Crystalline forms of NRTTI compounds

Abstract

The present disclosure provides anhydrous crystalline forms 1 and 4 of 4'-ethynyl-2-fluoro-2'-deoxyadenosine and pharmaceutical compositions thereof, each of which can be used to inhibit HIV reverse transcriptase, treat or prevent HIV infection and / or treat, prevent and / or delay the onset or progression of AIDS or ARC.

Description

Background of the Invention

[0002] Human immunodeficiency virus (HIV-1) infection is a serious condition that, if not treated, will eventually destroy the host's immune system to cause acquired immunodeficiency syndrome (AIDS) and premature death. Despite progress in antiretroviral therapy (ART), HIV remains a global epidemic and a global public health priority. In 2012, it was estimated that 35 million people in the world were infected with HIV (Global Report: UNAIDS report on the global AIDSepidemic2013.UNAIDS / JC2502 / 1 / E). In the United States, it is estimated that 1.2 million people are infected with HIV and about 50,000 people are newly infected each year. HIV seropositive individuals are initially asymptomatic, but usually develop AIDS-related syndrome (ARC), followed by AIDS. More than 650,000 people have died of AIDS in the United States and more than 14,000 new deaths are reported each year. Treatment can help HIV patients live longer, healthier lives, but currently only 30% of HIV patients in the United States successfully control their virus (Center for Disease Control and Prevention. Today's HIV / AIDS epidemic. July 2015).

[0003] Nucleoside and nucleotide reverse transcriptase inhibitors (NsRTI and NtRTI, or collectively referred to as NRTI) inhibit HIV reverse transcriptase and block HIV replication. They are one of the 6 classes of HIV antiretrovirals (ARVs) used as components of potent and long-lasting multidrug regimens, which typically combine two NRTIs with non-nucleoside reverse transcriptase inhibitors, integrase strand transfer inhibitors, or protease inhibitors. Combination therapy maximizes treatment response and minimizes the emergence of drug resistance.

[0004] Due to the fact that HIV replication is asynchronous, antiretroviral agents need to be continuously present in the patient's body to effectively suppress viremia. For most types of drugs, including protease inhibitors, integrase inhibitors and non-nucleoside reverse transcriptase inhibitors, efficacy depends on circulating drug concentrations and the purpose of administration is to provide a circulating drug concentration (i.e., Cmin) that exceeds the concentration required for inhibiting viral replication (i.e., IC50 or IC95) throughout the dosing interval. On the contrary, after entering the cell, NRTIs and nucleotide reverse transcriptase inhibitors (NtRTIs, such as tenofovir) enter the dedicated intracellular anabolic pathway to be converted into active phosphorylated forms, and it is their intracellular half-life rather than their plasma concentration that determines their sustained effect. Currently approved NRTIs are administered at least once a day.

[0005] 4'-Ethynyl-2-fluoro-2'-deoxyadenosine (EFdA, also known as MK-8591) is a nucleoside reverse transcriptase translocation inhibitor (NRTTI) that blocks HIV-1 and SIV viral replication in vitro (Kawamoto, A., Kodama, E., Sarafianos SF et al., Int. J. Biochem. Cell Biol.; 40(11):2410-20

[2008] ; Ohrui, H., Kohgo, S., Hayakawa, H. et al., Nucleosides, Nucleotides & Nucleic Acids, 26, 1543-1546

[2007] ) and in vivo (Hattori, S., Ide, K., Nakata, H. et al. Antimicrobial. Agents and Chemotherapy, 53, 3887-3893

[2009] ).

[0006] U.S. Patent No. 7,339,053 describes EFdA (referred to as 2'-deoxy-4'-C-ethynyl-2-fluoroadenosine in the '053 patent) and a synthetic method for preparing EFdA. EFdA has the following chemical structure:

[0007]

[0008] US Patent No. 7,339,053 describes the use of water as the final crystallization solvent in a synthetic process for preparing EFdA, which is understood to be for the preparation of the monohydrate crystalline form of EFdA.

[0009] EFdA is metabolized in cells to their active triphosphate anabolic metabolites, which inhibit HIV reverse transcriptase. Unlike NRTIs currently used to treat HIV infection that do not contain a 3'-OH group to block the incorporation of incoming nucleotides, EFdA retains the 3'-OH group and acts as a chain terminator by preventing primer: template translocation in the reverse transcriptase (RT) active site and preventing the binding of incoming deoxyribonucleotide triphosphates (dNTPs). In addition, it is believed that the pucker of the modified ribose ring of EFdA helps inhibit reverse transcriptase by placing the 3'-OH in the carrier, where phosphate transfer from incoming nucleotides is ineffective. (Michailidis E et al., Mechanism of inhibition of HIV-1reversetranscriptase by 4'-ethynyl-2-fluoro-2'-deoxyadenosine triphosphate, J BiolChem 284:35681–35691

[2009] ; Michailidis E et al., 4'-Ethynyl-2-fluoro-2'-deoxyadenosine(EFdA)inhibits HIV-1reverse transcriptase with multiplemechanisms,J Biol Chem 289:24533–24548

[2014] ).

[0010] In an in vitro HIV replication assay, EFdA is a potent antiretroviral drug and exhibits considerable antiviral activity against clinical isolates of all subtypes evaluated. It is rapidly anabolized to an active triphosphate in both lymphoid cell lines and peripheral blood mononuclear cells in vitro, and the intracellular half-life of EFdA triphosphate (EFdA-TP) exceeds 72 hours (Stoddart, CA, Galkina et al., Oral Administration of the Nucleoside EFdA (4'-Ethynyl-2-Fluoro-2'-Deoxyadenosine) Provides Rapid Suppression of HIV Viremia in Humanized Mice and Favorable Pharmacokinetic Properties in Mice and the Rhesus Macaque, Antimicrob Agents Chemother, 2015 Jul; 59(7): 4190–4198, published online on May 4, 2015).

[0011] Currently available drug therapies for HIV infection work in combination to suppress viremia, thereby keeping the virus under control. HIV drug therapy is lifelong, and strict adherence to the treatment regimen is critical to maintaining viral suppression, reducing the risk of drug resistance, and minimizing the risk of transmission. Effective and safe well-tolerated drugs that are easily taken with low dosing frequencies have the potential to improve patient compliance and long-term treatment success. For the prevention of HIV infection, the only currently available pre-exposure prophylaxis (PrEP) therapy approved by the U.S. Food and Drug Administration is for the prevention of HIV infection in uninfected humans. (Emtricitabine / Tenofovir DF).

[0012] Currently available oral antiretroviral drugs are administered once a day. Due to the need for sustained circulating drug concentrations, the use of long-acting release drug delivery methods such as implants is ideal. Less frequent dosing may help alleviate the practical challenges and cumulative psychological impact of daily HIV drug therapy. Long-acting antiretroviral therapy may help patients return to a more normal sense and provide flexibility that can affect their lives, work, travel, get along with others and the way they look at themselves. In addition, some patients are suitable for and may prefer weekly, monthly or longer interval dosing options, such as provided by long-acting parenteral (LAP) administration, which can lead to improved medication compliance.

[0013] It would be valuable to provide additional treatment options for HIV-infected people or people at risk of HIV infection that can be administered less frequently than daily dosing, such as long-acting parenteral (LAP) implantable formulations. For LAP implantable drug formulations, the most thermodynamically stable crystalline form of a small molecule agent is desired to avoid physical form transformation during storage and / or shelf life, which may alter the in vivo performance and efficacy of the drug (see, e.g., Chemburkar et al., Organic Process Research & Development 2000, 4, 413-417). However, when the known monohydrate crystalline form of EFdA is utilized, in-situ recrystallization occurs during the hot melt extrusion (HME) formulation process, which results in the conversion of the monohydrate crystalline form of EFdA into a mixture of multiple thermodynamically unfavorable anhydrous phases in the drug product. SUMMARY OF THE INVENTION

[0015] The present application discloses the discovery of anhydrous crystalline forms of EFdA, namely, anhydrous crystalline Form 1 of EFdA and anhydrous crystalline Form 4 of EFdA, which possess the necessary physical stability for LAP implantable drug formulations of EFdA.

[0016] The present disclosure also provides methods for inhibiting HIV reverse transcriptase, treating HIV or preventing HIV infection and / or treating, preventing and / or delaying the onset or progression of AIDS or ARC using anhydrous crystalline form 1 or form 4 of EFdA. The present disclosure further provides pharmaceutical compositions of each of the anhydrous crystalline form 1 and form 4 of EFdA, and methods of using each of the crystalline forms. Further embodiments include, but are not limited to, procedures for preparing each of the anhydrous crystalline forms 1 and 4 of EFdA. BRIEF DESCRIPTION OF THE DRAWINGS

[0018] Figure 1 is a graph of a powder X-ray diffraction ("PXRD") pattern of anhydrous crystalline Form 1 of EFdA generated using the apparatus and methods described herein. The graph plots peak intensity as defined by counts per second versus diffraction angle 2θ (2Θ) in degrees.

[0019] Figure 2 Depicted is the solid state of the anhydrous crystalline form 1 of EFdA 19 F NMR (nuclear magnetic resonance) spectrum.

[0020] Figure 3 is a thermogravimetric analysis ("TGA") graph of anhydrous crystalline Form 1 of EFdA. The graph plots weight (%) versus temperature (°C).

[0021] Figure 4 is a graph of a powder X-ray diffraction ("PXRD") pattern of anhydrous crystalline Form 2 of EFdA generated using the equipment and methods described herein. The graph plots peak intensity as defined by counts per second versus diffraction angle 2θ (2Θ) in degrees.

[0022] Figure 5 Depicted is the solid state of anhydrous crystalline form 2 of EFdA 19 F NMR (nuclear magnetic resonance) spectrum.

[0023] Figure 6 is a graph of a powder X-ray diffraction ("PXRD") pattern of anhydrous crystalline Form 3 of EFdA generated using the equipment and methods described herein. The graph plots peak intensity as defined by counts per second versus diffraction angle 2θ (2Θ) in degrees.

[0024] Figure 7 is a graph of a powder X-ray diffraction ("PXRD") pattern of anhydrous crystalline Form 4 of EFdA generated using the equipment and methods described herein. The graph plots peak intensity as defined by counts per second versus diffraction angle 2θ (2Θ) in degrees.

[0025] Figure 8 Depicted is the solid state of anhydrous crystalline form 4 of EFdA 19F NMR (nuclear magnetic resonance) spectrum.

[0026] Fig. 9 is a thermogravimetric analysis ("TGA") graph of anhydrous crystalline Form 4 of EFdA. The graph plots weight (%) versus temperature (°C).

[0027] Fig.10 is a graph of a powder X-ray diffraction ("PXRD") pattern of the monohydrate crystalline form MH of EFdA generated using the apparatus and methods described herein. The graph plots peak intensity as defined by counts per second versus the diffraction angle 2θ (2Θ) in degrees.

[0028] Fig.11 Depicted is the solid state of the monohydrate crystalline form MH of EFdA 19 F NMR (nuclear magnetic resonance) spectrum.

[0029] Fig.12 is a graph of an X-ray diffraction ("XRD") pattern of a composition of crystalline Form MH, monohydrate of EFdA and EVA polymer after being processed by hot melt extrusion using HME low shear process A such that Form MH in the composition is converted to anhydrous crystalline Form 1 of EFdA and anhydrous crystalline Form 2 of EFdA.

[0030] Fig.13 is an X-ray diffraction ("XRD") pattern of a composition of crystalline Form MH, monohydrate of EFdA and EVA polymer after being processed by hot melt extrusion using HME high shear process B such that Form MH in the composition is converted to anhydrous crystalline Form 4 of EFdA.

[0031] Fig.14 is a graph of an X-ray diffraction ("XRD") pattern of a composition of anhydrous crystalline Form 1 of EFdA and EVA polymer after being processed by hot melt extrusion using HME low shear process A so as to maintain anhydrous crystalline Form 1 of EFdA in the composition.

[0032] Fig.15 is a graphical representation of an X-ray diffraction ("XRD") pattern of a composition of anhydrous crystalline Form 4 of EFdA, EVA polymer and BaSO4 after being processed by hot melt extrusion using HME low shear process A so that anhydrous crystalline Form 4 of EFdA is retained.

[0033] Fig.16 is a graph depicting the solubility of anhydrous crystalline Forms 1, 2, and 4 of EFdA in acetonitrile as a function of temperature and the calculated ΔΔ Gibbs free energy (G) between anhydrous crystalline Forms 1 and 4 of EFdA. DETAILED DESCRIPTION OF THE INVENTION

[0035] The terms used herein have their ordinary meanings and the meanings of these terms are independent at each location. Nevertheless and unless otherwise indicated, the following definitions apply throughout this specification and claims.

[0036] “API” means active pharmaceutical ingredient.

[0037] “Figure” may be abbreviated as FIG., Fig or fig. and refers to the corresponding drawing.

[0038] "Patient" or "subject" includes humans and other mammals.

[0039] "Mammal" includes humans and other mammals.

[0040] "PXRD" means powder x-ray diffraction.

[0041] "TGA" means thermogravimetric analysis.

[0042] "LAP" refers to long acting parenteral.

[0043] "L / D" refers to barrel length / screw diameter.

[0044] "s" stands for seconds.

[0045] "Excipient" refers to an essentially inert substance used as a diluent or to impart form or consistency to a formulation.

[0046] Anhydrous crystalline Form 1 of EFdA may also be referred to herein as "anhydrous crystalline Form 1," "crystalline Form 1," "anhydrous Form 1," or "Form 1."

[0047] Anhydrous crystalline Form 2 of EFdA may also be referred to herein as "anhydrous crystalline Form 2," "crystalline Form 2," "anhydrous Form 2," or "Form 2."

[0048] Anhydrous crystalline Form 3 of EFdA may also be referred to herein as "anhydrous crystalline Form 3," "crystalline Form 3," "anhydrous Form 3," or "Form 3."

[0049] Anhydrous crystalline Form 4 of EFdA may also be referred to herein as "anhydrous crystalline Form 4," "crystalline Form 4," "anhydrous Form 4," or "Form 4."

[0050] The monohydrate crystalline Form MH of EFdA may also be referred to herein as "monohydrate crystalline Form MH", "crystalline Form MH", "monohydrate Form MH" or "Form MH".

[0051] The present disclosure provides pharmaceutically acceptable compositions of each of the anhydrous crystalline Form 1 and Form 4 of EFdA.

[0052] The term "composition" (or "pharmaceutical composition" or "pharmaceutically acceptable composition") as used herein is intended to include a product comprising the specified ingredients and, when applicable, the specified amounts, as well as any product resulting directly or indirectly from combining the specified ingredients. The term is intended to include a product comprising one or more active ingredients and one or more inert ingredients (if any) constituting the carrier, as well as any product resulting directly or indirectly from the combination, complexation or aggregation of any two or more ingredients or from the dissociation of one or more ingredients or from other types of reactions or interactions of one or more ingredients. The carrier, which may include a diluent and / or an excipient, may be any one or more inert ingredients suitable for the mode of delivery of the active pharmaceutical ingredient, such as any one or more inert ingredients for oral or parenteral administration, including but not limited to polymers suitable for implantable pharmaceutical formulations. Accordingly, the pharmaceutical compositions of the present disclosure include any composition made by mixing Form 1 with Form 2, Form 3 or Form 4 or mixing any mixture of the crystalline forms and a pharmaceutically acceptable carrier. "Pharmaceutically acceptable" means that the carrier must be compatible with the other ingredients of the formulation and not harmful to the recipient thereof.

[0053] The term "composition" (or "pharmaceutical composition" or "pharmaceutically acceptable composition") as used herein is also intended to include bulk compositions and / or individual dosage units. Bulk compositions are materials that have not yet been formed into individual dosage units. Bulk compositions and each individual dosage unit may contain a fixed amount of active agent. Non-limiting examples of dosage units include oral dosage units, such as tablets, pills, etc., and parenteral dosage unit formulations, such as implantable dosage units. Similarly, the methods described herein for treating patients by administering the pharmaceutical compositions of the present disclosure are also intended to include administering the above-mentioned bulk compositions and individual dosage units.

[0054] The present disclosure includes methods for inhibiting HIV reverse transcriptase, treating HIV or preventing HIV infection and / or treating, preventing and / or delaying the onset or progression of AIDS or ARC, comprising administering to a subject an effective amount of anhydrous crystalline Form 1 or Form 4 of EFdA. The present disclosure further provides methods for using each of the anhydrous crystalline Forms 1 and 4 of EFdA (1) in the preparation of a medicament useful (alone or with another active ingredient) for inhibiting HIV reverse transcriptase, treating HIV or preventing HIV infection and / or treating, preventing and / or delaying the onset or progression of AIDS or ARC; and (2) in methods for inhibiting HIV reverse transcriptase, treating HIV or preventing HIV infection and / or treating, preventing and / or delaying the onset or progression of AIDS or ARC. Further embodiments include, but are not limited to, procedures for preparing each of the crystalline Forms 1, 2, 3 and 4 of EFdA.

[0055] The discovery of anhydrous crystalline Form 4 of EFdA was unusual and unexpected due to the fact that Form 4 was not found in standard drug crystal polymorph screens, but was instead discovered during a study dedicated to investigating a range of processing conditions for preparing implantable drug formulations as a result of specific processing conditions during the HME preparation of such formulations.

[0056] In the following isolation procedure, EFdA monohydrate (Form MH) powder was prepared by organic synthesis and its conversion to anhydrous phase was studied under various conditions. The monohydrate and anhydrous forms were also studied in the preparation of long-acting parenteral HME.

[0057] Isolation of anhydrous crystalline forms 1, 2 and 4 of EFdA by hot melt extrusion (HME) processing: Anhydrous crystalline forms 1, 2 and 4 of EFdA were isolated by hot melt extrusion (HME) processing. Micronized ethylene vinyl acetate (EVA) polymer and monohydrate crystalline form MH of EFdA were blended using a Turbula T2F mixer at various ratios: 30, 35, 40, 45 and 50 wt% drug. The premix was hot melt extruded using an 18 mm Leistritz twin screw extruder at 25:1 L / D, 400 g / hr throughput, 100-140°C temperature, 40°C feed zone and 30 rpm screw speed. Two screw configurations were used, both consisting mainly of conveying elements with different mixing sections. The milder screw design contained a mixing section consisting of conveying kneading elements each 15 mm long with twist angles of 30°, 60° and 60° ("HME low shear process A"). The more aggressive screw design included an additional mixing zone after the mixing section used in the milder design: conveying kneading elements, each 15 mm long, with a twist angle of 90° ("HME High Shear Process B"). For both extrusion arrangements, the strands were subsequently air cooled and pelletized to form micropellets. The pellets were then extruded using a 1 / 2" American Kuhne single screw extruder at a temperature of 110-140°C, a feed zone of 25°C and a screw speed of 20-25 rpm to form filaments of 2±0.05 mm diameter, which were then cut into lengths of 40±2 mm.

[0058] Extrusion under milder HME low shear process A conditions resulted in the formation of a phase mixture of anhydrous Forms 1 and 2 in the extruded product. Extrusion under more aggressive HME high shear process B conditions resulted in the formation of anhydrous Form 4 in the extruded product. Thermodynamically stable anhydrous Form 4 was formed under aggressive HME high shear process B processing conditions of implant drug products and subsequent analytical identification indicated the first discovery of this new phase.

[0059] Anhydrous crystalline forms 1, 2, 3 and 4 of EFdA were isolated by solid powder differential scanning calorimetry (DSC) thermal processing: Anhydrous forms 1, 2, 3 and 4 were isolated by DSC processing. DSC experiments were performed using TA Instruments Q2000 with monohydrate form MH bulk compositions in unsealed and hermetically sealed containers. For samples heated in unsealed DSC pans, 5-10 mg of sample was placed in an aluminum pan, which was then covered without sealing the container. For hermetically sealed samples, 5-10 mg was placed in an aluminum pan, which was then sealed with a sealed lid. Experiments were performed normally under non-modulated DSC conditions. Experiments were performed under nitrogen flow (50 mL / min).

[0060] By thermal processing in an unsealed container, anhydrous Form 3 was generated by heating to 95° C. Anhydrous Form 3 was converted to anhydrous Form 2 by further heating to 120° C. These same phases were generated at varying heating rates (from 0.1° C. / min to 75° C. / min) in an unsealed container.

[0061] Anhydrous Form 1 or 4 was produced by thermal processing in a hermetically sealed container. The sample was heated from 20°C to 150°C. At a heating rate of 1°C / min, anhydrous Form 4 was produced. At a heating rate of 10°C / min, anhydrous Form 1 was produced.

[0062] Physical Characterization of Anhydrous Crystalline Forms 1, 2, 3 and 4 of EFdA

[0063] X-ray diffraction (XRD) pattern studies (including powder X-ray diffraction (PXRD) pattern studies), thermogravimetric analysis (TGA), and solid-state NMR (ssNMR) are widely used to characterize molecular structure, crystallinity, and polymorphism, and were each used to characterize Forms 1, 2, 3, and 4 of EFdA as indicated. One skilled in the art will recognize that the crystalline form of a substance can be further characterized by a combination of measured PXRD values, ssNMR, and / or TGA measurements. Thus, in another aspect, crystalline Forms 1, 2, 3, 4, and MH of EFdA can each be characterized by any combination of the techniques described herein.

[0064] X-ray diffraction (XRD) of EFdA+EVA implant composite : Using a Cu Kα radiation source ( EFdA crystal phase analysis was performed by X-ray diffraction from 2-40 2θ with a scan step size of 0.0167° using a Phillips X'Pert PW3040-PRO transmission instrument (45 kV, 40 mA). The counting time for XRD of the EFdA / EVA extrudate sample was 158.750 seconds.

[0065] Powder X-ray Diffraction (PXRD) of Anhydrous Crystalline Forms 1, 2, 3, 4 and MHPowder X-ray diffraction data of anhydrous polymorphic phases of EFdA and Form MH phase were acquired on a Panalytical X-pert ProPW3040 System configured in a Bragg-Brentano configuration and equipped with a Cu radiation source (monochromatization to Kα using a Nickel filter). A fixed slit optical configuration was used for data acquisition. Data were acquired between 2 and 40° 2θ. Samples were prepared by gently pressing the powdered sample onto a shallow cavity zero background silicon holder. The counting time for powder X-ray diffraction (PXRD) of EFdA Forms 1, 2, 3, 4 and MH was 50.800 seconds using the EFdA powder sample.

[0066] Those skilled in the art will recognize that measurements of XRD peak positions for a given crystalline form of the same compound vary within margins of error. The margin of error for 2θ values ​​measured as described herein is typically + / -0.2 ° 2θ. Variability may depend on factors such as the system, method, sample, and conditions used for measurement. Also as a skilled crystallographer will recognize, the intensity of each peak reported in the figures herein may vary due to many factors, such as the directional effects of the crystal in the x-ray beam, the purity of the analyzed material, and / or the crystallinity of the sample. A skilled crystallographer will also recognize that measurements using different wavelengths will result in different displacements according to the Bragg-Brentano equation. These further XRD patterns generated using alternative wavelengths are considered to be alternative representations of the XRD patterns of the crystalline material of the present disclosure, and are therefore within the scope of the present disclosure.

[0067] Generate using the above equipment and program Figure 1 , 4 , 6, 7 and 10, respectively, for Form 1, Form 2, Form 3, Form 4 and Form MH of EFdA. The peak intensity (y-axis in counts per second) of each PXRD pattern is plotted against the 2θ angle (x-axis in ° 2θ). In addition, the data are plotted against the 2θ angle using the detector counts normalized to the collection time for each step. Fig.12 , 13 The XRD patterns in each of , 14 and 15 were generated as described above for the PXRD patterns, except that the sample used to obtain the XRD patterns was not a bulk composition but an EFdA / EVA composition in a solid dosage form (a long-acting polymer parenteral implant).

[0068] Solid State 19 FNMR (ssNMR) :All solid state 19 F nuclear magnetic resonance (NMR) spectra were acquired on a Bruker Avance III HD 9.4T spectrometer equipped with a 4.0 mm H / F / X magic angle spinning (MAS) probe. The probe was tuned to 19F / H double resonance mode of F (fluorine-19) experiment. During acquisition, the 1 H dipole decoupling and collection with a recycle delay of 60 s 19 F Direct polarization (DP) magic angle spinning (MAS) spectroscopy. For all experiments, the sample was spun at 12 kHz and kept at 294 K. 19 F, the typical pulse is 4.0us. 19 F chemical shift referenced to -122.0 ppm Teflon 19 F signal.

[0069] Using the above equipment and procedures, generate Figure 2 , 5 , 8 and 11 respectively show the EFdA form 1, form 2, form 4 and form MH of each 19 F NMR ssNMR spectra.

[0070] Thermogravimetric analysis (TGA) :

[0071] Thermogravimetric analysis (TGA) data were obtained using a Perkin Elmer model TGA 7 or equivalent. Experiments were performed to a maximum temperature of approximately 300°C under a nitrogen stream and using a heating rate of 10°C / min. After the balance was automatically tared, the appropriate amount of sample was added to a platinum pan, the furnace was raised and the heating program was started. Analysis of the results was performed by selecting the Delta Y function in the instrument software and selecting the temperature between which the weight loss was calculated. Weight loss was reported until decomposition / evaporation began.

[0072] TGA analysis was performed on Form 1 and Form 4 of EFdA using the thermogravimetric analysis equipment and procedure described above.

[0073] Anhydrous crystalline form 1 of EFdA

[0074] Anhydrous crystalline form 1 of EFdA: X-ray diffraction (PXRD) :

[0075] The PXRD pattern of Form 1 is shown in Figure 1 Therefore, in one aspect of the present disclosure, there is provided a method of Figure 1 The powder x-ray diffraction pattern shown in is characteristic of the anhydrous crystalline Form 1 of EFdA. The peak positions (on the 2θ x-axis) consistent with these profiles are shown in Table 1 (+ / - 0.2° 2θ). The positions of these PXRD peaks are characteristic of Form 1 of EFdA. Therefore, in another aspect, the anhydrous crystalline Form 1 of EFdA is characterized by a powder x-ray diffraction pattern having the respective peak positions (+ / - 0.2° 2θ) listed in Table 1.

[0076] Table 1

[0077] Thus, in one aspect, the anhydrous crystalline Form 1 of EFdA is characterized by a powder x-ray diffraction pattern having the peak positions listed in Table 1 (+ / - 0.2° 2θ).

[0078] In another aspect, the anhydrous crystalline Form 1 of EFdA is characterized by a powder x-ray diffraction pattern comprising 2 or more 2θ values ​​listed in Table 1 (+ / - 0.2° 2θ).

[0079] In another aspect, the anhydrous crystalline Form 1 of EFdA is characterized by a powder x-ray diffraction pattern comprising 3 or more 2θ values ​​listed in Table 1 (+ / - 0.2° 2θ).

[0080] In another aspect, the anhydrous crystalline Form 1 of EFdA is characterized by a powder x-ray diffraction pattern comprising 4 or more 2θ values ​​listed in Table 1 (+ / - 0.2° 2θ).

[0081] In another aspect, the anhydrous crystalline Form 1 of EFdA is characterized by a powder x-ray diffraction pattern comprising 6 or more 2θ values ​​listed in Table 1 (+ / - 0.2° 2θ).

[0082] In another aspect, the anhydrous crystalline Form 1 of EFdA is characterized by a powder x-ray diffraction pattern comprising 9 or more 2θ values ​​listed in Table 1 (+ / - 0.2° 2θ).

[0083] In another aspect, the anhydrous crystalline Form 1 of EFdA is characterized by a powder x-ray diffraction pattern comprising 12 or more 2θ values ​​listed in Table 1 (+ / - 0.2° 2θ).

[0084] In yet another aspect, the anhydrous crystalline form 1 of EFdA that best represents the EFdA of Table 1 and / or Figure 1 The PXRD peak positions shown in the table are grouped as "identification peak sets" to conveniently distinguish this crystalline form from other crystalline forms. A selection of such characteristic peaks is given in the column labeled "identification peak set" in Table 1.

[0085] Thus, in another aspect, there is provided anhydrous crystalline Form 1 of EFdA characterized by a powder x-ray diffraction pattern comprising each of the 2θ values ​​listed in Identifying Peak Set 1 in Table 1 (+ / - 0.2° 2θ).

[0086] In another aspect, an anhydrous crystalline Form 1 of EFdA is provided, characterized by a powder x-ray diffraction pattern comprising each of the 2θ values ​​listed in the Identification Peak Set 2 in Table 1 (+ / - 0.2° 2θ).

[0087] In another aspect, an anhydrous crystalline Form 1 of EFdA is provided, characterized by a powder x-ray diffraction pattern comprising each of the 2θ values ​​listed in the Identification Peak Set 3 in Table 1 (+ / - 0.2° 2θ).

[0088] In another aspect, an anhydrous crystalline Form 1 of EFdA is provided, characterized by a powder x-ray diffraction pattern comprising each of the 2θ values ​​listed in Identified Peak Set 1 and Identified Peak Set 2 in Table 1 (+ / - 0.2° 2θ).

[0089] In another aspect, an anhydrous crystalline Form 1 of EFdA is provided, characterized by a powder x-ray diffraction pattern comprising each of the 2θ values ​​listed in Identification Peak Set 1 and Identification Peak Set 3 in Table 1 (+ / - 0.2° 2θ).

[0090] In another aspect, an anhydrous crystalline Form 1 of EFdA is provided, characterized by a powder x-ray diffraction pattern comprising each of the 2θ values ​​listed in Identified Peak Set 1 and Identified Peak Set 2 and Identified Peak Set 3 in Table 1 (+ / - 0.2° 2θ).

[0091] In another aspect, the anhydrous crystalline Form 1 of EFdA is characterized by Figure 1 PXRD spectrum shown in .

[0092] In yet another aspect, the anhydrous crystalline Form 1 of EFdA is characterized by the above-mentioned PXRD characteristic peaks and / or Figure 1 The data shown in , alone or in combination with any other features of Form 1 of EFdA described herein.

[0093] Anhydrous crystalline form of EFdA 1: 19 F(Fluorine-19) Solid State NMR :

[0094] Use the above 19 F (fluorine-19) solid-state NMR equipment and procedures to obtain the solid-state form 1 of EFdA 19 F NMR spectrum. The spectrum shows Figure 2 Characteristic peaks of anhydrous crystalline Form 1 of EFdA were observed at -114.75, -117.09, and -118.92 ppm. This NMR measurement can be used alone or in combination with any other characteristics of Form 1 described herein to identify Form 1 of EFdA and distinguish it from other crystalline forms of EFdA.

[0095] Thus, in another aspect, the anhydrous crystalline Form 1 of EFdA is characterized by a solid phase having peaks at -114.75, -117.09 and -118.92 ppm. 19 F NMR spectrum.

[0096] In another aspect, the anhydrous crystalline Form 1 of EFdA is characterized by a solid state comprising any two of the following peaks19 FNMR spectrum: -114.75, -117.09 and -118.92 ppm.

[0097] In another aspect, the anhydrous crystalline Form 1 of EFdA is characterized by a solid state comprising at least the following peaks: 19 F NMR spectrum: -117.09 and -118.92 ppm.

[0098] In another aspect, the anhydrous crystalline Form 1 of EFdA is characterized by Figure 2 The solid state shown in 19 F NMR spectrum.

[0099] In yet another aspect, Form 1 of EFdA is characterized by the above-mentioned NMR characteristic peaks and / or Figure 2 The data shown in , alone or in combination with any other features of Form 1 of EFdA described herein.

[0100] Thus, in yet another aspect, Form 1 of EFdA is characterized by PXRD Peak Position Set 1 and / or PXRD Peak Position Set 2 and / or PXRD Peak Position Set 3, each as described in Table 1 above and each further characterized by:

[0101] 1) Solid with peaks at -114.75, -117.09 and -118.92 ppm 19 F NMR spectroscopy; or

[0102] 2) Solid with any two of the following peaks 19 F NMR spectrum: -114.75, -117.09 and -118.92 ppm; or

[0103] 3) Solid with peaks at -117.09 and -118.92 ppm 19 F NMR spectroscopy; or

[0104] 4) Basically Figure 2 The solid state shown in 19 F NMR spectrum.

[0105] Anhydrous crystalline form 1 of EFdA: Thermogravimetric analysis (TGA) :

[0106] Form 1 of EFdA was subjected to TGA analysis using the thermogravimetric analysis equipment and procedure described above. Figure 3A typical TGA analysis curve of Form 1 of EFdA is depicted. The data shows a 0.1 wt.% loss until 133°C, followed by thermal decomposition above 240°C. This TGA analysis can be used alone or in combination with any other characteristics of Form 1 described herein to identify Form 1 of EFdA and distinguish it from other crystalline forms of EFdA. Thus, in another aspect, Form 1 of EFdA is characterized by being substantially as Figure 3 In yet another aspect, Form 1 of EFdA is characterized by any of these TGA measurements and / or substantially as Figure 3 The TGA curve shown in FIG. 1 , alone or in combination with any one or more other features of Form 1 described herein, including various aspects of the PXRD characterization described above and / or the above description of Form 1 19 Various aspects of F ssNMR characterization.

[0107] Anhydrous crystalline form 2 of EFdA

[0108] Anhydrous crystalline form 2 of EFdA: PXRD pattern

[0109] The PXRD pattern of the anhydrous crystalline form 2 of EFdA is depicted in Figure 4 Therefore, in one aspect of the present disclosure, there is provided a method of Figure 4 The powder x-ray diffraction pattern shown in is characteristic of the anhydrous crystalline Form 2 of EFdA. The peak positions (on the 2θ x-axis) that coincide with these contours are shown in Table 2 (+ / - 0.2° 2θ). The positions of these PXRD peaks are characteristic of the anhydrous crystalline Form 2 of EFdA.

[0110] Thus, in another aspect, the anhydrous crystalline Form 2 of EFdA is characterized by a powder x-ray diffraction pattern having the peak positions listed in Table 2 (+ / - 0.2° 2θ).

[0111] Table 2

[0112]

[0113] In another aspect, the anhydrous crystalline Form 2 of EFdA is characterized by a powder x-ray diffraction pattern comprising 2 or more 2θ values ​​listed in Table 2 (+ / - 0.2° 2θ).

[0114] In another aspect, the anhydrous crystalline Form 2 of EFdA is characterized by a powder x-ray diffraction pattern comprising 3 or more 2θ values ​​listed in Table 2 (+ / - 0.2° 2θ).

[0115] In another aspect, the anhydrous crystalline Form 2 of EFdA is characterized by a powder x-ray diffraction pattern comprising 4 or more 2θ values ​​listed in Table 2 (+ / - 0.2° 2θ).

[0116] In another aspect, the anhydrous crystalline Form 2 of EFdA is characterized by a powder x-ray diffraction pattern comprising 6 or more 2θ values ​​listed in Table 2 (+ / - 0.2° 2θ).

[0117] Anhydrous crystalline form 2 of EFdA: 19 F(Fluorine-19) Solid State NMR

[0118] Use the above 19 F (fluorine-19) solid-state NMR equipment and procedures to obtain the solid state of the anhydrous crystalline form 2 of EFdA 19 FNMR spectrum of anhydrous crystalline form 2 19 F NMR spectrum shows Figure 5 Characteristic peaks of the anhydrous crystalline Form 2 of EFdA were observed at -114.73, -116.74 and -118.78 ppm. This NMR measurement can be used alone or in combination with any other characteristics of Form 2 described herein to identify Form 2 of EFdA and distinguish it from other crystalline forms of EFdA.

[0119] Thus, in another aspect, the anhydrous crystalline Form 2 of EFdA is characterized by a solid phase having peaks at -114.73, -116.74 and -118.78 ppm. 19 F NMR spectrum.

[0120] In another aspect, the anhydrous crystalline Form 2 of EFdA is characterized by a solid state comprising any two of the following peaks 19 FNMR spectrum: -114.73, -116.74 and -118.78 ppm. In another aspect, the anhydrous crystalline Form 2 of EFdA is characterized by a solid phase comprising the following peaks 19 F NMR spectrum: -116.74 and -118.78 ppm.

[0121] In another aspect, Form 2 of EFdA is characterized by Figure 5 The solid state shown in 19 F NMR spectrum.

[0122] In another aspect, the anhydrous crystalline Form 2 of EFdA is characterized by a solid phase having peaks at -114.73, -116.74 and -118.78 ppm in combination with any one or more of the PXRD peaks in Table 2 or aspects thereof as described above. 19 FNMR spectrum.

[0123] In another aspect, the anhydrous crystalline Form 2 of EFdA is characterized by any one or more PXRD peaks in Table 2 or combinations of aspects thereof as described above. Figure 5 The solid state shown in 19 F NMR spectrum.

[0124] Anhydrous crystalline form 3 of EFdA

[0125] Anhydrous crystalline form 3 of EFdA: PXRD pattern

[0126] The PXRD pattern of the anhydrous crystalline form 3 of EFdA is depicted in Figure 6 Therefore, in one aspect of the present disclosure, there is provided a method of Figure 6 The powder x-ray diffraction pattern shown in is characteristic of the anhydrous crystalline Form 3 of EFdA. The peak positions (on the 2θ x-axis) that coincide with these contours are shown in Table 3 (+ / - 0.2° 2θ). The positions of these PXRD peaks are characteristic of the anhydrous crystalline Form 3 of EFdA.

[0127] Thus, in another aspect, the anhydrous crystalline Form 3 of EFdA is characterized by a powder x-ray diffraction pattern having the peak positions listed in Table 3 (+ / - 0.2° 2θ).

[0128] Table 3

[0129]

[0130] In another aspect, the anhydrous crystalline Form 3 of EFdA is characterized by a powder x-ray diffraction pattern comprising 2 or more 2θ values ​​listed in Table 3 (+ / - 0.2° 2θ).

[0131] In another aspect, the anhydrous crystalline Form 3 of EFdA is characterized by a powder x-ray diffraction pattern comprising 3 or more 2θ values ​​listed in Table 3 (+ / - 0.2° 2θ).

[0132] In another aspect, the anhydrous crystalline Form 3 of EFdA is characterized by a powder x-ray diffraction pattern comprising 4 or more 2θ values ​​listed in Table 3 (+ / - 0.2° 2θ).

[0133] In another aspect, the anhydrous crystalline Form 3 of EFdA is characterized by a powder x-ray diffraction pattern comprising 6 or more 2θ values ​​listed in Table 3 (+ / - 0.2° 2θ).

[0134] Anhydrous crystalline form 4 of EFdA

[0135] Anhydrous crystalline form 4 of EFdA: Powder X-ray diffraction (PXRD)

[0136] The PXRD pattern of the anhydrous crystalline form 4 of EFdA is shown in Figure 7 Therefore, in one aspect of the present disclosure, there is provided a method of Figure 7 The powder x-ray diffraction pattern shown in FIG4 is characteristic of the anhydrous crystalline form of EFdA.

[0137] In another aspect, the anhydrous crystalline Form 4 of EFdA is characterized as follows Figure 7 PXRD spectrum shown in .

[0138] In yet another aspect, the anhydrous crystalline Form 4 of EFdA is characterized by Figure 7 The data shown in , alone or in combination with any other features of Form 4 of EFdA described herein.

[0139] Anhydrous crystalline form 4 of EFdA: 19 F(Fluorine-19) Solid State NMR :

[0140] Use the above 19 F (fluorine-19) solid-state NMR equipment and procedures to obtain the solid-state form 4 of EFdA 19 F NMR spectrum. The spectrum shows Figure 8 middle.

[0141] In one aspect, Form 4 of EFdA is characterized as follows Figure 8 The solid state shown in 19 F NMR spectrum.

[0142] In another aspect, Form 4 of EFdA is characterized by a solid state comprising the following peaks: 19 F NMR spectrum: -116.96 and -118.36 ppm.

[0143] Thus, in yet another aspect, Form 4 of EFdA is characterized by the above 19 F NMR characteristic peaks and Figure 8 The data shown in , alone or in combination with any other features of Form 4 of EFdA described herein.

[0144] Anhydrous crystalline form 4 of EFdA: Thermogravimetric analysis (TGA) :

[0145] Form 4 of EFdA was subjected to TGA analysis using the thermogravimetric analysis equipment and procedure described above. Fig. 9 A typical TGA analysis curve of Form 4 of EFdA is depicted. The data shows a 0.02 wt.% loss until 148°C, followed by thermal decomposition above 250°C. This TGA analysis can be used alone or in combination with any other characteristics of Form 4 described herein to identify Form 4 of EFdA and distinguish it from other crystalline forms of EFdA. Thus, in another aspect, Form 4 of EFdA is characterized by being substantially as Fig. 9 In yet another aspect, Form 4 of EFdA is characterized by any of these TGA measurements and / or substantially as Fig. 9 The TGA curve shown in FIG. 1 , alone or in combination with any one or more of the other characteristics described herein, including various aspects of the PXRD characterization described above and / or the above description of Form 419 Various aspects of F ssNMR.

[0146] PXRD pattern of monohydrate crystalline form MH:

[0147] The PXRD pattern of form MH is shown in Fig.10 Therefore, in one aspect of the present disclosure, there is provided a method of Fig.10 The powder x-ray diffraction pattern shown in Table 5 is characteristic of the monohydrate crystalline form MH of EFdA. The peak positions (on the 2θ x-axis) consistent with these contour lines are shown in Table 5 (+ / - 0.2° 2θ). The positions of these PXRD peaks are characteristic of form MH of EFdA. Therefore, in another aspect, the anhydrous crystalline form MH of EFdA is characterized by a powder x-ray diffraction pattern having the respective peak positions (+ / - 0.2° 2θ) listed in Table 5.

[0148] Table 5

[0149]

[0150]

[0151] Thus, in one aspect, the monohydrate crystalline Form MH of EFdA is characterized by a powder x-ray diffraction pattern having the peak positions listed in Table 5 (+ / - 0.2° 2θ).

[0152] In another aspect, the monohydrate crystalline Form MH of EFdA is characterized by a powder x-ray diffraction pattern comprising 2 or more 2θ values ​​listed in Table 5 (+ / - 0.2° 2θ).

[0153] In another aspect, the monohydrate crystalline Form MH of EFdA is characterized by a powder x-ray diffraction pattern comprising 3 or more 2θ values ​​listed in Table 5 (+ / - 0.2° 2θ).

[0154] In another aspect, the monohydrate crystalline Form MH of EFdA is characterized by a powder x-ray diffraction pattern comprising 4 or more 2θ values ​​listed in Table 5 (+ / - 0.2° 2θ).

[0155] In another aspect, the monohydrate crystalline Form MH of EFdA is characterized by a powder x-ray diffraction pattern comprising 6 or more 2θ values ​​listed in Table 5 (+ / - 0.2° 2θ).

[0156] In another aspect, the monohydrate crystalline Form MH of EFdA is characterized by a powder x-ray diffraction pattern comprising 9 or more 2θ values ​​listed in Table 5 (+ / - 0.2° 2θ).

[0157] In another aspect, the monohydrate crystalline Form MH of EFdA is characterized by a powder x-ray diffraction pattern comprising 12 or more 2θ values ​​listed in Table 5 (+ / - 0.2° 2θ).

[0158] In yet another aspect, the crystalline monohydrate form MH of EFdA that best represents Table 5 and / or Fig.10 The PXRD peak positions shown in and classified as "identification peak set" to easily distinguish this crystalline form from other crystalline forms. A selection of such characteristic peaks is given in Table 5 in the column labeled "identification peak set".

[0159] Thus, in another aspect, there is provided a monohydrate crystalline Form MH of EFdA characterized by a powder x-ray diffraction pattern comprising each of the 2θ values ​​listed in Identification Peak Set 1 in Table 5 (+ / - 0.2° 2θ).

[0160] In another aspect, there is provided a monohydrate crystalline Form MH of EFdA characterized by a powder x-ray diffraction pattern comprising each of the 2θ values ​​listed in Identification Peak Set 2 in Table 5 (+ / - 0.2° 2θ).

[0161] In another aspect, there is provided a monohydrate crystalline Form MH of EFdA characterized by a powder x-ray diffraction pattern comprising each of the 2θ values ​​listed in Identification Peak Set 3 in Table 5 (+ / - 0.2° 2θ).

[0162] In another aspect, there is provided a monohydrate crystalline Form MH of EFdA characterized by a powder x-ray diffraction pattern comprising each of the 2θ values ​​listed in Identification Peak Set 1 and Identification Peak Set 2 in Table 5 (+ / - 0.2° 2θ).

[0163] In another aspect, there is provided a monohydrate crystalline Form MH of EFdA characterized by a powder x-ray diffraction pattern comprising each of the 2θ values ​​listed in Identification Peak Set 1 and Identification Peak Set 3 in Table 5 (+ / - 0.2° 2θ).

[0164] In another aspect, there is provided a monohydrate crystalline Form MH of EFdA characterized by a powder x-ray diffraction pattern comprising each of the 2θ (+ / - 0.2° 2θ) values ​​listed in Identified Peak Set 1 and Identified Peak Set 2 and Identified Peak Set 3 in Table 5.

[0165] Form MH 19 F(Fluorine-19) Solid State NMR :

[0166] Use the above 19 F (fluorine-19) solid-state NMR equipment and procedures to obtain the solid-state MH of the monohydrate form of EFdA 19 FNMR spectrum of monohydrate form of MH 19 F NMR spectrum shows Fig.11 Characteristic peaks of Form MH were observed at -116.52 and -121.13 ppm. This NMR measurement can be used alone or in combination with the PXRD characterization data described in Table 5 to identify Form MH of EFdA and distinguish it from other crystalline forms of EFdA.

[0167] In another aspect, Form MH of EFdA is characterized by a solid phase having peaks at -116.52 and / or -121.13 ppm. 19 F NMR spectrum.

[0168] Thus, in another aspect, the monohydrate form MH of EFdA is characterized by Fig.11 The solid state shown in 19 FNMR spectrum.

[0169] In another aspect, Form MH of EFdA is characterized by the above-described characteristic NMR peaks in combination with any one or more of the PXRD peaks in Table 5, or aspects thereof as described above.

[0170] When the monohydrate Form MH is used, in-situ recrystallization occurs during the formulation process, which results in transformation into a mixture of multiple thermodynamically unfavorable anhydrous phases in the drug product. In contrast, preparation of the drug product using the thermodynamically stable Form 4 results in the same thermodynamically stable "Form 4" remaining unchanged throughout the process and in the drug product. Thus, unlike Form MH, Form 4 can be used to prevent EFdA from phase transformation into a thermodynamically unfavorable API phase during the HME formulation process for implantation into the drug product.

[0171] Similarly, unlike the monohydrate Form MH, "Form 1" is also physically stable in the implant HME preparation process, remaining unchanged throughout processing and in the drug product.

[0172] Figure 12-14 is a PXRD pattern generated for a LAP implant drug product composition composed of EFdA and EVA polymers ("EFdA / EVA Product") prepared by HME. Fig.12 The monohydrate form MH proved to be unsuitable for implant HME preparation as it changes the crystalline form to the undesirable and thermodynamically unfavorable Form 2 during the process. Fig.13 It was demonstrated that Form 4 was generated by an unusual and unexpected mechanism involving the use of high shear forces in the HME process. Fig.14 The thermal stability of Form 1 in the HME process was demonstrated. Fig.15 is a PXD pattern generated for a LAP implant drug product composition composed of EFdA, EVA polymer, and BaSO4 ("EFdA / EVA / BaSO4 Product") prepared by HME. Fig.15The thermal stability of Form 4 in the HME process was demonstrated, thereby achieving a long-acting implantable EFdA product composed of the thermodynamically stable phase EFdA.

[0173] Example 1

[0174] The Gibbs free energy (ΔG) of a phase is a fundamental thermodynamic parameter that determines the relative stability of a polymorph at a given temperature. The solubility of a particular polymorph is related to ΔG by the equation ΔG=-RTln(solubility). The polymorph with the lowest solubility is considered to be the thermodynamically stable phase at a given temperature. Table 6 lists the measured solubilities of Forms 1, 2, and 4 of EFdA in acetonitrile between 25.0 and 65.0°C. The data clearly demonstrate that the most thermodynamically stable phase of EFdA in this measured temperature range is Form 4, with Form 2 being the most unstable phase. The relatively low ΔΔG values ​​for converting Form 1 to Form 4 (i.e., the energy gap between Form 1 and Form 4 is small; 0.21-0.35 kJ / mol), combined with the rapid crystallization kinetics of Form 1 compared to Form 4, result in the difficulty of isolating Form 4 using conventional polymorph screening methods.

[0175] Table 6:

[0176] Temperature dependent solubility of EFdA forms 1, 2, and 4 in acetonitrile and calculation between forms 1 and 4 ΔΔ Gibbs free energy (G)

[0177]

[0178] The thermodynamic parameter for the stability of the monohydrate form MH and the anhydrous form 4 of EFdA is the critical water activity. Below this water activity, the anhydrous form 4 is thermodynamically preferred. Above this water activity, the monohydrate form MH is thermodynamically preferred. The solubility of EFdA monohydrate form MH and anhydrous forms 1 and 4 in acetonitrile was determined as a function of water content at 25.0°C. The data is plotted in Fig.16 The critical water activity of the EFdA monohydrate / Form 4 system at 25°C was determined to be 0.3.

[0179] Example 2: EFdA monohydrate crystalline form MH

[0180] A suitable starting amount of Form MH of EFdA can be obtained by the synthetic method described in US Pat. No. 7,339,053.

[0181] As will be appreciated by those skilled in the art, the use of seeds in the preparation of Form 1 and Form 4 as described in Examples 4 and 6 is not initially required, but is used for optimal preparation after initial amounts of crystalline Form 1 and Form 4, respectively, are prepared.

[0182] Example 3: Anhydrous Crystalline Form 1 of EFdA

[0183] EFdA Form 1 was prepared by adding 1.92 g of EFdA solid and 16.0 g of methanol (MeOH) to a clean reactor and heated to 65° C. while stirring. The mixture was cooled to 40° C. over 30 minutes, to 20° C. over 1 hour, and then stirred at 20° C. for ~14 hours. The slurry was filtered and the filter cake was dried by passing N2 through the filter cake at ambient temperature for 2 hours. 1.59 g of EFdA Form 1 was collected in ~88% isolated yield.

[0184] Example 4: Anhydrous Crystalline Form 1 of EFdA

[0185] EFdA Form 1 was prepared by dissolving 16.12 g of EFdA solid in dimethylformamide (DMF) with stirring at ambient temperature for 30 minutes. The resulting solution was filtered to remove undissolved material, the filter was washed twice with ~1 ml of DMF, and the filtrate and washings were combined. 294.4 g of isopropyl alcohol (IPA) was added to a clean reactor and heated to 50°C. While stirring the IPA, 15.0 g of the EFdA / DMF solution was added to the IPA. The solution was seeded with 152 mg of EFdA Form 1 and stirred for 20 minutes. 10.56 g of the EFdA / DMF solution was added to the reactor and stirred for 20 minutes. 10.79 g of the EFdA / DMF solution was added to the reactor and stirred for 15 minutes. The slurry was stirred at 50°C for 1 hour, cooled to 10°C over 12 hours, and stirred at 10°C for 2.5 hours. The slurry was filtered and the wet cake was washed twice with ~13 ml of IPA. The wet solid was dried by passing N2 through the filter cake at ambient temperature for 3 hours. 13.54 g of EFdA Form 1 was collected in -84% isolated yield.

[0186] Example 5: Anhydrous Crystalline Form 4 of EFdA

[0187] EFdA Form 4 was prepared by premixing 0.396 grams of water (HO) with acetonitrile (MeCN) to a total solvent weight of 31.66 grams. EFdA Form MH (monohydrate) (2.83 grams) and 31.02 grams of MeCN / HO solvent mixture were added to a clean reactor. The resulting slurry was stirred at 25°C for 5 minutes, then heated to 35°C over 30 minutes, and then heated to 40°C over 30 minutes. After stirring at 40°C for 45 minutes, the slurry was heated to 50°C over 2 hours, and then stirred at 50°C for 1 hour. After aging at 50°C for 1 hour, the slurry was cooled to 25°C over 8 hours. The resulting slurry was filtered and dried by passing nitrogen (N2) through the filter cake at ambient temperature for 24 hours. 2.47 grams of EFdA Form 4 were collected with a 93% isolated yield.

[0188] Example 6: Anhydrous Crystalline Form 4 of EFdA

[0189] EFdA Form 4 was prepared using critical water activity data by developing control over supersaturation by slowly heating a slurry of the monohydrate in a system where the amount of water was slightly below the critical water activity. Form 4 was prepared by premixing 0.9134 grams of water and 73.07 grams of acetonitrile in a bottle. 60.03 grams of acetonitrile / water mixture and 7.82 grams of EFDA monohydrate were added to a clean container. The suspension was stirred at 25°C for 30 minutes. After a 30-minute aging period, 0.80 grams of EFDA Form 4 seeds were added and the suspension was stirred at 25.0°C for 30 minutes. The suspension was heated linearly to 55°C over 10 hours. 42.5 milliliters of acetonitrile was linearly added to the slurry while stirring at 55°C. At the end of the acetonitrile addition, the slurry was stirred at 55°C for 1 hour. The slurry was then cooled linearly to 25.0°C over 4 hours and stirred at 25°C for another 2 hours. The slurry was filtered and washed with 20 mL of acetonitrile and the filter cake was dried by pulling nitrogen through it at ambient temperature for 24 hours. EFDA Form 4 (7.82 g) was collected to give a 95% yield after correction for seeding.

[0190] Example 7

[0191] EFdA / EVA products were made from Form MH of EFdA and EVA using HME Low Shear Process A. Although made from Form MH raw material of EFdA, the HME processing phase-converted Form MH into anhydrous Form 1 and anhydrous Form 2 in the drug product. Fig.12 The XRD pattern of the resulting mixture of Form 1 and Form 2 of EFdA in this EFdA / EVA product is shown. This result confirms that EFdA Form MH lacks stability under the manufacturing conditions required to prepare EFdA / EVA implant products, and therefore Form MH of EFdA is not suitable for use in implant products.

[0192] Example 8

[0193] EFdA / EVA products were made from Form MH of EFdA and EVA by processing with limited process parameter space using HME high shear process B. Although made from Form MH raw material of EFdA, the HME processing phase-converted Form MH into anhydrous Form 4. Fig.13 The XRD pattern of the resulting EFdA, Form 4, in this EFdA / EVA product is shown. This result confirms the unusual way in which Form 4 was found. This also confirms that Form MH of EFdA is not suitable for use in hot melt extruded EFdA / EVA products due to its instability under processing conditions.

[0194] Example 9

[0195] The EFdA / EVA product was made from Form 1 of EFdA and EVA using HME low shear process A. Under the process conditions, Form 1 of EFdA was retained throughout the HME process into the EFdA / EVA product. Fig.14 The XRD pattern of the resulting EFdA Form 1 in this EFdA / EVA product is shown. This confirms that EFdA Form 1 is superior to EFdA Form MH due to the thermal stability of Form 1 in the HME process.

[0196] Example 10

[0197] An EFdA / EVA product containing BaSO4 was produced from Form 4 of EFdA and EVA using HME low shear process A. Under the process conditions, Form 4 of EFdA was retained in the EFdA / EVA product throughout the HME process. Fig.15 The XRD pattern of the resulting EFdA Form 4 in this EFdA / EVA product is shown. This confirms that EFdA Form 4 is superior to EFdA Form MH due to the thermal stability of Form 4 in the HME process. This result also confirms the thermodynamic stability of Form 4 and also confirms the data shown in Example 1.

[0198] nature Crystalline Forms 1 and 4 of EFdA described and characterized herein exhibit excellent physical properties while minimizing the difficulties associated with drug product preparation and processing. For example, crystalline Forms 1 and 4 of EFdA exhibit unexpectedly improved thermal stability in drug products (solid dosage form long-acting parenteral implants) compared to monohydrate Form MH of EFdA while remaining a BCS Class I material. Despite its desirable properties, crystalline Form 4 of EFdA did not appear in conventional polymorph screening; it was surprisingly and advantageously created after making many batches of other crystalline forms (such as monohydrate MH and crystalline anhydrous Form II) at many manufacturing sites under various conditions using many synthetic routes.

[0199] The thermodynamic stability of crystalline Form 4 of EFdA was evaluated using competitive slurry experiments in various solvent systems. Crystalline Forms 1 and 4 of EFdA obtained as described above were slurried in various solvents at controlled temperatures for extended periods of time. At the end of the experiment, the solvent was removed and the remaining crystalline material was evaluated using powder X-ray diffraction (PXRD) to confirm the resulting form. The more stable form was generally left, and the less stable form was converted to the more stable form. In all cases, Crystalline Form 4 of EFdA was the only form left and was therefore the more stable form.

[0200] The use of crystalline forms of EFdA according to the invention enables the use of this formulation strategy to prepare long-acting parenteral formulations while maintaining a thermodynamically stable phase. This is of great interest because crystalline Form 4 of EFdA exhibits reduced physical stability risks compared to higher energy forms. This ultimately allows for a less protective and potentially cheaper packaging configuration.

[0201] Pharmaceutical composition

[0202] As described above, another embodiment provides a pharmaceutical composition comprising crystalline Form 1 or Form #4 of EFdA (as characterized by any of the features described herein, alone or in combination). In such a composition, crystalline Form 1 or Form 4 of EFdA constitutes the sole active agent, or optionally is present in combination with one or more additional therapeutic agents. In either case, the pharmaceutical composition may further comprise one or more pharmaceutically acceptable carriers, excipients, and / or diluents. Non-limiting examples of additional therapeutic agents that can be used in combination with crystalline Form 1 or Form 4 of EFdA are described, for example, in PCT Publication WO 2017 / 196697 and include those selected from drugs for treating or preventing HIV infection and / or treating, preventing, or delaying the onset of ARC or AIDS.

[0203] When used in combination with an additional therapeutic agent, crystalline Form 1 or crystalline Form 4 of EFdA and the one or more additional therapeutic agents may be administered together or sequentially as described above. When used simultaneously with one or more other drugs, pharmaceutical compositions containing unit dosage forms of these other drugs and crystalline Form 1 or crystalline Form 4 of EFdA are contemplated. However, combination therapy may also include therapies in which crystalline Form 1 or crystalline Form 4 and one or more other drugs are administered at different overlapping schedules. It is also contemplated that when used in combination with one or more other active ingredients, the dosages of crystalline Form 1 or crystalline Form 4 and the other active ingredients may be lower than when each is used alone. In addition, these other drugs may be administered simultaneously or sequentially with crystalline Form 1 or crystalline Form 4 by their commonly used routes and amounts. When crystalline Form 1 or crystalline Form 4 is used simultaneously with one or more other drugs, pharmaceutical compositions containing these other drugs in addition to crystalline Form 1 or crystalline Form 4 may be prepared without undue experimentation according to methods described herein and / or known in the art.

[0204] The weight ratio of crystalline form 1 or crystalline form 4 to the second active ingredient can vary and depends on the effective dose of each ingredient. Typically, the respective effective doses are used. Thus, for example, when crystalline form 1 or crystalline form 4 of EFdA is combined with another agent, the weight ratio of crystalline form 1 or crystalline form 4 to the second agent is typically from about 1000:1 to about 1:1000, such as from about 200:1 to about 1:200, wherein in each case an effective dose for the intended use is used. Such a combination may be administered separately or simultaneously, and the administration of one agent may be before, at the same time, or after the administration of the other agent.

[0205] For the preparation of the pharmaceutical compositions described herein, the pharmaceutically acceptable carrier can be a solid or liquid, or any other known dosage form, such as an aerosol or lotion. Non-limiting examples of solid form preparations include powders, tablets, dispersible granules, capsules, cachets, and suppositories. Powders and tablets can be composed of any weight % value of the active ingredients described herein and in any desired dosage (e.g., the dosages described herein).

[0206] Crystalline Form 1 or Crystalline Form 4 of EFdA can be conveniently presented in unit dosage form, which can be prepared by any method well known in the art of pharmacy. All methods include the step of combining Crystalline Form 1 or Crystalline Form 4 with carriers that constitute auxiliary ingredients. In general, pharmaceutical compositions are prepared by uniformly and intimately combining the active ingredient with liquid carriers or finely divided solid carriers or both, and then, if necessary, shaping the product into the desired formulation.

[0207] In the pharmaceutical composition, the active ingredient is included in an effective amount. The term "effective amount" as used herein refers to the amount of a compound that is sufficient to inhibit HIV reverse transcriptase, inhibit HIV replication, exert a preventive effect and / or exert a therapeutic effect after administration. One embodiment of an "effective amount" is a "therapeutically effective amount", which is an amount of a compound that effectively inhibits HIV reverse transcriptase, inhibits HIV replication (any of the foregoing may also be referred to herein as an "inhibitory effective amount"), treats HIV infection, treats AIDS, delays the onset of AIDS and / or slows down the progression of ARC or AIDS in HIV-infected patients. Another embodiment of an "effective amount" is a "preventive effective amount", which is an amount of a compound that effectively prevents HIV infection in subjects who are not infected with HIV or prevents ARC or AIDS in HIV-infected patients. It is to be understood that an effective amount can be both a therapeutically effective amount, such as for treating HIV infection, and a preventive effective amount, such as for preventing AIDS or reducing the risk of AIDS in subjects infected with HIV. The term "prevention" used herein with respect to HIV virus infection or AIDS refers to reducing the possibility or severity of HIV infection or AIDS. When crystalline Form 1 or Form 4 of EFdA is administered as a salt, the amount of the compound mentioned in milligrams or grams is based on the free form (ie, non-salt form) of the compound.

[0208] In the combination therapies of the present disclosure, an effective amount may refer to an individual agent or to the entire combination, wherein the amounts of all agents administered in the combination are effective together, but wherein the component agents of the combination may or may not be individually present in an effective amount - this refers to an amount that is considered effective for the component agent when administered alone.

[0209] Pharmaceutical compositions intended for parenteral use are contemplated herein, particularly long-acting implantable drug delivery devices suitable for providing effective amounts of crystalline Form 1 or Form 4 of EFdA over extended periods of time, such as, but not limited to, over 1 month, 3 months, 6 months, or 1 year or longer. Parenteral compositions may be prepared according to techniques known in the art and typically use sterile water as a carrier and optional other ingredients, such as a solubility aid.

[0210] Pharmaceutical compositions such as tablets or capsules consisting of crystalline Form 1 or crystalline Form 4 of EFdA intended for oral use can be prepared according to the methods described herein and other methods well known in the art for preparing pharmaceutical compositions. If pharmaceutically refined and / or palatable preparations are desired, such compositions may further contain active agents selected from sweeteners, flavoring agents, coloring agents and preservatives. Tablets or capsules may contain active ingredients mixed with non-toxic pharmaceutically acceptable excipients suitable for preparing tablets. Additional examples of dosage forms, formulations and pharmaceutically acceptable carriers and methods for preparing various compositions can be found in Remington's Pharmaceutical Sciences, 18th edition, edited by A.R. Gennaro, Mack Publishing Co., 1990; and Remington-The Science and Practice of Pharmacy, 22nd edition, published by Pharmaceutical Press and Philadelphia College of Pharmacy at University of the Sciences, 2012, ISBN 978085711-062-6 and previous versions.

[0211] Another embodiment provides suitable dosages and dosage forms of crystalline Form 1 or Form 4 of EFdA and uses in various methods described herein. A person skilled in the art, such as an attending physician, pharmacist or other skilled worker can easily determine the appropriate dosage for administering crystalline Form 1 or Form 4 to a patient, and can vary according to the patient's health, age, weight, frequency of administration, route of administration and / or duration of administration, use with other active ingredients and / or indications for administration of crystalline Form 1 or Form 4. Therefore, the dosage of the active ingredient in the composition of the present invention can be varied, but the amount of the active ingredient should be such that a suitable dosage form is obtained. These dosages can provide the optimal dosage for administration to patients in need of such treatment. The daily dosage can be, for example, about 0.01-10 mg per day - by sustained release from an implantable device over a period of 1 month, 3 months, 6 months, 1 year or more, or from a tablet or capsule containing a daily dosage of about 0.01-10 mg of crystalline Form 1 or Form 4 per day.

Claims

1. A crystalline anhydrous form 1 of EFdA, characterized in that A powder x-ray diffraction pattern having at least peaks at diffraction angles °2θ (+ / - 0.2°) 4.48, 11.79, 14.70, 8.99, 12.39 and 16.88 in a powder x-ray diffraction obtained using Cu Ka radiation.

2. The crystalline anhydrous form 1 of EFdA according to claim 1, characterized in that A powder x-ray diffraction pattern having peaks at diffraction angles °2θ (+ / - 0.2°) at least 4.48, 11.79, 14.70, 8.99, 12.39, 16.88, 10.39, 15.51, 18.09 and 20.

16.

3. The crystalline anhydrous Form 1 of EFdA according to claim 1, having a powder x-ray diffraction pattern as shown in FIG1.

4. Crystalline anhydrous form 1 of EFdA, characterized in that A solid state exhibiting any two of the following peaks 19 F NMR spectrum: -114.75, -117.09 and -118.92 ppm.

5. The crystalline anhydrous form 1 of EFdA according to claim 4, characterized in that The solid state exhibiting the following peaks 19 F NMR spectrum: -114.75, -117.09 and -118.92 ppm.

6. The crystalline anhydrous form 1 of EFdA according to claim 4, characterized in that As shown in Figure 2, the solid 19 F NMR spectrum.

7. The crystalline anhydrous Form 1 of EFdA according to any one of claims 1 to 3, further characterized by a solid phase exhibiting any two of the following peaks: 19 F NMR spectrum: -114.75, -117.09 and -118.92 ppm.

8. The crystalline anhydrous Form 1 of EFdA according to claim 7, further characterized by: 1) TGA data shows 0.1 wt.% loss up to 133°C, followed by thermal decomposition above 240°C, or 2) TGA curve as shown in Figure 3.

9. The crystalline anhydrous Form 1 of EFdA according to any one of claims 1 to 3, further characterized in that The solid state exhibiting the following peaks 19 F NMR spectrum: -114.75, -117.09 and -118.92 ppm.

10. The crystalline anhydrous Form 1 of EFdA according to claim 9, further characterized by: 1) TGA data shows 0.1 wt.% loss up to 133°C, followed by thermal decomposition above 240°C, or 2) TGA curve as shown in Figure 3.

11. The crystalline anhydrous Form 1 of EFdA according to any one of claims 1 to 3, further characterized in that As shown in Figure 2, the solid 19 F NMR spectrum.

12. The crystalline anhydrous Form 1 of EFdA according to claim 11, further characterized by: 1) TGA data shows 0.1 wt.% loss up to 133°C, followed by thermal decomposition above 240°C, or 2) TGA curve as shown in Figure 3.

13. The crystalline anhydrous form 1 of EFdA according to claim 12, characterized in that TGA data showed 0.1 wt. % loss until 133 °C, followed by thermal decomposition above 240 °C.

14. The crystalline anhydrous form 1 of EFdA according to claim 12, characterized in that The TGA curve is shown in Figure 3.

15. The crystalline anhydrous form 1 of EFdA according to claim 1, characterized in that A powder x-ray diffraction pattern comprising 6 or more 2-theta values ​​listed in Table 1 (+ / - 0.2° 2-theta).

16. Crystalline anhydrous form 4 of EFdA, characterized in that A solid state exhibiting at least the following peaks 19 F NMR spectrum: -116.96 and -118.36 ppm.

17. Crystalline anhydrous Form 4 of EFdA having a powder x-ray diffraction pattern as shown in FIG7.

18. Crystalline anhydrous form 4 of EFdA, characterized in that As shown in Figure 8, the solid 19 F NMR spectrum.

19. The crystalline anhydrous form 4 of EFdA according to any one of claims 16 to 18, characterized in that: 1) TGA data shows 0.02 wt.% loss up to 148°C, followed by thermal decomposition above 250°C, or 2) TGA curve as shown in FIG9 .

20. The crystalline anhydrous Form 4 of EFdA according to claim 16, further characterized by The powder x-ray diffraction pattern is shown in FIG7 .

21. The crystalline anhydrous Form 4 of EFdA of claim 20, further characterized by As shown in Figure 8, the solid 19 FNMR spectrum.

22. The crystalline anhydrous Form 4 of EFdA of claim 21, further characterized by: 1) TGA data shows 0.02 wt.% loss up to 148°C, followed by thermal decomposition above 250°C, or 2) TGA curve as shown in FIG9 .

23. Crystalline anhydrous form 4 of EFdA, characterized in that A powder x-ray diffraction pattern having peaks at diffraction angles °2θ (+ / - 0.2°) at 4.48, 11.79, 12.39, 14.70, 15.51, 18.09, 25.81 and 27.

42.

24. The crystalline anhydrous form 4 of EFdA according to claim 23, characterized in that A powder x-ray diffraction pattern comprising 6 or more 2-theta values ​​(+ / - 0.2° 2-theta) listed in Table 4.

25. A pharmaceutical composition comprising a crystalline anhydrous form of EFdA according to any one of claims 1 to 24 and a pharmaceutically acceptable carrier.

26. The pharmaceutical composition of claim 25, which is a parenteral implant composition.

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