharma) which has undergone Phase 1 clinical trials. TransCon Treprostinil is designed as a once-daily self-administered s.c. injection of treprostinil and is based on a polymer delivery system, especially a poly(oxazoline) or PEG-based polymer. TransCon Treprostinil is intended to offer the same efficacy as continuously-infused prostacyclin analogues, but with a safer and more convenient route of administration with reduce site reaction and bloodstream infection risks associated with current parenteral administration routes.

[00012] The poly-lactate, poly-glycolate, and poly-lactate-co-glycolate polymers typically used for degrading slow-release formulations are also the cause of some irritation in at least some patients. In particular, these polymers typically contain a certain proportion of acidic impurities such as lactic and glycolic acid, which will irritate the injection site on administration. When the polymer then breaks down, lactic acid and glycolic acid are the degradation products so that further irritation is caused.

[00013] Despite the potential advantages offered by TransCon Treprostinil in terms of patient comfort and somewhat less frequent (once daily) administration, even if a polymer such as a PEG is used which is not broken down into acidic impurities, polymer systems tend to be of high viscosity and consequently require injection through a wide needle and/or provide only a fairly short duration product. PEG-grafting to treprostinil typically increases the biological lifetime but may interfere with binding and cannot currently provide a product that will remain active for several days between injection. As a result of the combined effects of wide-needle administration and/or irritant contents, discomfort at the site of administration and the formation of connective scar tissue are often greater than desirable. This is increased in the case of the proposed treprostinil formulation since injection is at least daily, rather than weekly or longer periodicity. As a result, over a long treatment duration, either multiple irritant administrations must be made at a small number of sites, or a large number of sites utilised, with resultant widespread discomfort for the subject.

[00014] Evidently, it would be an advantage to provide a system of low viscosity, such as a homogeneous solution, dispersion of fine particles, or L2 phase, which could be administered easily through a narrow needle, thus decreasing the discomfort of the patient during the procedure and causing less site pain. This ease of administration is particularly significant where patients will be on a self-administration regime and may already be self-administering several times each day, as is the case with several existing treprostinil treatments. Providing a sustained formulation with a duration of a few days, but which is sufficiently complex to administer that it requires treatment by a healthcare professional will not be an advantage to all patients over twice-daily or daily selfadministration, and is likely to be more costly. Providing a formulation which gives sufficiently long duration to justify a visit to a health professional for administration and/or a preparation which can be self-administered easily would be a significant advantage. Reducing preparation time of health-care professionals or patients prior to the actual administration is also an important issue.

[00015] From a drug delivery point of view, polymer depot compositions also generally have the disadvantage of accepting only relatively low drug loads and having a “burst/lag” release profile. The nature of the polymeric matrix, especially when applied as a solution or pre-polymer, causes an initial burst of drug release when the composition is first administered. This is followed by a period of low release, while the degradation of the matrix begins, followed finally by an increase in the release rate to the desired sustained profile. This burst/lag release profile can cause the in vivo concentration of active agent to burst above the functional window immediately following administration, and then drop back through the bottom of the functional window during the lag period before reaching a sustained functional concentration for a period of time. Evidently, from a functional and toxicological point of view this burst/lag release profile is undesirable and could be dangerous. It may also limit the equilibrium concentration which can be provided due to the danger of adverse effects at the “peak” point. The presence of a lag phase may furthermore require supplementary dosing with repeat injections during the start-up period of depot treatment in order to maintain a therapeutic dose while the concentrations of active provided from the depot are sub -functional.

[00016] Controlled-release formulations are typically generated from bio-compatible polymers in the form of, for example, implants or injectable beads. Polymer microsphere formulations must generally be administered by means of a sizable needle, typically of 20-gauge or wider. This is necessary as a result of the nature of the polymeric dosing systems used, which are typically polymer suspensions. It would be an advantage to provide a system of low viscosity, such as a homogeneous solution, dispersion of fine particles, or L2 phase, which could be administered easily through a narrow needle, thus decreasing the discomfort of the patient during the procedure. Ease of administration is particularly significant when patients will be self-administering but also reduces the burden on healthcare professionals when they are conducting the administration.

[00017] The manufacture of PLGA microbeads and suspensions is additionally a considerable difficulty with certain existing depot systems. In particular, since the beads are particulate they cannot generally be sterile-filtered and furthermore, since the PLGA copolymer melts at elevated temperature, they cannot be heat-treated for sterility. As a result, the complex manufacturing process must be conducted aseptically.

[00018] Further issues with biodegradable polymer microspheres include complex reconstitution prior to injection and limited storage stability, due both to aggregation and degradation of the delivery system and/or active.

[00019] A lipid-based, slow-release composition has been described for certain peptides. For example, W02006/131730 discloses a lipid depot system for GLP-1 and analogues thereof. This is a highly effective formulation, but the concentration of active agent which can be included in the formulation is limited by its solubility. Evidently, a higher concentration of active agent allows for the possibility of longer duration depot products, products maintaining a higher systemic concentration, and products having a smaller injection volume, all of which factors are of considerable advantage under appropriate circumstances. It would thus be of considerable value to establish a way by which higher concentrations of active agents could be included in a lipid- based depot formulation and to identify combinations of active agent and delivery system which are particularly effective from the point of view of loading, stability, manufacturing, and/or controlled release. [00020] The present inventors have now established that by providing a pre-formulation comprising at least one neutral mono-, di-, or triacyl lipid and/or a tocopherol, optionally at least one phospholipid, at least one biocompatible organic mono-alcoholic solvent, and treprostinil or a salt thereof in a low viscosity phase, such as molecular solution or L2 (reversed micellar) phase, a pre-formulation may be generated addressing many of the shortfalls of known treprostinil formulations, and which may be applied to provide a controlled release of treprostinil or a salt thereof. By use of specific components in carefully selected ratios, a depot formulation can be generated having a combination of properties exceeding the performance of, and providing an advantage over, known treprostinil compositions such as Remodulin® or TransCon treprostinil. [00021] In particular, the pre-formulation shows a highly advantageous release profile, is easy to manufacture, may be sterile-filtered, has low viscosity (allowing easy and less painful administration typically through a narrow needle), allows a high level of treprostinil to be incorporated (thus potentially allowing a smaller amount of composition and/or treprostinil to be used), requires shallow injection, and/or forms a desired non-lamellar depot composition in vivo having a “low-burst” release profile. The compositions are also formed from materials that are non-toxic, biotolerable, and biodegradable, which can be administered by single i.m., or s.c. injection rather than central venous catheter or continuous s.c. injection, and are suitable for selfadministration. The pre-formulation may additionally have a very low level of irritation on injection and in preferred cases causes no irritation at the injection site (including transient irritation). The pre-formulations may be administered less frequently than even proposed “slow release” formulations, resulting in better compliance from the patient and/or less irritation due to repeated frequent administrations.

[00022] Advantages of the compositions of the present disclosure over polymer formulations, such as PLGA microspheres, include the ease of manufacture (including sterilization), handling and use properties combined with low initial release (“low-burst profile”) of treprostinil. This may be defined such that the area under a plasma concentration against time the curve during the first 24 hours of a one-week dosing period is less than 50% of the area under the curve for the entire curve (measured or extrapolated from time 0 to infinity or from time 0 to the last sampling time point), e.g., less than 40% and most preferable less than 30%. Furthermore, it may be defined such that the maximum plasma concentration of treprostinil in vivo following injection of the pre- formulation (Cmax) is no more than 10 times, preferably no more than 8 times and most preferably no more than 5 times the average plasma concentration during the therapeutic period (Cave) (i.e., Cmax/Cave < 10, preferably < 8, e.g., < 5).

SUMMARY

[00023] The present disclosure provides a pharmaceutical formulation comprising an appropriate combination of lipid excipients, organic alcoholic solvent, and treprostinil or a salt thereof and certain optional components, that can be used as a depot-precursor formulation (referred to herein for brevity as a pre-formulation) to address one or more of the needs described above. The inventors have established that by optimising these components, depot compositions of treprostinil, and corresponding precursor formulations with a highly advantageous combination of properties can be generated.

[00024] A first embodiment provides a pre-formulation comprising:

[00025] a) at least one of a mono-, di-, or tri-acyl lipid and/or a tocopherol;

[00026] b) optionally at least one phospholipid;

[00027] c) at least one biocompatible, organic solvent; and

[00028] d) treprostinil or a salt thereof;

[00029] wherein the pre-formulation optionally but preferably forms, or is capable of forming, at least one liquid crystalline phase structure upon contact with excess aqueous fluid.

[00030] In another embodiment, the prostacyclin analogue (free acid) is present at a level of 0.1 to 10 wt.% of the pre-formulation, preferably 0.2 to 6 wt.%. In an embodiment, the prostacyclin analogue (free acid) is present at a level such as 0.2 to 5 wt.%, 0.5 to 5 wt.%, especially 0.2 to 4 wt.% or 0.75 to 4 wt.%. The term “free acid” as used herein is to be interpreted as neutral molecule (e.g., neutral ester).

[00031] In another embodiment, component d) comprises or consists of treprostinil sodium salt. [00032] In another embodiment, component c) comprises or consists of at least one solvent selected from the group consisting of: alcohols, amines, amides, sulphoxides, and/or esters.

[00033] In another embodiment, component c) comprises or consists of ethanol or mixtures of ethanol and propylene glycol, preferably wherein the ratio of ethanol to PG is 1 : 1 to 10: 1, e.g., 1.5: 1 to 8: 1, most preferably 2: 1 to 5:1 (e.g., around 3: 1). [00034] In another embodiment, the pre-formulation has a stability after 3 months of at least 96%, preferably at least 97%, especially at least 98%, in terms of treprostinil assay as measured by HPLC, at 25°C and 60% RH, preferably after 6 months, especially after 12 months, as defined herein.

[00035] In another embodiment, the pre-formulation has a stability after 1 month of at least 96%, preferably at least 97%, especially at least 98%, in terms of treprostinil assay as measured by HPLC following storage at 40°C and 75% RH, after preferably after 3 months, especially after 6 months.

[00036] In an especially preferred embodiment:

[00037] component a) comprises or consists of GDO;

[00038] component b) comprises or consists of PC;

[00039] component c) comprises ethanol and optionally propylene glycol; and

[00040] component d) is treprostinil or a salt thereof (e.g., sodium).

[00041] A second aspect relates to the use a pre-formulation as defined herein in the sustained administration of treprostinil or a salt thereof.

[00042] Another aspect provides a pre-formulation according to the first embodiment or a composition derived by exposing said pre-formulation to excess aqueous fluid, for use as a medicament (e.g., for use in the treatment of the conditions described herein).

[00043] Another aspect provides a method for the treatment of a human or non-human mammalian subject comprising administering to said subject a pre-formulation as defined herein. [00044] In one embodiment, the method of treatment (as well as corresponding uses and other aspects) is a method for the treatment of a human or non-human mammalian subject (especially one in need thereof). In a further embodiment, the method of treatment (as well as corresponding uses and other aspects) is a method for the treatment of at least one condition selected from pulmonary artery hypertension (PAH), PAH-associated chronic obstructive pulmonary disease (COPD), severe Raynaud’s disease or phenomenon, ischemia, and related conditions. The method of treatment (as well as corresponding uses and other aspects) is a method for the treatment of Raynaud’s disease or phenomenon, e.g., including Raynaud’s phenomenon secondary to lupus, scleroderma, Buerger disease, Sjogren syndrome, rheumatoid arthritis, polymyositis, blood disorders, thyroid disorders, or pulmonary hypertension. [00045] In an embodiment, the method of treatment involves administration a pre-formulation as defined herein once per week for as long as needed, effective, and/or tolerated. In another embodiment, the pre-formulation is administered once every 7 (±1) days, twice every 14 (±2) days, four times every 28 (±3) days, eight times every 56 (±4) days, sixteen times every 112 (±5) days, 32 times every 224 (±6) days, and so forth. In another embodiment, the pre-formulation is administered once every 7 (±1) days (once weekly). In another embodiment, the pre-formulation is administered once every 14 (±1) days; or once every 4 weeks ((±3) days.

[00046] In an embodiment, the method of treatment involves administering treprostinil or salt thereof at a level of about 1.5 to 15 mg/week, such as 20 to 12.5 mg/week or 2.5 to 10 mg/week, or 3 to 7.5 mg/week (e.g., 2.5, 3, 3.5, 4, 4.5, 5, 5.5, 6, 6.5, 7, 7.5, 8, 8.5, 9, 9.5, 10, 10.5, 11, 11.5, 12, 12.5, 13, 13.5, 14, 14.5, 15 mg/week).

[00047] In an embodiment, the method of treatment involves administering treprostinil or salt thereof at a level of 0.005 to 2.5 mg/kg/week, preferably at a level of 0.01 to 1 mg/kg/week, especially 0.015 to 0.7 mg/kg/week.

[00048] Another aspect relates to a pre-formulation as described herein for use in a method of treatment as described herein (including all diseases, conditions, dosages, methods or administration, and administration protocols described herein).

[00049] Another aspect relates to the use of a pre-formulation as defined herein in the manufacture of a medicament for use in the in vivo formation of a depot for treatment of at least one condition selected from pulmonary artery hypertension (PAH), PAH-associated COPD, Raynaud’s disease or phenomenon, ischemia, and related conditions, preferably Raynaud’s disease or phenomenon, such as Raynaud’s phenomenon secondary to lupus, scleroderma, Buerger disease, Sjogren syndrome, rheumatoid arthritis, polymyositis, blood disorders, thyroid disorders, or pulmonary hypertension..

[00050] Another aspect provides a pre-filled administration device containing a pre-formulation as defined herein.

[00051] Another aspect relates to a kit comprising an administration device as defined herein, preferably including an auto-injector, cartridge, and/or pen.

BRIEF SUMMARY OF THE ATTACHED FIGURES [00052] FIG. 1A. In vitro release profiles of selected formulations from Table 1 as a function of time.

[00053] FIG. IB. In vitro release profiles of selected formulations from Table 1 as a function of the square root of time.

[00054] FIG. 2. Results of bodyweight change in rats during the pilot study dosing using formulations Bl and B2 (see Example 2 and Table 2).

[00055] FIG. 3. Viscosity of selected formulations L-AA (see Example 3 and Table 6)

[00056] FIG. 4A. In vitro release profiles of formulations N, P, Q, R and S (cumulative percentage release).

[00057] FIG. 4B. In vitro release profiles of formulations N, P, Q, R and S with the 0-20% release region in expanded view.

[00058] FIG. 5. X-ray diffractograms at 25°C, 37°C and 42°C of Formulations L-S after equilibration in aqueous medium.

[00059] FIG. 6. X-ray diffractograms at 25°C, 37°C and 42°C of Formulations T-AA after equilibration in aqueous medium.

[00060] FIG. 7. Mean plasma concentrations of TPN in rats following administration of Formulations EE, FF, GG or HH.

[00061] FIG. 8. In vitro release profiles (cumulative percentage release) of formulations FF, EE, X and HH.

[00062] FIG. 9. Mean Treprostinil Plasma Concentration-Time Profiles Following a Single Subcutaneous Injection of 3, 15, 22.5 and 30 mg TPN in pre-formulation to a Male and Female Beagle Dogs.

[00063] FIG. 10. Mean Treprostinil AUCo-i68hr Values Following A Single Subcutaneous Injection of 3, 15, 22.5, and 30 mg TPN in pre-formulation to Male and Female Beagle Dogs.

[00064] FIG. 11. Proposed investigation of a parallel arm, open-label trial designed to characterize the safety and tolerability of CAM2043 when subjects are given repeated, once- weekly subcutaneous (SC) injections at ascending or steady doses of CAM2043.

DETAILED DESCRIPTION [00065] The formulations of the present disclosure generate a non-lamellar liquid crystalline phase following administration. The use of non-lamellar phase structures (such as liquid crystalline phases) in the delivery of bioactive agents is now relatively well established. A most effective lipid depot system for general use is described in W02005/117830, and a suitable lipid matrix for use in the present disclosure is described in general terms in that document, the full disclosure of which is hereby incorporated herein by reference. For a description of the most favourable phase structures of such formulations, attention is drawn to the discussion in W02005/117830 and particularly to page 29 thereof.

[00066] All % are specified by weight herein throughout, unless otherwise indicated. Furthermore, the % by weight indicated is the % of the total pre-formulation including all of the components indicated herein where context allows. Weight percentages of treprostinil will be calculated on the basis of the weight of free acid irrespective of whether the acid or a salt thereof is used. The pre-formulations can optionally consist of essentially only the components indicated herein (including where appropriate additional optional components indicated herein below and in the attached claims) and in one aspect consist entirely of such components. Where a formulation is indicated as “consisting essentially of’ certain components herein, when the specified components provide the essential nature of that formulation, such as when the specified components constitute at least 95%, preferably at least 98%, of the formulation.

[00067] Preferably the pre-formulation according to the disclosure is a molecular solution or has an L2 phase structure (prior to administration). The pre-formulation forms a non-lamellar (e.g., liquid crystalline) phase following administration. Such a phase change is typically brought about by absorption of aqueous fluid from the physiological environment, as indicated herein. Although it has previously been established in W02012/160213 that a carefully controlled amount of water can be tolerated provided that a mono-alcoholic solvent is present, it will be understood that upon administration the pre-formulation is exposed to a large amount of aqueous fluid. Typically, the pre-formulation will form a non-lamellar phase upon contact with at least an equivolume amount of aqueous fluid.

[00068] The present inventors have now surprisingly established that by appropriate choice of types, absolute amounts, and ratios of lipid components along with treprostinil and a biocompatible organic solvent, the release properties of the depot compositions formed from the pre-formulations of the disclosure can be rendered highly advantageous and superior to existing depot formulations of treprostinil. In particular, the release duration of a single administration of treprostinil is far beyond that of existing treprostinil depots, with the maximum plasma concentration in vivo being only a small multiple of the average or even minimum concentration during the dosing period.

[00069] Component a) - Acyl Lipid / Tocopherol

[00070] Preferable ranges for component a) are 15-85 wt.%, preferably 20-80 wt.%, preferably 30-60 wt.%, preferably 35-55 wt.%, such as 38-52 wt.%, especially 38 to 52 wt.%. Levels of around 43 wt.% (e.g., 41 to 45 wt.%) are particularly useful in some embodiments.

[00071] Preferable ranges for component b) are 15-85 wt.%, preferably 20-80 wt.%, preferably 30-60 wt.%, preferably 35-55 wt.%, such as 38-52 wt.%, especially 38 to 52 wt.%. Levels of around 43 wt.% (e.g., 41 to 45 wt.%) are particularly useful in some embodiments.

[00072] Ratios of a:b are typically 40:60 to 60:40, preferably 45:55 to 55:45 and, e.g., 47:53 to 53:47. Ratios of around 50:50 (e.g., ±2) are highly effective.

[00073] Component “a” as indicated herein comprises one or more of a mono- or di-acyl lipid and/or a tocopherol. Most preferably component a) comprises or consists of a mono- or diacyl lipid and thus has one or two non-polar “tail” groups. Acyl glycerols for use in the present disclosure (e.g., mono- or di-acyl glycerols) will generally not form a non-lamellar liquid crystalline phase structures as a pure compound in water at 25°C.

[00074] In one embodiment, component a) may be a mono-acyl lipid. Mono-acyl lipids contain a polar “head” group and one non-polar “tail group”. The “head” group may be glycerol, diglycerol, sugar moieties (such as inositol and glucosyl based moieties), and esters of polyols, such as acetate or succinate esters.

[00075] In a particular embodiment, component a) comprises or consists of at least one diacyl lipid, preferably a diacyl glycerol (DAG). A diacyl lipid comprises a polar head group as described above and two apolar tail groups, preferably linked to the polar head group via an ester linkage. The most preferred polar head group for diacyl lipids is glycerol.

[00076] The non-polar group(s) may have the same or a differing number of carbon atoms and may each independently be saturated or unsaturated. Examples of non-polar groups include Ce- C32 alkyl and alkenyl groups, which are typically present as the esters of long chain carboxylic acids. These are often described by reference to the number of carbon atoms and the number of unsaturations in the carbon chain. Thus, CX:Z indicates a hydrocarbon chain having X carbon atoms and Z unsaturations. Examples particularly include lauroyl (C12:0), myristoyl (C14:0), palmitoyl (C16:0), phytanoyl (C16:0), palmitoleoyl (C16: l), stearoyl (C18:0), iso-stearoyl (C18:0), oleoyl (C18: l), elaidoyl (C18: l), linoleoyl (C18:2), linolenoyl (C18:3), arachidonoyl (C20:4), behenoyl (C22:0), and lignoceroyl (C24:9) groups. Thus, typical non-polar chains are based on the fatty acids of natural ester lipids, including caproic, caprylic, capric, lauric, myristic, palmitic, phytanic, palmitolic, stearic, oleic, elaidic, linoleic, linolenic, arachidonic, behenic or lignoceric acids, or the corresponding alcohols. Preferable non-polar chains are palmitic, stearic, oleic, and linoleic acids, particularly oleic acid.

[00077] Mixtures of any number of mono- or diacyl lipids may be used as component a). Preferably this component will include at least a portion of C18 lipids (e.g., DAG having one or more (i.e., one or two) C18:0, C18: l, C18:2, or C18:3 non-polar groups), such as sorbitan monooleate (SMO), glycerol dioleate (GDO), and/or glycerol dilinoleate (GDL). A highly preferred example is DAG comprising at least 50%, preferably at least 80%, and even comprising at least 93%, GDO.

[00078] Since GDO and other mono- and di -acyl glycerols are products derived from natural sources, there is generally a certain proportion of “contaminant” lipid having other chain lengths etc. In one aspect, GDO as used herein is thus used to indicate any commercial grade of GDO with concomitant impurities (i.e., GDO of commercial purity). These impurities may be separated and removed by purification but providing the grade is consistent this is rarely necessary. If necessary, however, “GDO” may be essentially chemically pure GDO, such as at least 80% pure, such as at least 90% pure and, e.g., at least 93% pure GDO.

[00079] Component b) - Phospholipid

[00080] Optional component “b” in the preferred lipid matrices of the present disclosure is at least one phospholipid. As with component a), this component comprises a polar head group and at least one non-polar tail group. The difference between components a) and b) lies principally in the polar group. The non-polar portions may thus suitably be derived from the fatty acids or corresponding alcohols considered above for component a). The phospholipid (e.g., PC) will contain two non-polar groups. Again, C18 groups are preferred and may be combined with any other suitable non-polar group, particularly C16 groups. The phospholipids for use herein may be those which do not form a non-lamellar liquid crystalline phase structures as a pure compound in water at 25°C. Alternatively, the phospholipids for use herein may be those which form a non- lamellar liquid crystalline phase structure, e.g., a hexagonal liquid crystalline phase, in water at 25°C.

[00081] The phospholipid portion, even more suitably than any diacyl glycerol portion, may be derived from a natural source. Suitable sources of phospholipids include egg, heart (e.g., bovine), brain, liver (e.g., bovine), and plant sources including soybean. Such sources may provide one or more constituents of component b), which may comprise any mixture of phospholipids.

[00082] Suitable polar head groups for component b) include phosphatidylcholine, phosphatidylethanolamine, phosphatidylserine, and phosphatidylinositol. Most preferred are phosphatidylcholine (PC) and/or phosphatidylethanolamine (PE).

[00083] In the present disclosure component b) comprises or consists of one or more PCs. For instance, at least 50% of the head groups of component b) should be PC, preferably more that 65% of the head groups, especially more than 85% or more than 90%. Any single PC or mixture of PCs from these or other sources may be used, but mixtures comprising soy PC or egg PC are suitable. The PC component may contain at least 50% soy PC or egg PC, e.g., at least 75% soy PC or egg PC.

[00084] In one embodiment, applicable to all aspects and embodiments disclosed herein, component b) comprises or consists of PC. The PC may be derived from soy. The PC comprises 18:2 fatty acids as the primary fatty acid component with 16:0 and/or 18: 1 as the secondary fatty acid components. These are preferably present in the PC at a ratio of between 1.5: 1 and 6: 1. PC having approximately 60-65% 18:2, 10 to 20% 16:0, 5-15% 18: 1, with the balance predominantly other 16 carbon and 18 carbon fatty acids is typical of soy PC. Phosphatidyl choline may also contain lysophosphatidylcholine (NMT 3%) and triglycerides (NMT 2%). Generally, the phosphatidyl choline used in the lipid composition should contain at least about 94% phosphatidyl choline.

[00085] In an alternative but equally preferred embodiment, also applicable to all aspects and embodiments disclosed herein, the PC component may comprise synthetic dioleoyl PC (DOPC). This is believed to provide increased stability and so will be particularly preferable for compositions needing to be stable to long term storage, and/or having a long release period in vivo. In this embodiment the PC component contains at least 50% synthetic dioleoyl PC, e.g., at least 75% synthetic dioleoyl PC, and most preferably essentially pure synthetic dioleoyl PC. Any remaining PC may be soy or egg PC as above.

[00086] In one embodiment, the precursor formulations of the present disclosure are comprised at least partially of synthetic DOPC (i.e., PC having at least 95% PC head groups and at least 90% oleoyl (C18: 1) acyl groups) and has a stability to storage at 15-25°C, defined as less than 5% active agent degradation, as assayed by HPLC, after at least 6 months, e.g., at least 12 months and such as at least 24 months.

[00087] Since the pre-formulations are to be administered to a subject for the controlled release of treprostinil, it is important that the components are biocompatible. In this regard, the preferred lipid matrices for use in the pre-formulations of the present disclosure are highly advantageous since both PC and DAGs are well tolerated and are broken down in vivo into components that are naturally present in the mammalian body.

[00088] Synthetic or highly purified PCs, such as dioleoyl phosphatidylcholine (DOPC) and palmitoyl oleoyl phosphatidylcholine (POPC), as well as the other various high-purity PCs described herein, are highly appropriate as all or part of component b).

[00089] In one embodiment, the absolute amount of component a) will be 40 to 47 wt.%, the absolute amount of component b) will be 40 to 47%, the ratio of a:b will be 48:52 to 52:48, the amount of component c) will be 5 to 20 wt.%, preferably 8 to 12 wt.% wherein component c) consists of ethanol and propylene glycol at a ratio of 2.5: 1 to 3.5: 1, and component d) will be treprostinil sodium at 1.5 to 15 mg/ml (based on free acid), such as 5 to 12 mg/mL (based on free acid), such as 10 mg/mL (based on free acid).

[00090] Component c) - Biocompatible organic solvent

[00091] Component c) of the pre-formulations is a biocompatible organic solvent. Since the pre-formulation is to generate a depot composition following administration (e.g., in vivo), typically upon contact with aqueous fluid, it is desirable that this solvent be tolerable to the subject and be capable of mixing with the aqueous fluid, and/or diffusing or dissolving out of the preformulation into the aqueous fluid. Solvents having at least moderate water solubility are thus preferred. [00092] Component c) comprises or consists of a biocompatible organic solvent selected from the group consisting of: alcohols including mono-alcoholic solvents and di- and polyalcoholic solvents, amines, amides, sulphoxides, or esters. It is particularly preferred that component c) comprises or consists of a mono-alcoholic solvent.

[00093] Component c) may comprise two or more components from the list of solvents above, particularly a mono-alcoholic solvent and a solvent selected from amides, sulphoxides, or dialcoholic solvents. Any solvent(s) which is not a mono-alcoholic solvent may be referred to herein as the co-solvent. Where two or more solvents are present especially preferred combinations are ethanol and an amide (such as ethanol and NMP), ethanol and a sulphoxide (such as ethanol and DMSO), or ethanol and a di- or poly-alcoholic solvent (such as ethanol and PG). Some embodiments relate to combination of solvents of ethanol and PG, particularly where the ratio of ethanol to PG is 1 :5 to 20: 1, such as 1 : 1 to 10: 1, such as 1.5: 1 to 8: 1, such as 2: 1 to 5: 1 (e.g., around 3: 1, such as 2.8:1 to 3.2: 1). Component c) may comprise or consists of ethanol, propanol, iso-propanol, benzyl alcohol, or mixtures thereof. Most preferably component c) is ethanol, optionally containing PG.

[00094] The amount of component c) in the pre-formulation will have a considerable effect upon several features. In particular, the viscosity and the rate (and duration) of release will alter significantly with the solvent level. The amount of solvent will thus be at least sufficient to provide a low viscosity mixture but will additionally be determined so as to provide the desired release rate. This may be determined by routine methods in view of the Examples below. Typically a level of 1 to 30 wt.%, particularly 2 to 25 wt.% solvent will provide suitable release and viscosity properties. This will preferably be 4 to 22 wt.%, preferably 5 to 20 wt.%, preferably 6 to 18 wt.%. These levels include any co-solvent present as part of component c), as mentioned above.

[00095] As indicated above, the amount of component c) in the pre-formulations of the invention will be at least sufficient to provide a low viscosity mixture (e.g., a molecular solution) of components a), b), c), and d) and will be easily determined for any particular combination of components by standard methods.

[00096] A highly preferred combination for components a), b), and c) is GDO, PC, and ethanol, and optionally containing PG. Other preferred combinations include GDO/SPC/ethanol/DMSO, GDO/SPC/ethanol/NMP, and GDO/SPC/ethanol/PG. As indicated above, appropriate amounts of each component suitable for the combination are those amounts indicated herein for the individual components, in any combination.

[00097] Component c) as used herein may be a single solvent or a mixture of suitable solvents but will generally be of low viscosity. This is important because one of the key aspects of the present disclosure is that it provides pre-formulations that are of low viscosity and a role of a suitable solvent is to reduce this viscosity. This reduction will be a combination of the effect of the lower viscosity of the solvent and the effect of the molecular interactions between solvent and lipid composition. One observation of the present inventors is that the oxygen-containing solvents of low viscosity described herein have highly advantageous and unexpected molecular interactions with the lipid parts of the composition, thereby providing a non-linear reduction in viscosity with the addition of a small volume of solvent.

[00098] The viscosity of the “low viscosity” solvent component c) (single solvent or mixture) should typically be no more than 18 mPas at 20°C. This is preferably no more than 15 mPas, such as no more than 10 mPas, and most preferably no more than 7 mPas at 20°C.

[00099] Component d) - Treprostinil (TPN) or a salt thereof

[000100] The pre-formulations of the present disclosure contain treprostinil (TPN) or a salt thereof. Treprostinil is shown below.

[000101] In all embodiments in which the treprostinil is a pro-drug it is preferred that the prodrug is formulated in a pre-formulation comprising:

[000102] a) at least one of a mono-, di-, or tri-acyl lipid and/or a tocopherol;

[000103] b) optionally at least one phospholipid; and

[000104] c) at least one biocompatible, organic solvent; [000105] wherein the pre-formulation forms, or is capable of forming, at least one liquid crystalline phase structure upon contact with excess aqueous fluid.

[000106] In particular, in all embodiments in which the treprostinil is a pro-drug it is preferred that the pro-drug is formulated in a pre-formulation comprising:

[000107] a) a diacyl lipid, e.g., glycerol dioleate (GDO);

[000108] b) at least one phospholipid, e.g., phosphatidyl choline (PC); and

[000109] c) at least one biocompatible, organic solvent, e.g., ethanol, optionally containing

PG.

[000110] Any biologically acceptable salt of treprostinil may also be used. Where amounts of component d) are given as a percentage by weight, the weight based on the free acid is meant unless context allows otherwise. In a particularly preferred embodiment component d) comprises or consists of treprostinil free acid (TPN) or a salt thereof, most preferably treprostinil sodium salt (TPN(Na)). The use of ester derivatives, such as ethyl esters or esters of other biologically tolerable alcohols (including diols or poly-hydroxy alcohols such as propylene glycol or glycerol) is also envisaged and may provide a “prodrug” effect which may be useful in controlling the release of the treprostinil and/or its biological half-life.

[000111] Component d) is present in an amount of 0.1 to 15 wt.%, based on the treprostinil free acid, preferably 0.1 to 10 wt.%, such as 1 to 12 wt.%, especially 2 to 8%. In some embodiments, the level of treprostinil may be 4 to 8%.

[000112] Component d) is present in an amount of 5 to 30 mg/mL, based on treprostinil free acid, preferably 5 to 20 mg/mL, such as 7.5 to 15 mg/mL, e.g. 10 mg/mL based on treprostinil free acid. [000113] The release duration of treprostinil is dependent on both the amount of treprostinil and the nature of the solvent component c). Accordingly, the release properties of the depot may be tuned by varying one or more of these parameters.

[000114] In general, with depot formulations of mono- and/or diacyl lipids (such as GDO) with phospholipids such as phosphatidyl choline, the release of active agent is primarily controlled by the phase behaviour of the formulation, which in turn is primarily controlled by the nature and proportion of the lipid components. In the present case, however, the inventors have established that the release properties, and in particular the maximum in vivo concentration reached following administration (Cmax) can be usefully optimised by choice of solvent and solvent ratio. In one embodiment, for example, the present disclosure provides precursor formulations of the present disclosure where component c) comprises, comprises essentially of, or consists of ethanol and propylene glycol wherein the ratio of ethanol to PG is between 1 : 1 and 10: 1, such as 1.5: 1 to 8: 1, such as 2: 1 to 5: 1 (e.g., around 3: 1). In many aspects or embodiments disclosed herein, the ratio of ethanol to PG is about 3 : 1. In particular, formulations having both ethanol and PG (e.g., at least 0.5% of each) where the amount of ethanol is greater than the amount of PG may provide a lower Cmax (i.e., a lower “peak” in vivo concentration) than formulations where there is an equal or lesser amount of ethanol in comparison with PG.

[000115] With regard to the level of component d), Figure 4A illustrates the in vitro release of TPN from formulations containing between 1.5 to 7.8 wt.% TPN(Na) in a matrix of GDO/PC/EtOH (45:45: 10). At loadings of 7.8 wt.% TPN(Na) the formulations exhibit “burst” characteristics, i.e., about 50% of the TPN(Na) is released after a period of about 24 hours, whereas at levels of 6.17 wt.% and below, the release of TPN(Na) is much more gradual. Being able to alter the burst profile simply by appropriate choice of components is potentially a very useful feature of pre-formulations disclosed herein. In one embodiment, these may be affected by choice of solvent and solvent ratio.

[000116] In one embodiment, particularly where it is desired that the formulation provides a short-term release over 1 to 3 days, it may be desirable to operate with a level of TPN or TPN(Na) of 6.5% or more, especially 6.7% or more, or 7% or more. However, in general the level of TPN will typically be no more than 5% wt.%, preferably no more than 4% wt.% (e.g., 0.5 to 4% wt.%, such as around 1%, around 2% or around 3% wt.%).

[000117] Short term release depots providing an effective release over a period of 1 to 3 days may be formulated with ethanol as the only component c) a level of at least 11 wt.%, such as at least 12 wt.%, especially at least 13 wt.%.

[000118] Where it is desired to provide a more gradual release of treprostinil, for instance for a week-long or fortnightly or monthly duration depot, it may be desirable to operate with levels of d) of less than 6.5 wt.%, such as 6.2 wt.% or less, especially 5.5 wt.% or less or 5 wt.% or less may be desirable. As noted above, the level of treprostinil will typically be no more than 5% wt.%, such as no more than 4% wt.% (e.g., 0.5 to 4% wt.%, such as around 1%, around 2% or around 3% wt.%). Thus, a pre-formulation for once weekly or once fortnightly administration may preferably comprise 1 to 7 wt.% of TPN or TPN(Na), such as 1 to 3 wt%.

[000119] Long term release depots providing an effective release over a period of greater than 5 days, such as weekly or fortnightly may be formulated with ethanol as the only component c) at a level of less than 11%, such as 10% or less. Alternatively, a mixture of ethanol and a di- or polyalcoholic solvent, especially ethanol and PG or ethanol and water, may be used in an amount of 5 to 20%, especially 5 to 15%, with a ratio of ethanol :PG or ethanol: water in the range of 40:60 to 60:40 (w:w), levels of about 50:50 may be preferred. An amount of around 2.5 wt.% PG and around 7.5 wt.% ethanol is used in many aspects or embodiments in the present disclosure.

[000120] In one aspect, each of the embodiments herein can optionally contain an antimicrobial or microbial-static agent, which includes bacteriostatic agents and preservative. Such agents include benzalkonium chloride, m-cresol, benzyl alcohol, or other phenolic preservatives. Typical concentrations as known in the art can be used. In most aspects or embodiments according to the present disclosure there is no need for an antimicrobial or microbial-static agent such as those described above.

[000121] Additional components above those mentioned as components a) to d) will, where present at all, be present in an amount of 0 to 5% (e.g., 0.01% to 5%) wt.%, e.g., no more than 2% wt.%.

[000122] In one embodiment, components a) and b) (allowing for any impurity inherent in the nature of these components) make up at least 95% of the lipid components of the composition. Preferably at least 99% of the total lipid content of the pre-formulation consists of components a) and b). Preferably the lipid content of the pre-formulation is made up from a) and b), and wherein any additional lipids components originate from one or more impurity associated with component a) and/or b). Preferably the lipid content of the pre-formulation is made up from glycerol dioleate and phosphatidyl choline, and wherein any additional lipids components originate from one or more impurity associated with component a) and/or b).

[000123] Administration

[000124] The pre-formulations of the present disclosure are administered by subcutaneous (s.c.) injection. [000125] The administration will be by injection, which term is used herein to indicate any method in which the formulation is passed through the skin, such as by needle, catheter, or needleless (needle-free) injector. The s.c. injection comprising pre-formulations disclosed used herein has the advantage of being less painful to the subject than the i.m. injection and is technically most suitable in the present case as it combines ease of injection with low risk of local side effects.

[000126] As used herein, the term “low viscosity mixture” or “low viscosity pre-formulation” is used to indicate a mixture which may be readily administered to a subject and in particular readily administered by means of a standard syringe and needle arrangement. This may be indicated, for example by the ability to be dispensed from a 1 ml disposable syringe through a small gauge needle. Preferably, the low viscosity mixtures can be dispensed through a needle of 19 gauge, preferably smaller than 19 gauge, more preferably 23 gauge (or most preferably even 27 gauge) needle by manual pressure. In an embodiment, the low viscosity mixture should be a mixture capable of passing through a standard sterile filtration membrane such as a 0.22 pm syringe filter. A typical range of suitable viscosities would be, for example, 10 to 1000 mPas, such as 10 to 800 mPas and, e.g., 200 to 700 mPas at 20°C.

[000127] Upon administration, the lipid-based pre-formulations of the present disclosure undergo a phase structure transition from a low viscosity mixture to a high viscosity (generally tissue adherent) depot composition (depot).

[000128] The depot systems formed by the treprostinil formulations disclosed herein allow treprostinil to be released to the patient over an extended release period. The formulations disclosed herein thus may provide in vivo depots of treprostinil which require s.c. administration only once every 1 to 60 days. Typical administration intervals will be, for example, every 1, 2, 3, 7, 14, 21, 28, 30, or 60 days and may be varied either systematically or occasionally by small amounts (e.g., by ±3 days, or by ±20% in any appropriate case). In many aspects administration frequencies include every 7 (± 1) days or every 14 (±2) days, or every 30 (±3) days. In one embodiment, formulations containing a comparatively low level of treprostinil (e.g., 0.5 to 4.0 mg/mL) may be administered once weekly, once fortnightly, or once monthly. Where the preformulation optionally is self-administered, patient compliance may be aided by a weekly (e.g., every 7 days, optionally ±1 day) or monthly (e.g., every 28 or 30 days (optionally ±7 days)) administration. [000129] In one embodiment of the present disclosure, applicable to all aspects but particularly the methods of treatment and corresponding uses, administration dose and frequency may be gradually escalated to correspond with the progression of the underlying disease (such as any of those diseases indicated herein). Thus, dosages of 1.5 mg/week - 15 mg/week, or of treprostinil may be sufficient for a subject and these may be provided as weekly, fortnightly, or monthly administrations as needed (e.g., a 1 ml injection at 10 mg/ml every 4 weeks would give an average dose of 2.5 mg/week. Alternatively, 0.5 mL injection (at 10 mg/mL) every week (5 mg/week), 1 ml (at 10 mg/ml) every fortnight, (5 mg/week), or 0.75 ml every week (7.5 mg/week) and even 1.0 ml/week (10 mg/week) could be considered. Subsequent increases may then be achieved by higher concentrations of formulation, such as 1.0 ml of 30 mg/ml. In some aspects, the dose of treprostinil is selected from 1.5 mg/week, 2.0 mg/week, 2.5 mg/week, 3.0 mg/week, 3.5 mg/week, 4.0 mg/week, 4.5 mg/week, 5.0 mg/week, 5.5 mg/week, 6.0 mg/week, 6.5 mg/week, 7.0 mg/week, 7.5 mg/week, 8.0 mg/week, 8.5 mg/week, 9.0 mg/week, 9.5 mg/week, 10.0 mg/week, 11.0 mg/week, 12.0 mg/week, 13.0 mg/week, 14.0 mg/week, 15.0 mg/week. In some aspects of the present disclosure the patient is administered a dose of treprostinil selected from 2.5 mg/week, 5.0 mg/week, 7.5 mg/week, or 10.0 mg/week. In some aspects of the present disclosure, the patient is administered a dose of treprostinil of 2.5 mg/week. In some aspects of the present disclosure, the patient is administered a dose of treprostinil of 5.0 mg/week. In some aspects of the present disclosure, the patient is administered a dose of treprostinil of 7.5 mg/week.

[000130] In some aspects of the present disclosure, the patient is initially administered a dose of treprostinil of 2.5 mg/week, for 1-4 weeks, and thereafter the dose increased to 5.0, 7.5 or even 10 mg/week. Should the patient experience unacceptable levels of adverse events the dose may be reduced according to similar administration schemes.

[000131] A considerable advantage of the depot precursors of the present disclosure is that they are stable homogeneous phases. That is to say, they may be stored for considerable periods (e.g., at least 12 months) at room or refrigerator temperature, without phase separation. As well as providing advantageous storage and facile administration, this allows for the dose of treprostinil to be selected by reference to the species, age, sex, weight, and/or physical condition of the individual subject, by means of injecting a selected volume. [000132] The present disclosure thus provides for methods comprising the selection of a dosing amount specific to an individual, particularly by subject weight. The means for this dose selection is the choice of administration volume.

[000133] The pre-formulations of the present disclosure are highly advantageous in that they are stable to prolonged storage in their final “administration ready” form. As a result, they may readily be supplied for administration either by health professionals or by patients or their carers, who need not be fully trained health professionals and may not have the experience or skills to make up complex preparations. This is particularly important in long-duration, slow-effecting diseases such as diabetes.

[000134] Devices

[000135] In a yet further aspect, the present disclosure provides a disposable administration device (which is also to include a device component) pre-loaded with a measured dose of a preformulation of the present disclosure. Such a device will typically contain a single dose ready for administration, and will generally be sterile-packed such that the composition is stored within the device until administration. Suitable devices include cartridges, ampoules, and particularly syringes and syringe barrels, either with integral needles or with standard (e.g., luer) fittings adapted to take a suitable disposable needle. Similarly appropriate devices include a needle-less injector, a multi- or single-use autoinjector combined with a pre-filled syringe, a cartridge, optionally combined with a multi-use pen device, or a vial. Evidently, such pre-filled syringes and cartridges may be for any appropriate injecting device, such as a multi-use or single-use injector or needle-less injection unit.

[000136] The devices of the present disclosure may contain the pre-formulation of the present disclosure which delivers a dosage in the range of 5 to 30 mg/ml, preferably 5 to 20 mg/ml, such as 7 to 20 mg/ml, e.g., 10 mg/ml. Dose volumes will typically be no more than 2 ml (e.g., 0.1 to 2 ml), for example 0.25 to 1.5 ml or 0.5 to 1 ml.

[000137] The devices may contain a total volume for administration of no more than 2 ml, such as no more than 1 ml, e.g., no more than 0.5 ml.

[000138] The pre-filled devices may also suitably be included in an administration kit, which kit also forms a further aspect. Provided is also a kit for the administration of treprostinil, said kit containing a measured dose of a formulation and optionally an administration device or component thereof. Preferably the dose will be held within the device or component, which will be suitable for s.c. administration. The kits may include additional administration components such as needles, swabs, etc. and will optionally and preferably contain instructions for administration. Such instructions will typically relate to administration by a route as described herein and/or for the treatment of a disease indicated herein above.

[000139] Kits

[000140] The pre-filled administration device as indicated herein and a kit as indicated herein comprising a pre-formulation as described herein. Suitable kits may include a single- or multipleuse injection device such as an auto-injector or may include cartridges or components for use in such devices. Example of devices includes pre-filled syringes, pre-filled autoinjectors, and prefilled pens.

[000141] Kits of the present disclosure will optionally include any of the following components: [000142] i) an injection device such as a syringe, auto-injector, pen injector.

[000143] ii) a dose-measurement device (e.g., a graduated device for measuring or setting administration volume).

[000144] iii) a table, chart, phone app, or electronic calculator for calculating and/or setting dosage volume based on parameters such as subject weight and/or dose frequency. Factors such as disease progression and/or treprostinil concentration may be accounted for in such calculations, either explicitly or implicitly.

[000145] iv) instructions for dosing and/or for escalation of dosing according to factors such as subject weight and/or dose frequency, disease progression (e.g., mean pulmonary artery pressure) and/or treprostinil concentration.

[000146] Features and Combinations

[000147] Component a) comprises, consists essentially of, or preferably consists of GDO. Glycerol dioleate (GDO) may also contain monoglycerides (NMT 2%) and triglycerides (NMT 5%). Generally, the GDO used in the lipid composition should contain at least about 93% glycerol di oleate.

[000148] Component b) comprises, consists essentially of, or preferably consists of soy PC and/or “high purity PC” such as DOPC. Phosphatidyl choline may also contain lysophosphatidylcholine (NMT 3%) and triglycerides (NMT 2%). Generally, the phosphatidyl choline used in the lipid composition should contain at least about 94% phosphatidyl choline. [000149] Component c) is ethanol and optional propylene glycol.

[000150] The pre-formulation has a low viscosity as indicated herein, and less than 1000 mPas. [000151] The pre-formulation comprises forms a lipid depot upon in vivo administration.

[000152] The pre-formulation generates a depot following in vivo administration, which depot releases treprostinil over a period of at least 3 days, such as at least 5 days. In some aspects the depot releases pharmaceutically relevant levels of treprostinil for about 7 days.

[000153] The pre-formulation generates a depot following in vivo administration to a subject, which depot releases treprostinil such that the plasma concentration of treprostinil in said subject at the end of the seventh day following administration is no less that 10'⁴, or 10'³, preferably no less than 10'², more preferably no less than 10'¹ times the plasma concentration of treprostinil in said subject at the end of the first day following administration (i.e., at 24 hours after administration).

[000154] In combination with the features and preferred features indicated herein, the method(s) of treatment of the present disclosure may have one or more of the following features independently or in combination:

[000155] The method comprises the administration of at least one formulation with one or more features as indicated above;

[000156] The method comprises the administration of at least one formulation as indicated herein by s.c. injection;

[000157] The method comprises administration by means of a pre-filled administration device as indicated herein;

[000158] The method comprises administration through a needle no larger than 20 gauge, such as smaller than 20 gauge, and an example is 23 gauge;

[000159] The method comprises a single administration every 3 to 10 days, such as every 5 to 8 days, e.g., once weekly (e.g. 1.5 to 15 mg/week).

[000160] They method contains a homogeneous mixture of a composition of the present disclosure in ready -to-inject form. [000161] They contain a formulation of components a) to c) for combination with treprostinil whereby to form a pre-formulation.

[000162] They contain a total volume for administration of no more than 2 ml, e.g., no more than 1.5 ml.

[000163] In combination with the features and preferred features indicated herein, the kits of the present disclosure may have one or more of the following features independently or in combination:

[000164] They contain a preferred formulation as indicated herein;

[000165] They contain a pre-filled device as indicated herein;

[000166] They contain a needle smaller than 20 gauge, preferably no larger than 23 gauge;

[000167] They contain a single dose of 1 to 100 mg of treprostinil (as described herein), preferably 2 to 75 mg, e.g., 1.5 to 15 mg, e.g. 2.5, 5, 7.5 or 10 mg.

[000168] They contain a “two compartment kit” comprising at least two vessels containing a lipid formulation disclosed herein and treprostinil, respectively.

[000169] They contain a total volume for administration of no more than 5 ml, preferably no more than 3 ml, for example no more than 2 ml, e.g., no more than 1.5 ml.

[000170] They contain instructions for administration by a route and/or at a frequency as indicated herein.

[000171] They contain instructions for administration for use in a method of treatment as described herein.

[000172] As used herein, the term “about,” “around,” “substantially,” or “approximately” in relation to a number or a range of numbers will generally indicate that the number or range specified is preferred but that such a number may be varied to a certain extend without materially affecting the properties of the relevant material, composition or similar product. The skilled worker will typically be able to readily establish the extent by which such numbers may be varied without prejudicing the key advantages of the present disclosure. As a general guide, such numbers or the ends of such ranges may be varied by ±10%, or ±5% and or ±1%. A corresponding meaning may be attributed to compositions “consisting essentially of’ certain components, which may include up to 10%, such as up to 5% and, e.g., up to 1% of other components in addition to those specified. Where a chemical group, chain or other moiety is described herein as optionally substituted, such substitution may be absent or one or more atoms in the moiety (typically one or more hydrogens and/or carbons) may be substituted with groups such as halide (e.g., F, Cl, Br, I) groups, oxygenbased moieties such as ethers, alcohols, esters carboxylic acids, or epoxides, nitrogen-based groups such as amines, amides, nitriles, or nitro groups, or sulphur-based groups such as thiols, disulphides, thioesters, etc. Up to around 10 such substitutions may be made where context allows, but typically 3 or few substitutions, such as 1, 2, or 3 substitutions with independently selected substituent groups will be typical.

[000173] The disclosure will now be further illustrated by reference to the following non-limiting Examples and the attached Figures.

[000174] EXAMPLES

[000175] Materials

[000176] Treprostinil sodium salt (TPN(Na)) from Sanofi; soy phosphatidylcholine, SPC, Lipoid SI 00 from Lipoid; Glycerol di oleate, GDO, Cithrol GDO HP-SO-(LK) from Croda; di oleoyl phosphatidylcholine, DOPC, from NOF; Ethanol, EtOH (99.7% Ph. Eur), from Solveco; propylene glycol, PG (Ph. Eur), from Fischer; N-methyl pyrrolidone, NMP, and dimethylsulfoxide, DMSO, from Sigma-Aldrich were used as received. All other chemicals were of analytical grade purity.

[000177] Preparation of pre-formulations

[000178] Lipid stock mixtures were prepared by weighing appropriate amounts of SPC, GDO, and solvents into sterilized glass vials. Sealed vials were then placed on a roller mixer at room temperature (RT) until mixed completely into clear homogeneous liquid solution (<24 hours). TPN(Na) powder was added to the respective lipid placebo formulations in new glass vials. Vials were then sealed and placed on a roller mixer at RT until mixed completely into clear homogeneous liquid solution (<24 hours). Prepared formulations were stored at RT in the dark until further experiments. For explorative stability evaluation, formulations were divided into sterilized 2R glass vials (1 g of formulation per vial). Vials were sealed and placed in controlled environment storage cabinets. At predefined sampling points two vials of formulation were withdrawn from each storage cabinet, placed at room temperature for 1 hour and analyzed for content and purity using gradient HPLC with UV detection.

[000179] Example 1: Evaluation of TPN(Na) solubility and in vitro release [000180] The solubility was assessed by adding TPN(Na) to respective lipid stock mixtures followed by mixing on a roller mixer at room temperature (RT) until mixed completely into clear homogeneous liquid solution. During preparation samples were visually inspected. Results showed that TPN(Na) has good solubility in a variety of pre-formulations and that a drug load of at least 7 wt.% (~ 78 mg TPN(0)/mL) is feasible. As shown in Table 1, measured viscosities of the formulations range between 185-628 mPas depending on co-solvent type, concentration, and composition.

Table 1. Viscosities of various pre-formulations containing 7 wt.% TPN(Na).

[000181] In vitro release testing of TPN(Na) was performed using a straightforward assay based on UV/VIS spectroscopy for quantification. In the test, depots were prepared by injecting 0.03- 0.10 g (target 0.1 g) of the respective pre-formulation into 10 mL of PBS (pH 7.4) kept in 20R glass injection vials. The exact amount of formulation added to each vial was determined by weighing. The vials were sealed with rubber stoppers and aluminum crimp caps and placed on shaking in an incubator held at 37°C. Release media were sampled at scheduled time points, diluted and transferred to quartz cuvettes and analyzed on a Perkin Elmer Lambda 25, double-beam, UV- VIS spectrophotometer at 273 nm.

[000182] The results from the in vitro release measurements are displayed in Figure 1A. From the results it is evident that both solvent amount and composition affect the initial in vitro release of treprostinil. Formulations comprising PG as co-solvent have a slower initial release (24 h) than formulations with DMSO. Also, when comparing formulations with EtOH as only solvent, the initial release is faster for higher solvent content. Figure IB further indicates that the release in vitro is biphasic, after the initial phase where solvent and drug is released concomitantly, the release is linear with square root of time as expected by a diffusion-controlled release mechanism from a monolithic depot matrix.

[000183] Example 2: Administration of pre-formulations with TPN(Na) in rats: formulations and body weight change

[000184] The primary objective of this pilot study was to evaluate the tolerability of TPN, both locally and systemically following single subcutaneous injections of pre-formulations with TPN(Na) to rats (formulation compositions are given in Table 2). The study was designed as a dose escalation study, with doses of administration of 3, 9 and 27 mg/kg TPN (Table 3).

Table 2. Formulation compositions used in the pilot rat study.

Table 3. Treatment groups and doses of TPN used in the pilot study.

[000185] Figure 2 shows the mean relative body weight change during the study. Formulations

Bl and B2 were monitored in the pilot study for injection site erythema/edema as follows:

[000186] The extent of erythema and edema formation following administration of formulations

Bl and B2 is indicated in the Table 4 below:

Table 4. Summary of erythema and edema at the injection sites during the pilot study.

[000187] Formulations Bl and B2 were monitored in the pilot study for angiogenesis / haemorrhage as follows:

[000188] The level of angiogenesis was defined by a range from 0 to 3:

[000189] 0 for no angiogenesis.

[000190] 1 for minor angiogenesis. Limited growth of blood vessels.

[000191] 2 for medium angiogenesis. Expanded growth of blood vessels

[000192] 3 for major angiogenesis. Extensive growth of blood vessels.

[000193] The level of haemorrhage was defined by a range from 0 to 3:

[000194] 0 for no haemorrhage.

[000195] 1 for minor haemorrhage. Diffuse redness area or areas.

[000196] 2 for medium haemorrhage. At least one well defined red area.

[000197] 3 for major haemorrhage. Several well defined red areas.

[000198] The extent of angiogenesis and haemorrhage following administration of formulations Bl and B2 is indicated in Table 5 below.

Table 5. Summary of injection site findings at necropsy in the pilot study.

[000199] Example 3: Effect on nanostructure of the fully hydrated pre-formulations as a function of different amounts of TPN(Na)

[000200] Formulations L to AA below were prepared which compositions and measured viscosities are given in Table 6 and Figure 3, respectively.

Table 6. Formulation codes and compositions used for nanostructural evaluation.

[000201] The nanostructure of the fully hydrated formulations from Table 6 was evaluated using small angle X-ray diffraction. Briefly, about 100 mg of the formulation was injected into 5 mL PBS buffer and left to equilibrate at ambient RT in still standing vials for 8 days before SAXD measurements. The nanostructure of fully hydrated formulations as a function of TPN(Na) concentration was studied using synchrotron SAXD measurements, performed at the 1911-4 beamline at MAX IV laboratory (Max II electron accelerator operating at 1.5 GeV, Lund University, Sweden), using a IM PILATUS 2D detector (Dectris) containing a total of 981 x 1043 pixels. Samples were mounted between thin polyimide films in a custom made steel sample holder at the sample to detector distance of 1919.5 mm. Diffractograms were recorded with a X-ray wavelength of 0.91 A and the beam cross-section of 0.25 x 0.25 mm (full width at the halfmaximum) at the sample. Temperature control was achieved using computer controlled Lauda RE 420G thermostat (Lauda-Brinkmann, LP). The experiments were performed successively at 25, 37, and 42°C with a 60 s exposure time at each temperature and a wait of 10 minutes between temperature steps. The resulting CCD images were integrated and analyzed using the Fit2D software provided by ESRF (European Synchrotron Radiation Facility, France). Silver behenate calibrated sample-to-detector distance and detector positions were used.

[000202] Figure 5 and Figure 6 show obtained SAXD results of the nanostructure of the fully hydrated Lipid/EtOH (90/10 wt.%) and Lipid/EtOH/PG (85/7.5/7.5) formulations as a function of TPN concentration and temperature. Data in Figure 5 show that in the temperature region of 37- 42°C fully hydrated 10% EtOH based formulations form mixtures of reversed hexagonal (H2) and reversed micellar cubic (Fd3m) phases up to 3.1 wt.% of TPN. At 4.65 and 6.2 wt.% of TPN a single H2 is formed which at even higher concentrations of TPN starts to transform into disordered micellar solution (L2). In addition, with increasing TPN concentration the lattice parameter for Fd3m phase remains unchanged whereas it starts to increase for the H2 phase. The increase of the lattice parameter starting from 4.65 wt.% of TPN(Na) is likely related to the increased mechanical softness of the depots observed in gelling experiments. Overall, the observed transformation from mixture of Fd3m and H2 to single H2 and further to a mixture of H2 and L2 phases with increasing concentration of TNP correlates with the obtained in vitro release results where drastic increase in released TPN is found at 6.2 and 7.75 wt.% of TPN (Figures 4A, 4B).

[000203] As a comparison, Figure 6 shows SAXD results obtained for the fully hydrated formulations prepared at 7.5/7.5 wt.% of EtOH/PG mixture. Here, in the temperature region of 37- 42°C, clearly pronounced mixture of Fd3m and H2 phases is formed only up to 1.55 wt.% of TPN. With increasing TPN concentration between 2.33 and 6.2 wt.% of TPN an H2 phase mixture with L2 phase is observed. Furthermore, at 7.75 wt.% of TPN a mixture of lamellar (La) and L2 phase is formed. In addition, the lattice parameter for the hexagonal phase starts to increase already at 3.10 wt.% of TPN(Na). Based on these results, it may be concluded that fully hydrated formulations prepared using EtOH/PG 7.5/7.5 can accommodate less of TPN before transforming into more swollen liquid crystalline phases (especially La) which are less favourable from the perspective of sustained release.

[000204] In-vitro release profiles for formulations N, P, Q, R, and S were measured and are shown in Figures 4A and 4B (cumulative % release).

[000205] Example 4: Physical and chemical stability of pre-formulations containing TPN(Na)

[000206] The storage stability of formulations BB, CC, and DD (Table 7 and 8) was studied by HPLC under conditions of: < -25°C (frozen conditions); 25°C/60% RH; and 40°C/75% RH. Results from the explorative stability testing indicate good physical as well as chemical stability of TPN in the pre-formulations using both 10% EtOH as solvent and with a mixture of EtOH/PG. TPN shows good stability for at least up to 3 months when stored at 25°C/60% RH and 40°C/75% RH (Table 9).

Table 7. Formulation codes and compositions used for physical and chemical stability evaluation.

Table 8. Formulation codes and viscosities used for physical and chemical stability evaluation.

Table 9. TPN assay results (HPLC) in the formulations from the explorative stability study as a function of time at different storage conditions.

[000207] Example 5: Administration of pre-formulations with TPN(Na) in rats

[000208] Formulations EE-HH were prepared using different solvent compositions (Table 10) and were administered to a group of 24 rats and monitored for Erythema, Edema Angiogenesis and Haemorrhage using the scoring system of earlier Examples.

Table 10. Formulation compositions and TPN doses used in the rat PK study.

Table 11. Extent of erythema following administration of Formulations EE-HH.

Table 12. Extent of edema following administration of Formulations EE-HH.

Table 13. Injection site at necropsy following administration of Formulations EE-HH.

[000209] Example 6: Rat PK data of pre-formulations with TPN(Na).

[000210] Pharmacokinetic data was collected over 14 days for formulations EE to HH of Example 7 administered to rats. The data is illustrated graphically in Figure 7 and the corresponding in vitro release profiles in Figure 8.

[000211] Table 14. PK parameters for Formulations EE-HH in rats. The data is illustrated graphically in Figure 9 and percentage release profile in Figure 10.

[000212] Example 7: Subcutaneous injection of pre-formulations with TPN(Na) in dogs

[000213] The objective of this study was to assess the exposure to TPN following subcutaneous injection of the pre-formulation with TPN(Na) (formulation code JJ, composition TPN(Na)/GDO/SPC/EtOH/PG 3.38/43.31/43.31/7.50/2.50 wt.%) to beagle dogs in a maximum tolerated dose toxicity study. The TPN doses (calculated as TPN acid form) used in the dog study are given in Table 15. The obtained dose dependent PK profiles and exposure (AUC0-168h) values are presented in Figure 9 and Figure 10, respectively.

Table 15. TPN dose levels and volumes used in the dog PK study.

[000214] Example 8: An Exploratory Trial Assessing Finger Perfusion, Raynaud’s Condition Score and Safety, Following a Single Dose of CAM2043 (Treprostinil Subcutaneous Depot) in Patients with Raynaud’s Phenomenon Secondary to Systemic Sclerosis

[000215] Background

[000216] Prostacyclin analogues, although effective treatment for systemic sclerosis (SSc)- related Raynaud’s phenomenon (RP), have the disadvantage of requiring intravenous or subcutaneous (SC) infusion. Treprostinil subcutaneous depot (CAM2043) is a novel lipid-based SC formulation of treprostinil (e.g. 10 mg/mL, excipients GDO and PC ratio 50:50, solvent ethanol /7.5 wt.% and PG 2.5 wt.%). The aim of this trial was to explore the effect of a single SC dose of CAM2043 on skin temperature (indirect measure of perfusion), as evaluated by thermography following cold challenge.

[000217] Methods

[000218] This was an exploratory, open-label, single-dose Phase 2 trial. Ten female patients (mean age 54.9 years) with mean SSc duration of 11.5 years (all anticentromere antibody positive) and mean RP duration of 14.5 years, were recruited. To be included, patients had to experience >5 attacks of RP/week. Patients attended on 6 occasions: screening, baseline (Day 1), and on Days 2, 3, 8, and 15. At baseline, each patient received a single SC injection of 2.5 mg CAM2043 (except the first patient who received 5 mg CAM2043, due to non-serious systemic adverse drug reactions (ADRs) related to CAM2043 following treatment of the first patient, it was decided to lower the dose to 2.5 mg). A standard cold challenge test of the hands (15°C for one minute) was performed pre-dose and at 3, 6, 24, 72, 168, and 336 hours post-dose, with temperature responses over the subsequent 15 minutes measured by infrared thermography. The primary endpoint was the mean change from baseline to 6 hours post-dose in the area under the curve (AUCtherm) for rewarming (8 fingers). Secondary endpoints included: change from baseline in AUCtherm, maximum temperature after rewarming (MAX), and Raynaud’s Condition Score (RCS). tolerability were assessed throughout the trial.

[000219] Results

[000220] AUCtherm increased 6 hours post-dose, but not statistically significantly: mean increase 192.7°C*sec (95% CI: -727.1, 1112.6). AUCtherm was significantly greater at 24 hours than at baseline (mean increase 1175.8°C*sec (95% CI: 127.3, 2224.3)), with a trend towards mean AUCtherm values higher than baseline up to 168 hours post-dose (Day 8) and returning to baseline values at 336 hours (Day 15). A statistically significant increase from baseline was also seen for MAX at 24 hours: mean increase 1.4°C (95% CI: 0.1, 2.7). At baseline, mean (SD) RCS was 3.7 (1.3) units (10-point scale), and was significantly improved post dosing, including Day 8 (mean reduction 1.6 units (95% CI: -2.68, -0.52)). This positive treatment effect persisted at Day 15 (mean reduction 1.6 units (95% CI: -3.33, 0.19)). Adverse events were reported by all 10 patents: all reported erythema and pain at the injection site.

[000221] Conclusion

[000222] Finger skin temperature rose after CAM2043 treatment (significantly at 24 hours postdose), with significant improvements in RP symptoms (RCS) over the 15-day period following dosing indicating a positive treatment effect. Overall, the safety profile for CAM2043 was consistent with known safety profile for treprostinil, including injection site reactions. All patients had adverse events and there may be a dose-limiting response potentially caused by systemic and local reactions. The results indicate that CAM2043 could be further investigated in clinical trials of Raynaud's phenomena.

[000223] Example 9: An Exploratory Trial Assessing Vascular Digital Perfusion, Pharmacokinetics, Safety, and Tolerability Following a Single Dose of CAM2043 (Treprostinil Subcutaneous Depot) in Patients with Raynaud’s Phenomenon Secondary to Systemic Sclerosis

[000224] 1 INTRODUCTION

[000225] 1.1 CAM2043

[000226] CAM2043 (‘treprostinil subcutaneous depot’) is a novel and long-acting lipid-based pharmaceutical formulation of treprostinil for subcutaneous (SC) administration. The application of CAM2043, alleviating the need for continuous parenteral infusion, and may be an attractive alternative to increase patient convenience, safety, and tolerability. CAM2043 is designed to provide therapeutic levels of treprostinil over the target duration of 7 days following injection. The intended commercial product presentation is a ready -to-use, pre-filled syringe equipped with a needle stick prevention safety device compatible with room temperature storage. [000227] The PK, safety, and tolerability of CAM2043, after single and repeated doses, has been evaluated in a Phase 1 trial with healthy subjects.

[000228] 2 1 OBJECTIVES, TRIAL DESIGN, AND KEY SELECTION CRITERIA [000229] 2.1 Objectives

[000230] The following information relates to the objectives of the trial and rational behind their selection.

[000231] 2.1 Background and Rationale

[000232] 2.3 Trial Design

[000233] This is an exploratory, single-site, open-label, single-group, single-dose trial evaluating the effect of CAM2043 on digital vascular perfusion in patients with Raynaud’s phenomenon secondary to systemic sclerosis.

[000234] At Screening (Visit 1), patients must provide written informed consent to participate in the trial before any trial related procedures are performed. The patient’s eligibility will be confirmed, and the medical history and prior and concomitant medications will be recorded. Blood and urine samples will be collected for clinical laboratory testing, urine drug and alcohol screen, and serum pregnancy test. A complete physical examination, including assessments of vital signs, height, weight, body mass index (BMI) and electrocardiogram (ECG) will be performed. The patient’s Child-Pugh status will be recorded. Patients will be subjected to cold challenge followed by thermography to assess the change in skin temperature (i.e., reperfusion/rewarming). Thermography will be assessed twice during the Screening visit in order to be able to exclude patients with variability greater than ±20%. [000235] At the Baseline Visit (Visit 2), patients will be subjected to cold challen^ by assessment of digital vascular perfusion using thermography immediately predose. After imaging, patients will receive a single SC injection of 5 mg CAM2043. After dosing, the cold challenge will be repeated followed by thermography at 3, 6, 24, 72, and 168 hours postdose. After Week 1, the patients will enter a 1-week follow-up period. At Day 15 the patients will visit the clinic and be subjected to cold-challenge followed by thermography.

[000236] Pharmacokine tics:

[000237] Venous blood samples (4 mL each) will be collected. Samples will be collected, processed, and shipped according to instructions provided separately. The actual date and time of each blood sample collection will be recorded.

[000238] Proposed time-points for plasma sampling: Predose and at approximately 3, 6, 24, 72, 168 and 336 hours postdose.

[000239] Efficacy assessments

[000240] The effect of CAM2043 on patient reported outcomes will be assessed using Pain visual analogue scale (VAS), Raynaud’s Condition Score, and a Likert scale rating the patient’s perception of Raynaud’s phenomenon. Rating of the scales will be collected using patient diaries. The patient will fill out the diary in the morning and evening.

[000241] Trial Flow Chart

Day -14 to -1 Week 1 Week 2

Baseline Completion

(Day 1) (Day 15)

* dose changed to 2.5 mg as indicated above Schedule of Assessments

^a Predose, 3 and 6 hours after administration ^b Approximately 24 hours after administration ^c Approximately 72 hours after administration ^d Approximately 168 hours after administration ^e Approximately 336 hours after administration ^f Thermography will be performed 2 times during the Screening Visit. Patients who have a variability of more than ±20% between the thermography assessments are not eligible for the trial.

⁸ Patients will rate the Pain VAS and Raynaud’s Condition Score daily during the trial using Investigational Medicinal Product (IMP)

The actual date and time of the administered dose will be recorded.

[000242] Disallowed Medications

The following medication is prohibited throughout the trial:

[000243] Allowed Medications

[000244] The following medications are allowed throughout the trial:

[000245] Stable vasodilator therapy (4-weeks prior to screening): calcium channel blockers, phosphodiesterase Type 5 inhibitors, alpha-adrenergic blockers, nitrates, angiotensin converting enzyme inhibitors, or angiotensin receptor blockers. Investigators should keep those treatments constant during the study.

[000246] 1. Restrictions

[000247] Patients must refrain from vigorous exercise, caffeine, and alcohol for 24 hours prior to and during the entire visit to the clinic. Clinical Laboratory Tests

Women of childbearing potential will have a serum beta-hCG pregnancy test at screening and at completion/early termination. The results of the pregnancy test at screening must be reviewed and confirmed to be negative prior to enrollment to assess the patient’s eligibility for the trial.

Trial Design Constraints

The following information relates to the key elements of the trial design and where appropriate the rationale behind their inclusion.

Trial Specific Inclusion/Exclusion Criteria and Rationale

[000248] Example 10: CAM2043 IN RAYNAUD PHENOMENON

[000249] As concluded in Example 8 further studies are needed to continue the development of CAM2043

[000250] Example 10 aims to further investigate the safety and tolerability of CAM2043 when subjects are given repeated, once-weekly, SC injections of either ascending or steady doses of CAM2043, and to evaluate the impact on tolerability following repeated CAM2043 administration within a limited injection site area selected either in the buttock (right or left upper outer quadrant) or in the abdomen. With the present trial, the Sponsor aims to gather more knowledge on the repeat-dose regime, in particular after 4 SC doses 2.5 mg CAM2043 (or with a dose increase as shown below) given once weekly either in the buttock or the abdomen with the injection given in close proximity to each other

[000251] The investigation could be done as a parallel arm, open-label trial designed to characterize the safety and tolerability of CAM2043 when subjects are given repeated, once-weekly subcutaneous (SC) injections at ascending or steady doses of CAM2043. Healthy volunteers aged 18 to 65 (inclusive) will be included.

[000252] The estimated duration of the trial for each subject is 11 weeks, including a 2-week screening period:

• Screening Period: <2 weeks

• Treatment Period: 4 weeks

• Follow-up Period: 4 weeks

[000253] 24 subjects will be included in the trial and randomized to 2 dosing treatment schedules and, within each dosing schedules, to 2 different injection site locations, in a 1 : 1 : 1 : 1 ratio (6 subjects each). Randomized subjects will receive SC injections of either ascending (treatment arm A) or steady (treatment arm B) doses of CAM2043 limited to a pre-specified injection area in the abdomen (cohorts Al or Bl) or in the upper outer quadrant the buttock (either right or left) (cohorts A2 or B2). [000254] The trial design is visualized in Fig. 11.

[000255] Itemized list of Embodiments

[000256] El. A pre-formulation comprising: a) diacyl glycerol; b) phospholipid; c) ethanol and optionally propylene glycol; and d) treprostinil (TPN) or a salt thereof; wherein the pre-formulation is administered to a patient in need thereof in a dose of 1.5 mg/week to 15 mg/week.

[000257] E2. The pre-formulation of El, wherein component d) is treprostinil sodium (TPN(Na)).

[000258] E3. The pre-formulation of El or E2, wherein the TPN or salt thereof is the sole active agent in the pre-formulation.

[000259] E4. The pre-formulation of E4, wherein the pre-formulation is administered once every 7 (±1) days, twice every 14 (±2) days, four times every 28 (±3) days, eight times every 56 (±4) days, sixteen times every 112 (±5) days, or 32 times every 224 (±6) days.

[000260] E5. The pre-formulation of any one of E1-E4, wherein the treprostinil or salt thereof is administered at a level of 2.0 to 12.5 mg/week, such as 2.5 to 10 mg/week or 2.5 to 7.5 mg/week.

[000261] E6. The pre-formulation of any one of claims 1-5, comprising 10 mg/mL of component d) based on treprostinil free acid.

[000262] E7. The pre-formulation of any one of E1-E6, wherein component a) is glycerol dioleate.

[000263] E8. The pre-formulation of any one of E1-E7, comprising 20 to 80 wt.% component a), such as 35 to 55%, such as 38 to 52%, such as 40 to 50%.

[000264] E9. The pre-formulation of any one of E1-E8, wherein component b) comprises or consists of a phosphatidyl choline (PC), a phosphatidyl ethanolamine (PE) or a phosphatidyl inositol (PI), most preferably PC.

[000265] E10. The pre-formulation of any one of E1-E9, wherein component b) is PC.

[000266] El l. The pre-formulation of any one of E1-E10, comprising 30 to 60 wt.% component b), such as 35 to 55%, such as 38 to 52%, such as 40 to 50%. [000267] E12. The pre-formulation of any one of El-El l, wherein component c) comprises or consists of a mono-alcoholic solvent, preferably ethanol.

[000268] E13. The pre-formulation of any one of E1-E12, wherein component c) comprises or consists of ethanol, and optionally propylene glycol.

[000269] E14. The pre-formulation of any one of E1-E12, wherein component c) comprises or consists of ethanol or mixtures of ethanol and propylene glycol, wherein the ratio of ethanol to PG is 1 : 1 to 10: 1, such as 1.5: 1 to 8: 1, such as 2: 1 to 5: 1 (e.g., around 3: 1).

[000270] E15. The pre-formulation of any one of E1-E14, wherein component c) is present at a level of 1 to 30% by weight, such as 2 to 20% by weight, especially 5 to 15% by weight.

[000271] E16. The pre-formulation of any one of E1-E15, wherein the ratio of components a:b is in the range of 40:60 to 60:40, especially in the range of 45:55 to 55:45.

[000272] E17. The pre-formulation of any one of E1-E16, having a stability of at least 96%, preferably at least 97%, especially at least 98% in terms of treprostinil assay as measured by HPLC after storage at 25°C and 60% RH, for 6 months, especially after 12 months.

[000273] E18. The pre-formulation of any one of E1-E17, having a viscosity of 100 to 700 mPas at 20°C.

[000274] E19. The pre-formulation of El, wherein: component a) comprises or consists of GDO; component b) comprises or consists of PC; component c) is ethanol; and component d) is TPN or a salt thereof.

[000275] E20. The pre-formulation of E19, further comprising a co-solvent selected from the group consisting of PG, DMSO, and or NMP, wherein the ratio of ethanol : co-solvent is in the range of 30:70 to 70:30 (w/w).

[000276] E21. The pre-formulation of E19 or E20, wherein component d) is treprostinil sodium TPN(Na).

[000277] E22. The pre-formulation of E1-E21 for treatment of Raynaud's phenomena, optionally secondary to lupus, scleroderma, Buerger disease, Sjogren syndrome, rheumatoid arthritis, polymyositis, blood disorders, thyroid disorders, or pulmonary hypertension, and wherein a subject is administered a dose of 2.0 mg/week - 10 mg/week as a single dose.

[000278] E23. The pre-formulation of E1-E22 for treatment of Raynaud's phenomena, optionally secondary to lupus, scleroderma, Buerger disease, Sjogren syndrome, rheumatoid arthritis, polymyositis, blood disorders, thyroid disorders, or pulmonary hypertension, and wherein a subject is administered a dose of 2.0 mg/week - 10 mg/week as a repeated weekly dose.

[000279] E24. The pre-formulation of E1-E23, wherein the dose is 2.5 mg/week, 5.0 mg/week, or 7.5 mg/week.

[000280] E25. The pre-formulation of E1-E24, wherein the dose is administered in the buttocks and/or abdomen.

[000281] E26. The pre-formulation of E1-E25, wherein each dose is administered in the abdomen.

[000282] E27. The pre-formulation of E1-E26, wherein each dose is administered in the buttocks.

[000283] E28. Use of the pre-formulation of any one of E1-E27 in the sustained administration of treprostinil.

[000284] E29. A medicament comprising the pre-formulation of any one of E1-E27 for use as a medicament.

[000285] E30. A method for the treatment of a human or non-human mammalian subject comprising administering to the subject the pre-formulation of any one of El- E28.

[000286] E31. The method of E30 for the treatment of a human or non-human mammalian subject in need thereof to treat at least one condition selected from pulmonary artery hypertension (PAH), severe PAH, Raynaud’s disease or phenomenon, ischemia, and related conditions.

[000287] E32. The method of E31 for the treatment of a human or non-human mammalian subject in need thereof to treat Raynaud’s disease or Raynaud’s phenomenon.

[000288] E33. The method of E32 for the treatment of a human or non-human mammalian subject in need thereof to treat Raynaud’s phenomenon secondary to lupus, scleroderma, Buerger disease, Sjogren syndrome, rheumatoid arthritis, polymyositis, blood disorders, thyroid disorders, or pulmonary hypertension.

[000289] E34. The method of E33 for the treatment of a human or non-human mammalian subject in need thereof to treat Raynaud’s phenomenon secondary to scleroderma.

[000290] E35. The method of any one of E27-E34, wherein the administration route is subcutaneous injection.

[000291] E36. The method of E30-E35, wherein the dose is 2.5 mg/week, 5.0 mg/week, or 7.5 mg/week.

[000292] E37. The method of E30-E35, wherein the dose is administered in the buttocks and/or abdomen.

[000293] E38. The method of E30-E35, wherein each dose is administered in the abdomen.

[000294] E39. The method of E30-E35, wherein each dose is administered in the buttocks.

[000295] E40. Use of the pre-formulation of any one of E1-E27 in the manufacture of a medicament for use in the in vivo formation of a depot for treatment of at least one condition selected from pulmonary artery hypertension (PAH), severe PAH, Raynaud’s disease or phenomenon, ischemia, and related conditions.

[000296] E41. The use of E40 in the manufacture of a medicament for use in the in vivo formation of a depot for treatment of Raynaud’s disease or phenomenon.

[000297] E42. The use of E41 in the manufacture of a medicament for use in vivo formation of a depot for treatment of Raynaud’s phenomenon secondary to lupus, scleroderma, Buerger disease, Sjogren syndrome, rheumatoid arthritis, polymyositis, blood disorders, thyroid disorders, or pulmonary hypertension.

[000298] E43. The use of E42 in the manufacture of a medicament for use in vivo formation of a depot for treatment of Raynaud’s phenomenon secondary to scleroderma.

[000299] E44. A pre-filled administration device containing the pre-formulation of any one of E1-E27.

[000300] E45. The pre-filled administration device of E44 comprising an injector comprising a needle having a thickness of 23G or less. [000301] E46. The pre-filled administration device of E44 or E45 comprising < 1 mL, of the pre-formulation of any one of E1-E24.

[000302] E47. The pre-filled administration device of any one of E44-E46, wherein the pre-filled administration device is ready -to-use.

[000303] A kit comprising the administration device of any one of E44-E47.

[000304] INCORPORATION BY REFERENCE

[000305] All publications and patents mentioned herein, including those items listed below, are hereby incorporated by reference in their entirety for all purposes as if each individual publication or patent was specifically and individually incorporated by reference. In case of conflict, the present application, including any definitions herein, will control.

EQUIVALENTS

[000306] While specific embodiments of the subject disclosure have been discussed, the above specification is illustrative and not restrictive. Many variations of the disclosure will become apparent to those skilled in the art upon review of this specification. The full scope of the disclosure should be determined by reference to the claims, along with their full scope of equivalents, and the specification, along with such variations.

[000307] Unless otherwise indicated, all numbers expressing quantities of ingredients, reaction conditions, and so forth used in the specification and claims are to be understood as being modified in all instances by the term “about.” Accordingly, unless indicated to the contrary, the numerical parameters set forth in this specification and attached claims are approximations that may vary depending upon the desired properties sought to be obtained by the present disclosure.

Claims

1. A pre-formulation comprising: a) diacyl glycerol; b) phospholipid; c) ethanol and optionally propylene glycol; and d) treprostinil (TPN) or a salt thereof; wherein the pre-formulation is administered to a patient in need thereof in a dose of 1.5 mg/week to 15 mg/week.

2. The pre-formulation of claim 1, wherein component d) is treprostinil sodium (TPN(Na)).

3. The pre-formulation of claim 1 or claim 2, wherein the TPN or salt thereof is the sole active agent in the pre-formulation.

4. The pre-formulation of claim 1, wherein the pre-formulation is administered once every 7 (±1) days, twice every 14 (±2) days, four times every 28 (±3) days, eight times every 56 (±4) days, sixteen times every 112 (±5) days, or 32 times every 224 (±6) days.

5. The pre-formulation of any one of claims 1-4, wherein the treprostinil or salt thereof is administered at a level of 2.0 to 12.5 mg/week, such as 2.5 to 10 mg/week or 2.5 to 7.5 mg/week.

6. The pre-formulation of any one of claims 1-5, comprising 10 mg/mL of component d) based on treprostinil free acid.

7. The pre-formulation of any one of claims 1-6, wherein component a) is glycerol dioleate.

8. The pre-formulation of any one of claims 1-7, comprising 20 to 80 wt.% component a), such as 35 to 55%, such as 38 to 52%, such as 40 to 50%.

9. The pre-formulation of any one of claims 1-8, wherein component b) comprises or consists of a phosphatidyl choline (PC), a phosphatidyl ethanolamine (PE) or a phosphatidyl inositol (PI), most preferably PC.

10. The pre-formulation of any one of claims 1-8, wherein component b) is PC.

11. The pre-formulation of any one of claims 1-10, comprising 30 to 60 wt.% component b), such as 35 to 55%, such as 38 to 52%, such as 40 to 50%.

12. The pre-formulation of any one of claims 1-11, wherein component c) comprises or consists of a mono-alcoholic solvent, preferably ethanol.

13. The pre-formulation of any one of claims 1-12, wherein component c) comprises or consists of ethanol, and optionally propylene glycol.

14. The pre-formulation of any one of claims 1-11, wherein component c) comprises or consists of ethanol or mixtures of ethanol and propylene glycol, wherein the ratio of ethanol to PG is 1 : 1 to 10: 1, such as 1.5: 1 to 8: 1, such as 2: 1 to 5: 1 (e.g., around 3: 1).

15. The pre-formulation of any one of claims 1-14, wherein component c) is present at a level of 1 to 30% by weight, such as 2 to 20% by weight, especially 5 to 15% by weight.

16. The pre-formulation of any one of claims 1-15, wherein the ratio of components a:b is in the range of 40:60 to 60:40, especially in the range of 45:55 to 55:45.

17. The pre-formulation of any one of claims 1-16, having a stability of at least 96%, preferably at least 97%, especially at least 98% in terms of treprostinil assay as measured by HPLC after storage at 25°C and 60% RH, for 6 months, especially after 12 months.

18. The pre-formulation of any one of claims 1-17, having a viscosity of 100 to 700 mPas at 20°C.

19. The pre-formulation of claim 1, wherein: component a) comprises or consists of GDO; component b) comprises or consists of PC; component c) is ethanol; and component d) is TPN or a salt thereof.

20. The pre-formulation of claim 19, further comprising a co-solvent selected from the group consisting of PG, DMSO, and or NMP, wherein the ratio of ethanol : co-solvent is in the range of 30:70 to 70:30 (w/w).

21. The pre-formulation of claim 19 or claim 20, wherein component d) is treprostinil sodium TPN(Na).

22. The pre-formulation of claim 1-21 for treatment of Raynaud's phenomena, optionally secondary to lupus, scleroderma, Buerger disease, Sjogren syndrome, rheumatoid arthritis, polymyositis, blood disorders, thyroid disorders, or pulmonary hypertension, and wherein a subject is administered a dose of 2.0 mg/week - 10 mg/week as a single dose.

23. The pre-formulation of claim 1-21 for treatment of Raynaud's phenomena, optionally secondary to lupus, scleroderma, Buerger disease, Sjogren syndrome, rheumatoid arthritis, polymyositis, blood disorders, thyroid disorders, or pulmonary hypertension, and wherein a subject is administered a dose of 2.0 mg/week - 10 mg/week as a repeated weekly dose.

24. The pre-formulation of claims 1-23, wherein the dose is 2.5 mg/week, 5.0 mg/week, or 7.5 mg/week.

25. The pre-formulation of claims 1-24, wherein the dose is administered in the buttocks and/or abdomen.

26. The pre-formulation of claims 1-24, wherein each dose is administered in the abdomen.

27. The pre-formulation of claims 1-24, wherein each dose is administered in the buttocks.

28. Use of the pre-formulation of any one of claims 1-27 in the sustained administration of treprostinil.

29. A medicament comprising the pre-formulation of any one of claims 1-27 for use as a medicament.

30. A method for the treatment of a human or non-human mammalian subject comprising administering to the subject the pre-formulation of any one of claims 1- 27.

31. The method of claim 30 for the treatment of a human or non-human mammalian subject in need thereof to treat at least one condition selected from pulmonary artery hypertension (PAH), severe PAH, Raynaud’s disease or phenomenon, ischemia, and related conditions.

32. The method of claim 31 for the treatment of a human or non-human mammalian subject in need thereof to treat Raynaud’s disease or Raynaud’s phenomenon.

33. The method of claim 32 for the treatment of a human or non-human mammalian subject in need thereof to treat Raynaud’s phenomenon secondary to lupus, scleroderma, Buerger disease, Sjogren syndrome, rheumatoid arthritis, polymyositis, blood disorders, thyroid disorders, or pulmonary hypertension.

34. The method of claim 33 for the treatment of a human or non-human mammalian subject in need thereof to treat Raynaud’s phenomenon secondary to scleroderma.

35. The method of any one of claims 27-34, wherein the administration route is subcutaneous injection.

36. The method of claims 30-35, wherein the dose is 2.5 mg/week, 5.0 mg/week, or 7.5 mg/week.

37. The method of claims 30-35, wherein the dose is administered in the buttocks and/or abdomen.

38. The method of claims 30-35, wherein each dose is administered in the abdomen.

39. The method of claims 30-35, wherein each dose is administered in the buttocks.

40. Use of the pre-formulation of any one of claims 1-27 in the manufacture of a medicament for use in the in vivo formation of a depot for treatment of at least one condition selected from pulmonary artery hypertension (PAH), severe PAH, Raynaud’s disease or phenomenon, ischemia, and related conditions.

41. The use of claim 40 in the manufacture of a medicament for use in the in vivo formation of a depot for treatment of Raynaud’s disease or phenomenon.

42. The use of claim 41 in the manufacture of a medicament for use in vivo formation of a depot for treatment of Raynaud’s phenomenon secondary to lupus, scleroderma, Buerger disease, Sjogren syndrome, rheumatoid arthritis, polymyositis, blood disorders, thyroid disorders, or pulmonary hypertension.

43. The use of claim 42 in the manufacture of a medicament for use in vivo formation of a depot for treatment of Raynaud’s phenomenon secondary to scleroderma.

44. A pre-filled administration device containing the pre-formulation of any one of claims 1-27.

45. The pre-filled administration device of claim 44 comprising an injector comprising a needle having a thickness of 23G or less.

46. The pre-filled administration device of claim 44 or claim 45 comprising < 1 mL, of the pre-formulation of any one of claims 1-24.

47. The pre-filled administration device of any one of claims 44-46, wherein the pre-filled administration device is ready-to-use.

48. A kit comprising the administration device of any one of claims 44-47.