Pharmaceutical composition for herpes virus

Abstract

The present invention relates to methods and compositions for treating and / or inhibiting the development or progression of a disease or disorder caused by or associated with herpes virus infection. In particular, the present invention provides a long acting injectable depot pharmaceutical composition comprising a helicase-eliciting enzyme inhibitor; a method for manufacturing the same; and the use of said pharmaceutical composition as a medicament and for the treatment of diseases or disorders caused by or associated with herpes virus.

Description

Technical Field

[0001] Provided herein are compositions and methods for treating and / or inhibiting the development or progression of a disease or disorder caused or associated with herpes simplex virus infection. Specifically, provided herein are long-acting injectable reservoir-type pharmaceutical compositions comprising helicase-primase inhibitors; methods for making the same; and uses of the pharmaceutical compositions as medicines and for treating a disease or disorder caused or associated with herpes simplex virus infection. Background Art

[0002] Human herpesviruses are large enveloped double-stranded DNA viruses that have the common characteristic of infecting humans for life. This is due to their ability to persist in the host as an asymptomatic latent infection, where the virus lies dormant, or, after activation, as a lytic infection with associated symptoms. These viral infections are widespread worldwide, and it is noteworthy that more than 90% of humans are chronically infected with more than one human herpesvirus.

[0003] Human herpesviruses are divided into three subfamilies (α, β and γ) based on their biological characteristics. The family consists of eight members, namely herpes simplex virus subtypes 1 and 2 (HSV1, HSV2), varicella-zoster virus (VZV), Epstein-Barr virus (EBV), cytomegalovirus (CMV) and human herpesvirus 6-8 (HHV 6-8).

[0004] Infection with HSV1 and 2 can cause disease in immunocompetent individuals. Both subtypes cause skin genital / anal and oral / nasal (cold sores) lesions, although HSV2 is usually associated with the former and HSV1 with the latter. >80% of genital infections are believed to be caused by HSV2. Globally, more than 500 million people are infected with genital herpes, and approximately 50% to 80% of the world's population is infected with oral HSV, which is the main cause of cold sores. HSV, especially HSV1, can also cause lesions on the fingers (Whitlows) and other skin sites.

[0005] The vast majority of people infected with HSV do not experience any noticeable symptoms. However, some people experience repeated (and often severe) outbreaks of the infection. In the United States, 20% to 40% of the population will have recurring HSV lesions on the lips. Of note, oral herpes and Whitlow's provide a very easy way for the virus to spread to other people, which can lead to rare but more serious HSV-related conditions. For example, HSV-related keratitis is a leading cause of blindness, and HSV can also cause neonatal encephalitis, a life-threatening condition. Other diseases believed to be caused by HSV include herpes gladiatorum, Molaret's meningitis, and possibly Bell's palsy.

[0006] Primary infection or reactivation of an existing herpesvirus infection may be the main cause of illness in immunocompromised individuals. The main high-risk groups include patients undergoing solid organ or stem cell transplantation, patients undergoing cancer treatment, HIV / AIDS patients, and ICU patients.

[0007] Currently, there is no cure for HSV. Drugs have been developed that can reduce the incidence and / or shorten the duration of symptoms to some extent, but there is still a need for improved treatments.

[0008] Currently, nucleoside analogs (such as acyclovir and its prodrugs, such as valacyclovir and famciclovir) are used as drugs against herpes viruses (such as HSV). In order to exert their effects, these nucleoside analogs must be phosphorylated by viral thymidine kinase (TK) and subsequently converted to nucleoside triphosphates by cellular kinases, thereby inhibiting the activity of viral DNA polymerase. If the virus does not have a functionally active TK, for example, in the case of resistant HHV1 mutants or TK-negative viruses, nucleoside analogs cannot exert their effects.

[0009] Nucleoside analogs are clinically administered at very high doses, for example, doses of up to several hundred milligrams to several grams are often administered daily. Even at these high doses, which are often administered during long-term treatment, these drugs cannot completely prevent the recurrence of symptoms of HSV infection. Nucleoside analogs are also powerless to address the problem of viral shedding, which can promote the spread of HSV to more people asymptomatically. Certain nucleoside analogs, especially when used in high doses, also raise safety concerns. For example, because these drugs can be integrated into the host's genomic DNA by host DNA polymerases, their mutagenicity is a concern, as documented for the nucleoside analog ganciclovir (Aoki, Chapter 45 in Mandell, Douglas and Bennett's Principles and Practice of Infectious Diseases (Eighth Edition) 2015).

[0010] Given the inadequacy of existing treatments, there is an urgent need to develop improved, well-tolerated anti-herpes therapies.

[0011] One class of compounds currently under investigation is the helicase-primase inhibitors. Helicase-primase inhibitors are antiviral drugs with a novel mechanism of action. They inhibit the viral heterotrimeric complex composed of helicase, primase, and cofactor subunits, which has functions that are essential for viral DNA replication. These drugs are not nucleoside analogs and do not require TK phosphorylation to inhibit HSV replication. Therefore, they have potential activity against TK-deficient HSV, which, as mentioned above, is the main mechanism of resistance to nucleoside analogs.

[0012] Two examples of helicase-primase inhibitors are BILS-179BS (Crute et al., (2002) Nature Medicine 8, pp. 386-391) and aminavir (Katsumata et al., (2018) Biochem Pharm 158, pp. 201-206).

[0013] BILS-179BS has been used for oral administration in humans, but was suspended in early clinical trials due to adverse events (Ruebsamen et al., (2019) Med. Chem. Commun., DOI: 10.1039 / C9MD00233B). Similarly, aminavir has also been used for oral administration in humans, but was suspended in early HSV clinical trials due to adverse events. In a dose-exploring placebo-controlled study, 437 patients with recurrent genital herpes took aminavir orally at doses of 100 mg, 200 mg, 400 mg, and 1200 mg. The study found that the time to lesion healing (the primary endpoint of the study) was significantly different only between the highest dose 1200 mg single-dose group and the placebo group (Aoki, Mandell, Douglas and Bennett's Principles and Practice of Infectious Diseases (Eighth Edition) Chapter 45, 2015).

[0014] Another example of a helicase-primase inhibitor is pritelivir, a thiazole amide derivative with the chemical name N-methyl-N-(4-methyl-5-sulfamoyl-1,3-thiazol-2-yl)-2-[4-(pyridin-2-yl)phenyl]acetamide. This compound has been disclosed in WO 2001 / 47904.

[0015] In a human clinical study, the inhibitory effect of pralivir on genital herpes was studied in 156 subjects (Wald et al., (2014) New England Journal of Medicine 370, pp. 201-210). Subjects received one of three oral daily doses or one weekly oral pralivir or placebo for 28 days. The four pralivir dosing regimens were: a loading dose of 20 mg, followed by a daily dose of 5 mg; a loading dose of 100 mg, followed by a daily dose of 25 mg; a loading dose of 300 mg, followed by a daily dose of 75 mg; and a weekly dose of 400 mg. At high doses, pralivir was found to reduce the genital HSV shedding rate, which was the primary endpoint of the study. Pralivir also reduced the number of lesion days in healthy men and women with genital herpes. An oral daily dose of 75 mg had the best antiviral effect and was found to be superior to a 400 mg dose with a lower weekly dosing frequency.

[0016] As highlighted in the study report, although HSV shedding was reduced at the highest daily dose of 75 mg compared with placebo, breakthrough shedding still occurred. In discussing this finding, Wald et al. ((2014) New England Journal of Medicine 370, pp. 201-210) explained that persistent low-level shedding has also been observed with nucleoside therapy, and that the pathogenesis of breakthrough viral shedding during adequate antiviral treatment with nucleoside analogs is unclear; it is not related to lack of adherence to the treatment regimen or viral resistance. The authors also raised the question of whether further increasing the daily dose of pralivir would completely eliminate viral shedding, which should be addressed in subsequent clinical studies.

[0017] Following this recommendation, a subsequent clinical study using a higher daily oral dose was indeed conducted. In a 28-day phase II clinical study in 91 patients with recurrent genital HSV-2, daily oral administration of 100 mg of pralivir (after a loading dose of 400 mg) resulted in HSV shedding in 2.4% of genital swabs analyzed, while daily administration of 500 mg of valacyclovir resulted in HSV shedding in 5.3% of swabs. The results of this study were reported by Wald et al. (2016) (J. Am. Med. Assoc. 316(23), p. 2495-2503), which states that the selection of a 100 mg daily pralivir dose (after a loading dose of 400 mg) was based on previous trial results showing that a 75 mg daily dose was highly effective in suppressing viral shedding.

[0018] In the current Phase III clinical trial, a 400 mg oral loading dose followed by 100 mg oral pritelivir daily for up to 28 days or until all mucocutaneous HSV lesions heal, whichever comes first, is being studied (https: / / www.clinicaltrials.gov / ct2 / show / NCT03073967?term=pritelivir+f&draw=2&rank=2).

[0019] As can be seen above, for all of the above small molecule-based antiviral therapies, the main route of administration studied is the oral route. Efforts to improve treatment, such as attempts to further reduce or prevent HSV shedding and viral reactivation, have mainly focused on using higher oral drug doses (including high loading doses) and more frequent dosing (e.g., multiple daily dosing).

[0020] Efforts to increase systemic exposure via the oral route are also evident from various attempts to improve the oral bioavailability of nucleoside analogs and helicase-primase inhibitors. For example, valacyclovir was developed as a prodrug of acyclovir to provide higher oral bioavailability; the bioavailability of valacyclovir in humans after oral administration is about 54%, while the bioavailability of oral acyclovir is only 10-20% (Murray (1995) Antiviral Chem. Chemotherapy 6 (1), p. 34-38). For pralivir, efforts have been made to improve the solubility of salt forms for oral administration. For example, WO 2013 / 045491 discloses that an increase in oral bioavailability can be achieved by using a crystalline monomethanesulfonate monohydrate salt of pralivir with specific particle characteristics. The disclosure describes pralivir free base as having unfavorable drug release and absorption characteristics compared to the monomethanesulfonate monohydrate salt due to poor solubility. The free base is also described as being unsuitable for long-term stable formulations. WO 2013 / 045479 discloses a specific crystalline monomesylate monohydrate salt of priligyvir and reports that the salt form has superior long-term stability and improved systemic exposure after oral administration compared to priligyvir free base.

[0021] There is a continuing need for new and improved methods of treating HSV infection and the present invention has been designed with the foregoing in mind. Summary of the invention

[0022] In a first aspect, the present invention provides a long-acting injectable depot pharmaceutical composition comprising a helicase-initiation enzyme inhibitor or a pharmaceutically acceptable salt thereof; and one or more pharmaceutically acceptable excipients; wherein the helicase inhibitor has a terminal biological half-life in humans of 10 hours or more.

[0023] Surprisingly, when a helicase-initiation enzyme inhibitor or a pharmaceutically acceptable salt thereof having a human biological terminal half-life of 10 hours or more is formulated into a long-acting injectable depot pharmaceutical composition according to the present invention, such a composition provides beneficial properties, including certain release characteristics and properties that allow plasma levels to be achieved for a long time.

[0024] It is noteworthy that, in some embodiments, the long-acting injectable reservoir composition of the present invention is capable of stably and continuously releasing the helicase-priming enzyme inhibitor for at least 14 days after administration. This provides the possibility of less frequent administration and / or using lower doses compared to other routes of administration. This is an unexpected discovery. As described above, the main route of administration for small molecule antiviral therapy research is the oral route, and efforts to improve treatment are mainly focused on increasing oral drug doses (including the use of high loading doses) and using more frequent drug dosing regimens (e.g., multiple daily doses).

[0025] In some embodiments, it has also been found that certain long-acting injectable depot compositions of the present invention exhibit high drug loading capacity. This provides a number of advantages, including the possibility of administering the composition using a low injection volume.

[0026] The present invention addresses the need for new treatments for HSV infection that may provide improvements in efficacy and / or safety and / or patient use.

[0027] In a second aspect, the invention provides a method of forming a pharmaceutical composition according to the first aspect. Various techniques can be used to form the injectable depot composition of the invention, however, certain methods have been found to have specific advantages. In addition, it has also been found that certain chemical and / or physical forms of the helicase-initiase inhibitor (e.g., forms with low water solubility and low dissolution rate) are very suitable for being formulated into the injectable depot composition of the invention.

[0028] In a third aspect, the present invention provides a method for treating or preventing a herpes infection in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of a helicase-primase inhibitor or a pharmaceutically acceptable salt thereof, wherein the helicase-primase inhibitor or a pharmaceutically acceptable salt thereof is formulated as a pharmaceutical composition according to the first aspect. BRIEF DESCRIPTION OF THE DRAWINGS

[0030] The abstract and the following detailed description of the invention can be further understood by reading in conjunction with the accompanying drawings. In order to illustrate the disclosed compositions and methods, exemplary embodiments of the compositions and methods are shown in the drawings; however, the compositions and methods are not limited to the specific embodiments disclosed. In the drawings:

[0031] Figure 1 Shown is the XRPD diffraction pattern of Priligy free base dihydrate.

[0032] Figure 2 Shown are the mean plasma concentration-time profiles of priligy following subcutaneous administration of the aqueous suspension formulation disclosed in Example 1 at 33.3 (open triangles) and 166.7 (open squares) mg / kg in male Sprague Dawley rats (n=3 / group).

[0033] Figure 3 Shown are the mean plasma concentration-time profiles of priligy following subcutaneous administration of the aqueous suspension formulation disclosed in Example 2 at 10 mg / kg in male Sprague Dawley rats (n=3).

[0034] Figure 4 Shown are the mean plasma concentration-time profiles of priligy following subcutaneous administration of the aqueous suspension formulation disclosed in Example 3 at 10 mg / kg in male cynomolgus monkeys (n=3).

[0035] Figure 5 Shown are the mean plasma concentration-time profiles of priligy following subcutaneous administration of the formulation disclosed in Example 4 at 10 mg / kg in male non-naive beagle dogs (n=3).

[0036] Figure 6 Shown are the mean plasma concentration-time profiles of priligy following intravenous (IV) administration at 1 mg / kg and subcutaneous (SC) administration at 5 mg / kg of the solution formulation disclosed in Example 5 in male Sprague Dawley rats (n=3 / group).

[0037] Figure 7 Shown are mean plasma concentration-time profiles of Compound 1 of Example 7 following subcutaneous administration of an aqueous suspension formulation disclosed in Example 7 (triangles) at 10 mg / kg in male Sprague Dawley rats (n=3).

[0038] Figure 8 Shown are the mean plasma concentration-time profiles of Compound 1 of Example 7 following subcutaneous administration of an aqueous suspension formulation disclosed in Example 7 (triangles) at 10 mg / kg in male cynomolgus monkeys (n=3).

[0039] Fig. 9 Shown are mean plasma concentration-time profiles of Example 7 Compound 1 following subcutaneous administration of an aqueous suspension formulation disclosed in Example 7 (triangles) at 10 mg / kg in male non-naive beagle dogs (n=3).

[0040] Fig.10Shown are mean plasma concentration-time profiles of Compound 1 (Form A) following subcutaneous administration of 10 mg / kg of the aqueous suspension formulation disclosed in Example 10 (triangles) in male Sprague Dawley rats (n=3).

[0041] Fig.11 Shown are mean plasma concentration-time profiles of Compound 1 (Form A) following subcutaneous administration of an aqueous suspension formulation disclosed in Example 11 (triangles) at 10 mg / kg in male cynomolgus monkeys (n=3).

[0042] Fig.12 Shown are mean plasma concentration-time profiles of Compound 1 (Form A) following subcutaneous administration of an aqueous suspension formulation disclosed in Example 12 (diamonds) at 10 mg / kg in male non-naive beagle dogs (n=3).

[0043] Fig.13 Shown are the mean plasma concentration-time profiles of Compound 1 following intravenous administration of the solution formulation disclosed in Example 13 (open triangles) at 1 mg / kg in male Sprague Dawley rats (n=3).

[0044] Fig.14 Shown are the mean plasma concentration-time profiles of Compound 1 following intravenous administration of the solution formulation disclosed in Example 13 (open triangles) at 0.25 mg / kg in male cynomolgus monkeys (n=3).

[0045] Fig.15 Shown are mean plasma concentration-time profiles of Compound 1 following intravenous administration of the solution formulation disclosed in Example 13 (open triangles) at 0.15 mg / kg in male non-naive beagle dogs (n=3).

[0046] Fig.16 The XRPD diffraction pattern of priligyr free base sesquihydrate is shown.

[0047] Fig.17 The XRPD diffraction pattern of anhydrous Form A of Compound 1 free form is shown.

[0048] Fig.18 The XRPD diffraction pattern of anhydrous Form C of Compound 1 free form is shown.

[0049] Fig.19Shown are the mean plasma concentration-time curves of Compound 1 after intravenous injection of the solution formulation disclosed in Example 13A to the following animals: (A) male Sprague Dawley rats (n=3) at a dose of 0.2 mg / kg; (B) male non-naive beagle dogs (n=3) at a dose of 0.15 mg / kg; (C) male non-naive cynomolgus monkeys (n=3) at a dose of 0.2 mg / kg; and (D) naive Bama mini pigs (n=3) at a dose of 0.25 mg / kg.

[0050] Fig. 20 A SEM image of the microsuspension described in Example 14 is shown.

[0051] Fig.21 An SEM image of the nanosuspension described in Example 14 is shown.

[0052] Fig. 22 Shown are mean plasma concentration-time profiles of priligy following subcutaneous administration of solution (filled circles), microsuspension (filled triangles), and nanosuspension (filled squares) formulations disclosed in Example 14 at 10 mg / kg in male non-naive beagle dogs (n=3).

[0053] Fig.23 Shown are the mean plasma concentration-time profiles of Compound 1 following subcutaneous administration of the solution formulation disclosed in Example 15 (solid triangles) at 100 mg / kg in male Sprague Dawley rats (n=3).

[0054] Fig.24 Shown are mean plasma concentration-time profiles of Compound 1 following subcutaneous administration of the solution formulation disclosed in Example 16 at 10 mg / kg, 0.1 ml / kg (solid triangles) and 10 mg / kg, 0.2 ml / kg (solid diamonds) in male non-naive beagle dogs (n=3).

[0055] Fig.25 A SEM image of the microsuspension described in Example 17 is shown.

[0056] Fig.26 SEM images of the nanosuspension described in Example 17 are shown.

[0057] Fig. 27 Shown are the mean plasma concentration-time profiles of Compound 1 (Form C) following subcutaneous administration of the microsuspension (filled squares) and nanosuspension (filled triangles) formulations disclosed in Example 17 at a dose of 20 mg / rat (-80 mg / kg) in male Sprague Dawley rats (n=3).

[0058] Fig.28Shown are the mean plasma concentration-time profiles of Compound 1 (Form C) following intramuscular administration of the microsuspension (filled squares) and nanosuspension (filled triangles) disclosed in Example 17 at a dose of 20 mg / rat (-80 mg / kg) in male Sprague Dawley rats (n=3). DETAILED DESCRIPTION OF THE INVENTION

[0060] The disclosed compositions, uses thereof and methods may be more readily understood by reference to the following detailed description in conjunction with the accompanying drawings, which form a part of the present disclosure. It should be understood that the disclosed compositions and methods are not limited to the specific compositions and methods described and / or shown herein, and that the terminology used herein is only used to describe specific embodiments by way of example and is not intended to limit the claimed compositions and methods.

[0061] definition

[0062] Unless otherwise stated, the following terms used in the specification and claims have the meanings listed below.

[0063] Unless the context clearly dictates otherwise, references to a particular numerical value include at least that particular value. When expressing a range of values, another embodiment includes from one particular value and / or to another particular value. In addition, references to the values ​​described in a range include each and every value within that range. All ranges are inclusive and combinable.

[0064] It should be understood that, for clarity, certain features of the disclosed compositions and methods described herein in the context of separate embodiments may also be provided in combination in a single embodiment. Conversely, for brevity, various features of the disclosed compositions and methods described herein in the context of a single embodiment may also be provided individually or in any sub-combination.

[0065] As used herein, the singular form "a," "an," and "the" include plural forms.

[0066] As used herein, C max refers to the geometric mean maximum concentration of the active agent. This can be measured in vivo after administering a composition of the invention to a subject and measuring the plasma levels of the drug at various time points after administration.

[0067] As used herein, AUC refers to the area under the curve, which is the definite integral of the concentration of the active agent in plasma over time.

[0068] As used herein, "patient" or "subject" refers to mammals, including domestic animals, animals kept as livestock, and zoo animals. Conveniently, the "patient" or "subject" is a human.

[0069] It is understood that "treatment" or "therapy" includes prevention as well as alleviation of established symptoms of a disease. Thus, "treatment" or "therapy" of a disease, disorder or condition includes: (1) preventing or delaying the clinical symptoms of the disease, disorder or condition in a person who may have or is susceptible to the disease, disorder or condition but does not yet experience or display clinical or subclinical symptoms of the disease, disorder or condition, (2) inhibiting the disease, disorder or condition, i.e., preventing, reducing or delaying the development of the disease or its recurrence (in the case of maintenance therapy) or at least one clinical or subclinical symptom thereof, or (3) alleviating or relieving the disease, i.e., causing the disease, disorder or condition or at least one clinical or subclinical symptom thereof to disappear. "Treatment" and similar terms as used herein may specifically include reducing the severity and / or frequency of HSV-induced symptoms, eliminating HSV-induced symptoms and / or the underlying causes of the symptoms, reducing the frequency or likelihood of HSV-induced symptoms and / or their underlying causes, delaying, preventing and / or slowing the progression of HSV-induced conditions, and ameliorating or remedying damage caused directly or indirectly by HSV infection. The term "prevention" as used herein with respect to HSV infection or HSV-related diseases refers to reducing the likelihood of HSV infection. In one embodiment, "treating" or "treating" a state, disorder or condition refers to inhibiting the state, disorder or condition, i.e., preventing, reducing or delaying the development of the disease or its recurrence (in the case of maintenance treatment) or at least one clinical or subclinical symptom thereof, or (3) alleviating or attenuating the disease, i.e., causing the state, disorder or condition or at least one clinical or subclinical symptom thereof to subside. Conveniently, "treating" and similar terms refer to reducing the severity and / or frequency of HSV-induced symptoms, eliminating HSV-induced symptoms and / or potential causes of the symptoms, reducing the frequency or likelihood of HSV-induced symptoms and / or potential causes thereof, delaying, preventing and / or slowing the progression of HSV-induced conditions, and / or ameliorating or remedying damage caused directly or indirectly by HSV infection.

[0070] "Therapeutically effective amount" or "therapeutically effective dose" refers to the amount of a compound that, when administered to a patient or subject for treating a disease, is sufficient to achieve treatment of the disease. As used herein, the phrase "therapeutically effective dose" or "therapeutically effective amount" may specifically refer to an amount of a helicase-initase inhibitor administered to a patient or subject using the long-acting injectable pharmaceutical composition described herein, which is effective to achieve a particular biological or therapeutic outcome, such as, but not limited to, the biological or therapeutic outcomes disclosed, described, or exemplified herein. The therapeutically effective dose may vary depending on factors such as the disease state, age, sex, and weight of the individual, and the ability of the composition to elicit a desired response in the subject. Such outcomes include, but are not limited to, alleviation, remission, and / or regression of conditions caused by or associated with HSV, or prevention of the development of conditions caused by or associated with HSV, as determined by any suitable method in the art.

[0071] When values ​​are expressed as approximations using the antecedent "about," it is understood that the particular value forms another embodiment. Furthermore, unless otherwise stated or inferred, the term "about" refers to a variation of ±10% from the nominal value.

[0072] When the term "about" is used to refer to a numerical range, a cutoff value, or a specific value, it is used to indicate that the value may vary from the listed value by up to 10%. Since many of the numerical values ​​used herein are determined experimentally, it will be understood by those skilled in the art that such determinations may and usually will vary between different experiments. The values ​​used herein should not be considered to be overly limited due to such inherent variations. Therefore, the term "about" is used to cover variations of ±10% or less, ±5% or less, ±1% or less, ±0.5% or less, or ±0.1% or less from the specified value.

[0073] In different places of this specification, numerical value is disclosed in the form of group or range. In particular, the specification is intended to include all individual subcombinations of members of such groups and ranges and any combination of various endpoints of such groups or ranges. For example, the integer in the range of 0 to 40 is particularly intended to disclose 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39 and 40, and the integer in the range of 1 to 20 is particularly intended to disclose 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 and 20 individually.

[0074] As used herein, when a composition is described as having, including, or comprising particular components, or when a method is described as having, including, or comprising particular process steps, it is contemplated that the compositions of the present teachings also consist essentially of or consist of those components, and that the methods of the present teachings also consist essentially of or consist of those process steps.

[0075] Any and all examples or exemplary language used herein, such as "for example," "including," or "such as," are intended only to better illustrate the present teachings and are not intended to limit the scope of the present invention unless otherwise stated. No language in the specification should be construed as indicating any non-stated element is essential to the practice of the present teachings.

[0076] Helicase-primase inhibitors

[0077] The expression "helicase-primase inhibitor" in the context of the present invention means any compound or agent capable of reducing viral replication by inhibiting the viral complex consisting of DNA helicase, DNA primase and cofactor subunits. The helicase-primase complex is utilized by herpes viruses, and thus a helicase-primase inhibitor will have antiviral activity against one or more herpes viruses, such as one or more of herpes simplex virus subtypes 1 and 2 (HSV-1, HSV-2), varicella zoster virus (VZV), Epstein-Barr virus (EBV), cytomegalovirus (CMV) and human herpes virus 6-8 (HHV 6-8).

[0078] In one embodiment, the helicase-primase inhibitor has antiviral activity against HSV-1 and / or HSV-2.

[0079] In one embodiment, the helicase-primase inhibitor has an in vitro EC of 1.5 for HSV-1 and / or HSV-2 (suitably for both HSV-1 and HSV-2). 50 The value is less than about 0.1 μM. Suitably, the helicase-primase inhibitor has an in vitro EC of 50 The value is less than about 0.075 μM or less than about 0.05 μM. More suitably, the helicase-primase inhibitor has an in vitro EC of 50 More preferably, the helicase-primase inhibitor has an in vitro EC of less than about 0.030 μM for HSV-1 and / or HSV-2. 50 The value is less than about 0.020 μM. In vitro EC against HSV-1 and / or HSV-2 50 The value can be determined according to methods known to those skilled in the art, such as the method disclosed in Field et al. (2013, Antiviral Res. 100, pp. 297-299). 50 The value can also be determined according to the determination method described in the Examples section of this application.

[0080] In vivo, helicase-primase inhibitors may bind to plasma proteins. In one embodiment, the EC of a helicase-primase inhibitor (e.g., priligy) is 50 Values ​​may correspond to unbound plasma concentrations of the active agent.

[0081] In one embodiment, the helicase-primase inhibitor shows no or low levels of carbonic anhydrase inhibition, e.g., inhibition of carbonic anhydrase I and / or carbonic anhydrase II. Carbonic anhydrase inhibition can be measured using a carbonic anhydrase I assay as described by Katritzky et al. (J. Med. Chem. 1987, 30: 2058) and a carbonic anhydrase I assay as described by Iyer et al. (J. Biomol. Screen 2006, 11: 782). In one embodiment, the helicase-primase inhibitor has an IC of greater than 2.0 μM, greater than 3.0 μM, and most conveniently greater than 5.0 μM when measured in either or both assays. 50 .

[0082] The human biological terminal half-life of the helicase-initiase inhibitor used in the composition of the present invention is 10 hours or longer. The human biological terminal half-life of the helicase-initiase inhibitor can be derived from the terminal elimination phase of the pharmacokinetic curve after intravenous administration of the helicase-initiase inhibitor. The human biological terminal half-life of priligy is up to 80 hours (Wald et al., (2014) New England Journal of Medicine 370, p.201-210). In one embodiment, the human biological terminal half-life of the helicase-initiase inhibitor is 20 hours or longer, such as 30 hours or longer, 40 hours or longer, 50 hours or longer, 60 hours or longer, 70 hours or longer, or 80 hours or longer. Suitably, the human biological terminal half-life of the helicase-initiase inhibitor is 30 hours or longer.

[0083] In one embodiment, the helicase-primase inhibitor or its pharmaceutically acceptable salt has low water solubility. Conveniently, the solubility of the compound in water or aqueous solvent system (measured at room temperature) is less than 100 μg / ml, less than 50 μg / ml, suitably less than 25 μg / ml, more suitably less than 20 μg / ml. Suitably, the solubility of the compound in water at about pH 7.0 (measured at room temperature) is less than 100 μg / ml, less than 50 μg / ml, suitably less than 25 μg / ml, more suitably less than 20 μg / ml.

[0084] Examples of helicase-primase inhibitors include:

[0085] N-methyl-N-(4-methyl-5-sulfamoyl-1,3-thiazol-2-yl)-2-[4-(pyridin-2-yl)phenyl]acetamide (priligyvir; WO 01 / 47904);

[0086] (S)-2-(2′,5′-difluoro-[1,1′-biphenyl]-4-yl)-N-methyl-N-(4-methyl-5-(S-methylsulfonimidosulfon-imidoyl)thiazol-2-yl)acetamide (IM-250; WO 2017 / 174640 and WO 2019 / 068817A1); and

[0087] N-[2-[4-(2-Aminothiazol-4-yl)anilino]-2-oxo-ethyl]-N-[(1S)-1-phenylethyl]pyridine-4-carboxamide (BILS-179BS; WO 97 / 24343).

[0088] The helicase-primase inhibitors of the present invention include the parent compound and any pharmaceutically acceptable salt of the parent compound.

[0089] In cases where the helicase-primase inhibitor is sufficiently basic or acidic to form stable pharmaceutically acceptable acid or base salts, it may be appropriate to prepare and administer the helicase-primase inhibitor as a pharmaceutically acceptable salt.

[0090] Pharmaceutically acceptable salts can be obtained using standard procedures well known in the art, for example by reacting a sufficiently basic compound (e.g., an amine) with a suitable acid providing a physiologically acceptable anion. Alkali metal (e.g., sodium, potassium, or lithium) or alkaline earth metal (e.g., calcium) salts of carboxylic acids can also be prepared.

[0091] Examples of pharmaceutically acceptable salts are inorganic and organic acid addition salts formed with acids which form physiologically acceptable anions, such as malate, oxalate, chloride, bromide, iodide, nitrate, sulfate, bisulfate, phosphate, acid phosphate, isonicotinate, acetate, lactate, salicylate, citrate, tartrate, oleate, tannate, pantothenate, bitartrate, ascorbate, succinate, maleate, gentisinate, fumarate, gluconate, glucaronate, saccharate, formate, benzoate, glutamate, methanesulfonate, ethanesulfonate, benzenesulfonate, p-toluenesulfonate and pamoate (i.e., 1,1'-methylene-bis-(2-hydroxy-3-naphthoate)) salts.

[0092] Helicase-primase inhibitors that are acidic in nature may be able to form base salts with various pharmacologically acceptable cations. Examples of such salts include alkali metal or alkaline earth metal salts, particularly calcium, magnesium, sodium, lithium, zinc, potassium and iron salts.

[0093] It is also to be understood that certain inhibitors of the invention may exhibit polymorphism, and that the invention encompasses all such forms (including anhydrous / unsolvated forms, solvates, and hydrates).

[0094] In one embodiment, the helicase-initiase inhibitor is present in the composition in the form of a crystalline solid. A crystalline solid form is a solid material in which the components of the solid material are arranged in a highly ordered microstructure to form a lattice extending in all directions. The crystalline solid can exist in any suitable form, such as an anhydrate, hydrate or solvate form. It should also be understood that the helicase-initiase inhibitor of the present invention can be pure, substantially pure, or have a purity level greater than 75% relative to other crystalline forms, such as greater than 85%, greater than 90%, greater than 95%, greater than 97%, greater than 98% or greater than 99% (e.g., measured by X-ray powder diffraction).

[0095] It has been found that certain crystalline solid forms of helicase-primase inhibitors, such as certain crystalline solid forms of priligy, are surprisingly well suited for the long-acting injectable depot compositions of the present invention.

[0096] In one embodiment, the helicase-primase inhibitor is priligy, or a pharmaceutically acceptable salt thereof. Priligy is N-methyl-N-(4-methyl-5-sulfamoyl-1,3-thiazol-2-yl)-2-[4-(pyridin-2-yl)phenyl]acetamide, having the following structure:

[0097]

[0098] It is understood that priligy has potent antiviral activity against HSV-1 and HSV-2, with EC 50 The values ​​are 0.026 μM and 0.029 μM, respectively (Field et al., (2013) Antiviral Res. 100, pp. 297-299).

[0099] In one embodiment, the helicase-initiation enzyme inhibitor is priligy free base. The free base of priligy has low water solubility and is considered unsuitable for providing a long-term stable formulation (WO 2013 / 045491). However, the applicant has found that the free base compound is particularly suitable for being formulated into a long-acting injectable reservoir composition according to the present invention.

[0100] In suitable embodiments, the helicase-primase inhibitor is priligy free base and the free base is present in the composition as a crystalline solid. In one embodiment, the priligy crystalline free base is anhydrous (anhydrate) or hydrated form. Suitably, the priligy crystalline free base is a hydrate. Suitably, the priligy crystalline free base is a hemihydrate, a dihydrate or a sesquihydrate. More suitably, the priligy crystalline free base is a dihydrate or a sesquihydrate. Most suitably, the priligy crystalline free base is a sesquihydrate, as disclosed in the embodiments.

[0101] In one embodiment, the helicase-primase inhibitor is compound 1 represented below:

[0102]

[0103] or a pharmaceutically acceptable salt thereof.

[0104] The compound shown above is 2-(3-(2',5'-difluoro-[1,1'-biphenyl]-4-yl)-2-oxotetrahydropyrimidin-1(2H)-yl)-4-methylthiazole-5-sulfonamide (also referred to herein as 'Compound 1') and is a potent helicase-primase inhibitor.

[0105] In vitro EC of compound 1 against HSV-1 50 The in vitro EC50 value for HSV-1 and / or HSV-2 is about 0.019 μM, and the in vitro EC50 value for HSV-2 is about 0.011 μM. The in vitro EC50 value for HSV-1 and / or HSV-2 can be determined according to methods known to those skilled in the art. 50 Values, for example, the method disclosed in Field et al. (2013, Antiviral Res. 100, pp. 297-299). In vitro EC can also be determined according to the assay described in the Examples section of this application. 50 value.

[0106] Compound 1 was found to have a predicted human terminal biological half-life of more than 180 hours (see Examples 13 and 13A).

[0107] Compound 1 shows no or low levels of carbonic anhydrase inhibition, such as inhibition of carbonic anhydrase I and / or carbonic anhydrase II. Carbonic anhydrase inhibition can be measured using the carbonic anhydrase I assay described by Katritzky et al. (J. Med. Chem. 1987, 30: 2058) and the carbonic anhydrase I assay described by Iyer et al. (J. Biomol. Screen 2006, 11: 782).

[0108] It will be appreciated that Compound 1 may exhibit polymorphic forms and that the present invention encompasses all such forms (including anhydrous / unsolvated forms, solvates and hydrates).

[0109] In one embodiment, compound 1 is present in the composition in the form of a crystalline solid. The crystalline solid can be present in any suitable form, such as an anhydrate, a hydrate, or a solvate. It should also be understood that compound 1 can be pure, substantially pure, or have a purity level greater than 75% relative to other crystalline forms, such as greater than 85%, greater than 90%, greater than 95%, greater than 97%, greater than 98% or greater than 99% (e.g., measured by X-ray powder diffraction).

[0110] In a convenient embodiment, the helicase-primase inhibitor present in the composition of the invention is a free form of compound 1, and the free form is present in the composition in the form of a crystalline solid. In a convenient embodiment, the crystalline free form of compound 1 is an anhydrous form. It has been found that certain crystalline solid forms of the helicase-primase inhibitor compound 1 are unexpectedly well suited for the long-acting injectable depot composition of the invention. Suitably, the crystalline free form of compound 1 is form A or form C, as described in the examples. More suitably, the crystalline free form of compound 1 is form C, as described in the examples.

[0111] Pharmaceutical composition of the present invention

[0112] In a first aspect, the present invention provides a long-acting injectable depot pharmaceutical composition comprising a helicase-initiase inhibitor or a pharmaceutically acceptable salt thereof; and one or more pharmaceutically acceptable excipients; wherein the helicase-initiase inhibitor has a terminal biological half-life in humans of 10 hours or more.

[0113] As used herein, a "long-acting injectable depot pharmaceutical composition" refers to a pharmaceutical composition which is formulated to be administered to a subject (suitably a human) by subcutaneous or intramuscular route, wherein upon administration a depot is formed from which the active ingredient (helicase-priming enzyme inhibitor) of the composition is released over an extended period of time.

[0114] In one embodiment, extended release refers to the continuous release of the active ingredient over a period of at least 10 days, such as at least 14 days, at least 21 days, at least 28 days, at least 56 days, at least 72 days, at least 96 days, at least 120 days, at least 180 days, at least 240 days, or at least 300 days. Extended release of the active ingredient means that the plasma level of the active ingredient can be maintained at a therapeutically effective plasma concentration for at least 10 days, such as at least 14 days, at least 21 days, at least 28 days, at least 56 days, at least 72 days, at least 96 days, at least 120 days, at least 180 days, at least 240 days, or at least 300 days after administration of the composition, which will be understood by those skilled in the art to correspond to "long-acting".

[0115] It will be appreciated that the rate of release of the helicase-primase may vary over the extended release period, e.g., a brief "initial burst" of active agent may be observed shortly after administration, followed by a period of lower release. However, extended release means that the plasma level of the helicase-primase inhibitor may be maintained at a therapeutically effective plasma concentration for most, if not all, of the extended release period.

[0116] In one embodiment, the pharmaceutical composition provides a brief "initial burst" of the active agent shortly after administration, followed by a period of lower release. Advantageously, Applicants have discovered that certain compositions of the present invention are able to provide a rapid initial release to achieve high plasma levels of the helicase-priming enzyme inhibitor, followed by a sustained release to maintain plasma levels at therapeutically effective concentrations for an extended period of time.

[0117] In one embodiment, the pharmaceutical composition releases less than 20% (suitably less than 10%) of the total weight of active agent contained in the pharmaceutical composition within 24 hours (suitably 48 hours) after administration.

[0118] Advantageously, the applicant has found that the extended release provided by certain compositions of the present invention and the continuous plasma levels of therapeutically effective concentrations produced over an extended period of time allow the use of lower than expected doses of the helicase-primase inhibitor. In one embodiment, the dose of the long-acting injectable reservoir pharmaceutical composition administered in a single administration is about 100 to 1200 mg (suitably 300 to 900 mg) of the helicase-primase inhibitor or a pharmaceutically acceptable salt thereof, which dose is capable of providing extended release over a period of at least 14 days (suitably at least 28 days, at least 56 days, at least 96 days, at least 120 days, at least 180 days, at least 240 days or at least 300 days).

[0119] In one embodiment, the dose of the long-acting injectable depot pharmaceutical composition capable of providing extended release over a period of at least 1 month is about 100 to 900 mg (suitably 100 to 600 mg) of the helicase-primase inhibitor or a pharmaceutically acceptable salt thereof administered in a single administration.

[0120] In one embodiment, the dose of about 200 to 900 mg (suitably 200 to 600 mg) of the helicase-primase inhibitor or a pharmaceutically acceptable salt thereof administered in a single administration of a long-acting injectable depot pharmaceutical composition capable of providing extended release over a period of at least 2 months is about 200 to 900 mg (suitably 200 to 600 mg).

[0121] In one embodiment, the dose of the long-acting injectable depot pharmaceutical composition capable of providing extended release over a period of at least 3 months is about 300 to 900 mg (suitably 300 to 600 mg) of the helicase-primase inhibitor or a pharmaceutically acceptable salt thereof administered in a single administration.

[0122] In one embodiment, the dose of the long-acting injectable depot pharmaceutical composition capable of providing extended release over a period of at least 6 months is about 300 to 1200 mg (suitably 300 to 900 mg) of the helicase-primase inhibitor or a pharmaceutically acceptable salt thereof administered in a single administration.

[0123] In some embodiments, it has been found that some long-acting injectable reservoir compositions of the present invention exhibit high drug loading capacity. This provides many advantages, including that the composition can be administered with a low injection volume. In one embodiment, the dosage is administered in a single administration of a long-acting injectable reservoir pharmaceutical composition with an injection volume of about 0.1 to about 5 mL. Conveniently, the dosage is administered with an injection volume of about 0.1 to about 3 mL, about 0.1 to about 1.5 mL, or about 0.1 to about 0.75 mL. More conveniently, the dosage is administered with an injection volume of about 0.1 to about 0.5 mL.

[0124] In a convenient embodiment, the pharmaceutical composition is designed for subcutaneous or intramuscular injection. In another aspect of the invention, the pharmaceutical composition is for subcutaneous administration. In another aspect of the invention, the pharmaceutical composition is for intramuscular administration. Suitably, the dosage of the helicase-initiase inhibitor is administered subcutaneously with an injection volume of about 0.1 to about 3 mL, or about 0.1 to about 1.5 mL. Suitably, the dosage of the helicase-initiase inhibitor is administered intramuscularly with an injection volume of about 0.1 to about 5 mL, or about 0.1 to about 3 mL.

[0125] The benefits of the present invention are not limited to a specific type of long-acting injectable depot pharmaceutical composition with a specific drug release mechanism. Pharmaceutical compositions suitable for subcutaneous and / or intramuscular administration can be aqueous-based depots, aqueous-based suspension depots, formulations based on solutions in organic solvents, depots based on suspensions in organic solvents, gel-based depots, in situ gelling depot formulations, oil-based depots, emulsion-based depots, depots based on monolithic polymers, depots based on microparticle polymers, or solid implant depots.

[0126] In one embodiment, the pharmaceutical composition of the present invention comprises a rate control agent to help provide an extended release of the helicase-initiase inhibitor. In one embodiment, the rate control agent comprises at least one biodegradable polymer. Suitable biodegradable polymers are polymers that can be decomposed under physiological conditions so that the polymer and its degradation products do not cause any unacceptable toxicity or immune response. Such polymers can be natural or synthetic in origin. Examples of natural biodegradable polymers include polysaccharides, such as chitosan, alginate and dextran, or polyesters such as polyhydroxyalkanoates. Examples of synthetic biodegradable polymers include polymers of lactic acid and glycolic acid and copolymers thereof. In one embodiment, the biodegradable polymer is poly (lactic acid-co-glycolic acid) or polylactic acid, wherein each polymer is terminated with an acid or ester group. Suitably, the biodegradable polymer is poly (lactic acid-co-glycolic acid). Poly (lactic acid-co-glycolic acid) is also referred to herein as PLGA. Conveniently, the biodegradable polymer is PLGA with an acid group at the end. In one embodiment, the biodegradable polymer is PLGA, wherein the PLGA has a lactic acid content of about 5 to 95% by weight, with the remainder being glycolic acid.

[0127] In one convenient embodiment, the pharmaceutical composition of the invention is an aqueous-based suspension, a formulation based on a solution in an organic solvent, or an in situ gelling depot formulation.

[0128] In a preferred embodiment, the pharmaceutical composition of the present invention is a water-based suspension reservoir. Surprisingly, it has been found that certain water-based suspension reservoir compositions provide active ingredient plasma levels of therapeutically effective plasma concentrations for a long time without the need for a rate controller. This can be particularly advantageous because it allows high drug loads, smaller dosages, and lower excipient levels. In one embodiment, the pharmaceutical composition of the present invention does not include a rate controller or includes less than 20% of the total weight of the composition or less than 15, 10, 5, 3, 2, or 1% of the rate controller. Conveniently, the pharmaceutical composition of the present invention does not include a rate controller such as a biodegradable polymer.

[0129] It has also been found that release may be affected by the particle size of the helicase-priming enzyme inhibitor in a water-based suspension depot composition. Suitable methods for determining the particle size distribution of a sample will be clear to those skilled in the art, including techniques such as laser diffraction. The term Dx (or Dv x ) refers to the maximum particle size (inclusive) that accounts for x% of the total volume of the material in the sample. For example, D50 (or Dv 50 ) of 10 μm means that 50% of the sample has a particle size of 10 μm or less.

[0130] In one embodiment, the particle size volume distribution of the suspended particles of the active agent (helicase-initiase inhibitor) in the water-based suspension reservoir composition is such that when measured by laser diffraction analysis, less than 10% of the particles have a size greater than 9 microns. In one embodiment, the water-based suspension reservoir composition contains active agent particles having a D10 of 0.5 to 2.5 μm, a D50 of 2 to 5 μm, and / or a D90 of 4 to 13 μm. In one embodiment, the water-based suspension reservoir composition contains active agent particles having a D10 of 0.5 to 2.5 μm, a D50 of 3 to 5 μm, and / or a D90 of 8 to 13 μm. In one embodiment, the water-based suspension reservoir composition contains active agent particles having a D10 of 0.5 to 2.0 μm, a D50 of 2 to 3 μm, and / or a D90 of 4 to 8 μm. In one embodiment, the aqueous suspension depot composition comprises active agent particles having a D10 of 1 to 2 μm, a D50 of 4 to 5 μm, and / or a D90 of 10 to 11 μm. In one embodiment, the aqueous suspension depot composition comprises active agent particles having a D90 of 10 to 5000 nm, such as 20 to 2000 nm, or 50 to 1000 nm. Suitably, the helicase-inhibitor is priligy.

[0131] In one embodiment, the helicase-primase inhibitor is compound 1 and the aqueous suspension reservoir composition comprises active agent particles having a D10 of 0.3 to 1.5 μm, a D50 of 2 to 4 μm, and / or a D90 of 8 to 16 μm. In one embodiment, the aqueous suspension reservoir composition comprises active agent particles having a D10 of 0.3 to 0.7 μm, a D50 of 2 to 3 μm, and / or a D90 of 12 to 16 μm.

[0132] In one embodiment, the helicase-primase inhibitor is priligy and the pharmaceutical composition is a water-based micron suspension reservoir composition. Suitably, the particle size volume distribution of the priligy suspended particles in the water-based micron suspension reservoir composition is such that when measured by laser diffraction analysis, less than 10% of the particles have a size greater than 20 μm. In one embodiment, the priligy particles contained in the water-based micron suspension reservoir composition have a D90 of 4 to 20 μm, suitably 5 to 15 μm, more suitably 6 to 10 μm. In one embodiment, the priligy particles contained in the water-based micron suspension reservoir composition have a D10 of 0.3 to 3 μm, a D50 of 2 to 6 μm, and / or a D90 of 5 to 15 μm. In one embodiment, the aqueous microsuspension depot composition comprises priligy particles having a D10 of 1.5 to 2.5 μm, a D50 of 3 to 5 μm, and / or a D90 of 6 to 10 μm.

[0133] In one embodiment, the helicase-primase inhibitor is priligy and the pharmaceutical composition is a water-based nanosuspension reservoir composition. Suitably, the particle size volume distribution of the priligy suspended particles in the water-based nanosuspension reservoir composition is such that when measured by laser diffraction analysis, less than 10% of the particles have a size greater than 700 nm. In one embodiment, the priligy particles contained in the water-based nanosuspension reservoir composition have a D90 of 1 to 700 nm, suitably 1 to 500 nm, more suitably 1 to 300 nm.

[0134] In one embodiment, the helicase-primase inhibitor is compound 1 and the pharmaceutical composition is an aqueous micron suspension reservoir composition. Suitably, the particle size volume distribution of the suspended particles of compound 1 in the aqueous micron suspension reservoir composition is such that when measured by laser diffraction analysis, less than 10% of the particles have a size greater than 15 μm. In one embodiment, the aqueous micron suspension reservoir composition contains compound 1 particles having a D90 of 4 to 13 μm, suitably 4 to 10 μm, more suitably 4 to 8 μm. In one embodiment, the aqueous micron suspension reservoir composition contains compound 1 particles having a D10 of 0.3 to 2 μm, a D50 of 2 to 4 μm, and / or a D90 of 4 to 13 μm. In one embodiment, the aqueous micron suspension reservoir composition contains compound 1 particles having a D10 of 0.3 to 1.5 μm, a D50 of 2 to 3.5 μm, and / or a D90 of 4 to 8 μm.

[0135] In one embodiment, the helicase-primase inhibitor is compound 1 and the pharmaceutical composition is a water-based nanosuspension reservoir composition. Suitably, the particle size volume distribution of the suspended particles of compound 1 in the water-based nanosuspension reservoir composition is such that when measured by laser diffraction analysis, less than 10% of the particles have a size greater than 500nm. In one embodiment, the compound 1 particles contained in the water-based nanosuspension reservoir composition have a D90 of 1 to 400nm, suitably 1 to 300nm, more suitably 1 to 200nm. In one embodiment, the compound 1 particles contained in the water-based nanosuspension reservoir composition have a D10 of 1 to 100nm, a D50 of 1 to 200nm, and / or a D90 of 1 to 300nm. In one embodiment, the compound 1 particles contained in the water-based nanosuspension reservoir composition have a D10 of 1 to 100nm, a D50 of 1 to 150nm, and / or a D90 of 1 to 200nm. In one embodiment, the water-based nanosuspension depot composition comprises Compound 1 particles having a D10, D50, and D90 of 1 to 100 nm.

[0136] In one embodiment, the pharmaceutical composition of the present invention is a preparation based on a solution in an organic solvent. The organic solvent can be any suitable organic solvent, which can dissolve the helicase-initiase inhibitor and has biocompatibility. Suitable biocompatible organic solvents diffuse from the preparation after the preparation is injected into the body, causing the preparation to solidify. Suitably, the biocompatible solvent is nontoxic, miscible with water, and will not cause severe tissue irritation or necrosis at the injection / implantation site. Examples of these solvents include benzyl alcohol, benzyl benzoate, N-methyl-2-pyrrolidone, 2-pyrrolidone, ethanol, propylene glycol, acetone, methyl acetate, ethyl acetate, methyl ethyl ketone, dimethylacetamide, dimethylformamide, dimethyl sulfoxide, tetrahydrofuran, caprolactam, oleic acid and l-dodecylazacycloheptane-2-one. In one embodiment, the organic solvent is selected from N-methylpyrrolidone (NMP), dimethylacetamide (DMA), dimethyl sulfoxide (DMSO), benzyl alcohol, benzyl benzoate and mixtures thereof. Formulations based on solutions in organic solvents may optionally further comprise an aqueous medium (eg, aqueous buffer or saline) and / or a surfactant (eg, a polyethylene glycol fatty ester, for example, PEG-15 hydroxystearate).

[0137] In one embodiment, the helicase-primase inhibitor is compound 1 and the pharmaceutical composition is a formulation based on a solution in an organic solvent. In one embodiment, the formulation based on a solution in an organic solvent comprises compound 1 dissolved in an organic solvent selected from the group consisting of N-methylpyrrolidone (NMP), dimethylacetamide (DMA), dimethyl sulfoxide (DMSO), benzyl alcohol, benzyl benzoate, and mixtures thereof, preferably N-methylpyrrolidone (NMP), dimethylacetamide (DMA) and dimethyl sulfoxide (DMSO), such as NMP. In one embodiment, the formulation based on a solution in an organic solvent comprising compound 1 may optionally further comprise an aqueous medium (such as an aqueous buffer or saline) and / or a surfactant (such as a polyethylene glycol fatty ester, e.g., PEG-15 hydroxystearate). In one embodiment, the composition comprises 5-95% v / v (e.g., 5-75%, 5-50%, 5-20%, or 5-15% v / v) of an organic solvent selected from the group consisting of N-methylpyrrolidone (NMP), dimethylacetamide (DMA), dimethyl sulfoxide (DMSO), and mixtures thereof. In one embodiment, the composition comprises compound 1 at a concentration of 1-500 mg / mL, such as 10-400, 100-400, or 200-300 mg / mL. In one embodiment, the composition comprises compound 1 at a concentration of 100-400 mg / mL and 5-95% v / v of NMP as an organic solvent.

[0138] In one embodiment, the helicase-primase inhibitor is priligy free base and the pharmaceutical composition is a formulation based on a solution in an organic solvent. In one embodiment, the formulation based on a solution in an organic solvent comprises compound 1 dissolved in an organic solvent selected from the group consisting of N-methylpyrrolidone (NMP), dimethylacetamide (DMA), dimethyl sulfoxide (DMSO), benzyl alcohol, benzyl benzoate and mixtures thereof, preferably N-methylpyrrolidone (NMP), dimethylacetamide (DMA) and dimethyl sulfoxide (DMSO). In one embodiment, the formulation based on a solution in an organic solvent comprising priligy free base may optionally further comprise an aqueous medium (such as an aqueous buffer or saline) and / or a surfactant (such as a polyethylene glycol fatty ester, e.g., PEG-15 hydroxystearate). In one embodiment, the composition comprises 5-95% v / v (such as 5-75%, 5-50%, 5-20%, or 5-15% v / v) of an organic solvent selected from the following: N-methylpyrrolidone (NMP), dimethylacetamide (DMA), dimethyl sulfoxide (DMSO), and mixtures thereof. In one embodiment, the composition comprises 1-500 mg / mL of priligy free base, such as 10-400, 50-300 or 50-200 mg / mL. In one embodiment, the composition comprises 10-400 mg / mL (such as 100-400 mg / mL) of priligy free base and 5-95% v / v of NMP as an organic solvent.

[0139] In one embodiment, pharmaceutical composition of the present invention is an in situ gelation reservoir preparation.Term used herein " in situ gelation reservoir preparation " refers to the preparation comprising helicase-initiator inhibitor, biodegradable polymer (for example, biodegradable polymer described as rate control agent above) and biocompatible solvent, and the preparation is delivered to the patient in the form of injectable liquid, but as liquid solvent diffuses in vivo, the preparation can solidify into solid reservoir preparation.Suitable biocompatible solvent refers to any solvent that the component of preparation can dissolve, and after the preparation is injected into the body, the solvent can diffuse from the preparation, thereby causing the preparation to solidify.Conveniently, the solvent of biodegradable polymer is nontoxic, water miscible, and is biocompatible in other aspects.Solvent must also be biocompatible, so that they can not cause serious tissue irritation or necrosis at the implantation site.In addition, solvent should be miscible with water, so that it can diffuse into body fluid quickly, and allow water to penetrate into the polymer solution and solidify or solidify it. Examples of such solvents include benzyl alcohol, N-methyl-2-pyrrolidone, 2-pyrrolidone, ethanol, propylene glycol, acetone, methyl acetate, ethyl acetate, methyl ethyl ketone, dimethylformamide, dimethyl sulfoxide, tetrahydrofuran, caprolactam, oleic acid and 1-dodecylazacycloheptan-2-one.

[0140] The composition of the present invention includes one or more pharmaceutically acceptable excipients. The long-acting injectable reservoir pharmaceutical composition of the present invention includes at least one excipient selected from solvents, cosolvents, wetting agents or suspending agents, isotonic agents, pH regulators, stabilizers, emulsifiers and viscosity regulators.

[0141] Suitable wetting agents or suspending agents included in the formulations of the present invention include cellulose derivatives, polyvinyl pyrrolidone, polysorbate 20 and polysorbate 80, lecithin, polyoxyethylene sorbitan fatty acid esters and polyoxyethylene sorbitan fatty acid esters. Polysorbate 20 is used alone or in combination. Suitably, polysorbate 80 is present in the composition. Suitably, the composition comprises about 0.2% by weight to about 5% by weight of polysorbate 80, more suitably about 0.2% by weight to about 2% by weight of polysorbate 80.

[0142] The isotonic agent that can be included in the long-acting injectable reservoir pharmaceutical composition of the present invention is, for example, sodium chloride, dextrose, mannitol, sorbitol, lactose, and / or sodium sulfate. Suitably, the isotonic agent is sodium chloride. Suitably, the isotonic agent is mannitol.

[0143] The pH adjuster contained in the long-acting injectable depot-type pharmaceutical composition of the present invention may be hydrochloric acid or sodium hydroxide, but is not limited thereto.

[0144] The depot preparation of the present invention may further contain a preservative, which may be selected from benzoic acid, benzyl alcohol, butylated hydroxyanisole, butylated hydroxytoluene, chlorobutanol, gallic acid esters, hydroxybenzoic acid esters, EDTA, phenol, parabens, methylparaben, propylparaben, butylparaben, benzalkonium chloride, thimerosal, m-cresol or chlorobutanol, etc.

[0145] In some embodiments, the long-acting injectable reservoir pharmaceutical composition of the present invention comprises at least one viscosity modifier. In some cases, the viscosity modifier is selected from sodium carboxymethylcellulose, methylcellulose, hydroxypropyl cellulose, carboxymethylcellulose calcium, cross-linked carboxymethylcellulose, hydroxyethyl cellulose, hydroxypropyl methylcellulose, polyvinyl alcohol, gum arabic, gelatin and polyvinyl pyrrolidone. Suitably, the composition comprises about 0.1wt% to about 5wt% of viscosity modifier. In some embodiments, the viscosity modifier is selected from sodium carboxymethylcellulose and / or hydroxypropyl cellulose and / or polyvinyl pyrrolidone.

[0146] The long-acting injectable depot pharmaceutical composition of the present invention can be stored by freeze drying, for example in a suitable vial, and can be resuspended for injection immediately before injection. The composition can be stored in a pre-filled syringe.

[0147] Advantageously, the compositions of the present invention can exhibit good storage stability. Thus, in one embodiment, the composition is stable for at least four weeks, e.g., at least eight weeks or at least twelve weeks. In this context, "stable" may refer to the physical and / or chemical stability of the active agent (helicase-initiase inhibitor) in the long-acting injectable reservoir pharmaceutical composition. "Stable" may also refer to the release profile or pharmacokinetics of the composition being maintained during the indicated storage period. "Stable" may also refer to the appearance of the composition; for example, for a water-based suspension reservoir composition, it may refer to the uniformity of the suspension being maintained during the indicated storage period.

[0148] Physical stability involves the active agent remaining in the same physical form. When the long-acting injectable depot pharmaceutical composition is an aqueous suspension and the active agent is suspended as a crystalline solid, the physical form of the suspended solid (as determined by XRPD) does not change during the stated storage period.

[0149] Chemical stability relates to the low levels of impurities formed during the stated storage period. Typically, these impurities are associated with the active agent and can be measured by suitable techniques (eg HPLC or LC-MS).

[0150] The release of the active agent from the composition of the present invention can be determined by methods known in the art. For example, the release rate can be determined using an in vitro dissolution test that simulates an aqueous physiological environment. The term "aqueous physiological environment" as used herein refers to the body of a warm-blooded animal, particularly the human body. In a specific embodiment, the composition of the present invention is designed for subcutaneous or intramuscular administration, and the term "aqueous physiological environment" of such a composition refers to the subcutaneous or intramuscular environment of such a body. These conditions can be simulated in vitro by placing the composition in an aqueous dissolution medium, which is optionally buffered to physiological pH and at a temperature of 35 to 40° C. The amount of active agent released in a given time period can be determined by sampling the dissolution medium and measuring the concentration of the active agent using a suitable analytical method (e.g., HPLC).

[0151] The release of the active ingredient in the composition of the present invention can also be determined by in vivo methods known in the art. For example, the in vivo release can be tested by measuring the plasma concentration over a predetermined time period to obtain a plasma concentration versus time curve of the target compound.

[0152] Other tests can also be used to determine the amount of active agent released in vivo. Animals (e.g., mice, rats, dogs, etc.) can be used as models to study release characteristics. For example, for subcutaneous or intramuscular compositions, the animal can receive the composition under study, and after a specified period of time, the animal can be killed, and the subcutaneous or intramuscular composition can be extracted / recovered and analyzed. By determining the content of the active agent remaining in the composition within a specified period of time, the amount and degree of release can be calculated.

[0153] Advantageously, the compositions of the present invention are capable of maintaining therapeutically effective plasma concentrations of the drug over an extended period of time.

[0154] Pharmacokinetics

[0155] The present invention provides a long-acting injectable depot pharmaceutical composition which has certain advantages over prior art compositions, namely, providing a stable and continuous level of plasma helicase-priming enzyme inhibitors over a long period of time, thereby reducing or even eliminating viral shedding and viral reactivation.

[0156] In one embodiment, the long-acting injectable depot pharmaceutical composition of the present invention maintains a geometric mean plasma concentration of the helicase-initase inhibitor of at least 25 ng / mL in the subject for at least 14 days after administration to a subject in need of treatment.

[0157] In one embodiment, the long-acting injectable reservoir pharmaceutical composition according to the present invention comprises a helicase-primase inhibitor, or a pharmaceutically acceptable salt thereof, which produces a plasma concentration of the helicase-primase inhibitor in the subject after administration of at least 25 ng / mL (such as at least 50 ng / mL, or at least 100 ng / mL), and lasts for at least 80% of the dosing interval (such as at least 85%, at least 90%, at least 95%, at least 98%, at least 99%, or substantially all). Suitably, the dosing interval is at least 10 days (such as at least 14 days, at least 21 days, at least 28 days, at least 56 days, at least 72 days, at least 96 days, at least 120 days, at least 180 days, at least 240 days, or at least 300 days). In one embodiment, the dosing interval is at least 14 days or at least 28 days. Suitably, the dosing interval is at least 56 days, at least at least 96 days, at least 120 days, at least 180 days, at least 240 days, or at least 300 days.

[0158] In one embodiment, the long-acting injectable reservoir pharmaceutical composition according to the present invention comprises pralivir free base and produces a plasma concentration of pralivir in the subject after administration of at least 25 ng / mL (such as at least 50 ng / mL, or at least 100 ng / mL), which lasts for at least 80% of the dosing interval (such as at least 85%, at least 90%, at least 95%, at least 98%, at least 99%, or substantially all). Suitably, the dosing interval is at least 10 days (such as at least 14 days, at least 21 days, at least 28 days, at least 56 days, at least 72 days, at least 96 days, at least 120 days, at least 180 days, at least 240 days, or at least 300 days). In one embodiment, the dosing interval is at least 14 days or at least 28 days. Suitably, the dosing interval is at least 56 days, at least at least 96 days, at least 120 days, at least 180 days, at least 240 days, or at least 300 days.

[0159] In one embodiment, the long-acting injectable reservoir pharmaceutical composition according to the present invention comprises a free form of compound 1 and produces a plasma concentration of compound 1 in a subject after administration of at least 25 ng / mL (such as at least 50 ng / mL, or at least 100 ng / mL), which lasts for at least 80% of the dosing interval (such as at least 85%, at least 90%, at least 95%, at least 98%, at least 99%, or substantially all). Suitably, the dosing interval is at least 10 days (such as at least 14 days, at least 21 days, at least 28 days, at least 56 days, at least 72 days, at least 96 days, at least 120 days, at least 180 days, at least 240 days, or at least 300 days). In one embodiment, the dosing interval is at least 14 days or at least 28 days. Suitably, the dosing interval is at least 56 days, at least at least 96 days, at least 120 days, at least 180 days, at least 240 days, or at least 300 days.

[0160] Helicase-primase inhibitors are usually subject to plasma protein binding. Depending on the degree of plasma protein binding, the free portion (unbound active agent) may be relatively low relative to the protein-bound portion. For example, priligy is usually subject to 97-98% protein binding, so the free portion may only account for 2-3% of the total plasma concentration. In one embodiment, the above-mentioned plasma concentration refers to the unbound plasma concentration. Therefore, in one embodiment, the long-acting injectable reservoir pharmaceutical composition of the present invention is administered to a subject in need of treatment, and the geometric mean unbound plasma concentration of the helicase-primase inhibitor (e.g., priligy) is maintained in the subject for at least 14 days. At least 25ng / mL. In one embodiment, after the long-acting injectable reservoir pharmaceutical composition of the present invention is administered to a subject in need of its treatment, the geometric mean total (unbound and protein-bound) plasma concentration of the helicase-primase inhibitor (e.g., priligy) is maintained in the subject for at least 1,500ng / mL (e.g., at least 2,000ng / mL, or at least 2,500ng / mL).

[0161] In one embodiment, the long-acting injectable depot pharmaceutical composition according to the present invention, after administration to a subject in need of treatment, maintains a geometric mean total (unbound and protein-bound) plasma concentration of Compound 1 in the subject of at least 500 ng / mL (e.g., at least 1000 ng / mL, at least 1100 ng / mL, at least 1,500 ng / mL, at least 2,000 ng / mL, or at least 2,500 ng / mL) for most of the time (if not all of the time) throughout at least 14 days. Suitably, the long-acting injectable depot pharmaceutical composition of the present invention, after administration to a subject in need of treatment, maintains a geometric mean total (unbound and protein-bound) plasma concentration of Compound 1 in the subject of at least 14 days (if not all of the time) for most of the time (if not all of the time) throughout at least 14 days in the subject.

[0162] Specific compositions of the present invention

[0163] In one embodiment, the present invention provides a long-acting injectable depot pharmaceutical composition comprising:

[0164] a) a helicase-primase inhibitor; and

[0165] b) one or more pharmaceutically acceptable excipients;

[0166] Wherein the helicase-initiase inhibitor has a human biological terminal half-life of 10 hours or more; and wherein the helicase-initiase inhibitor has a solubility (measured at room temperature) of less than 100 μg / ml in water or an aqueous solvent system. Suitably, the long-acting injectable reservoir pharmaceutical composition continuously releases the helicase-initiase inhibitor at a certain rate for at least 1 month after subcutaneous or intramuscular injection, and the rate obtains the therapeutic plasma concentration of the helicase-initiase inhibitor. Suitably, the long-acting injectable reservoir pharmaceutical composition comprises about 1wt.% to about 80wt.% of the helicase-initiase inhibitor. Suitably, the long-acting injectable reservoir pharmaceutical composition comprises about 10wt.% to about 40wt.% of the helicase-initiase inhibitor. Suitably, the long-acting injectable reservoir pharmaceutical composition comprises 300 to 900mg of the helicase-initiase inhibitor and the long-acting injectable reservoir pharmaceutical composition is administered with an injection volume of about 0.1 to about 3mL. Suitably, the long-acting injectable depot pharmaceutical composition comprises a water-based suspension or a formulation based on a solution in an organic solvent. Suitably, the water-based suspension is a microsuspension or a nanosuspension. Suitably, the long-acting injectable depot pharmaceutical composition does not comprise a rate controller, such as a biodegradable polymer, or comprises a rate controller less than 20% or less than 15, 10, 5, 3, 2 or 1% weight of the total composition.

[0167] In one embodiment, the present invention provides a long-acting injectable depot pharmaceutical composition comprising:

[0168] a) Priligy free base form (suitably as a hydrate, such as a hemihydrate, sesquihydrate or dihydrate); and

[0169] b) one or more pharmaceutically acceptable excipients;

[0170] The long-acting injectable reservoir pharmaceutical composition continuously releases priligy at a rate that provides a therapeutic plasma concentration of a helicase-initiation enzyme inhibitor for at least one month after subcutaneous or intramuscular injection.

[0171] In one embodiment, the present invention provides a long-acting injectable depot pharmaceutical composition comprising:

[0172] a) Priligy free base form (suitably as a dihydrate or sesquihydrate); and

[0173] b) one or more pharmaceutically acceptable excipients;

[0174] The long-acting injectable reservoir pharmaceutical composition continuously releases priligy at a rate that provides a therapeutic plasma concentration of a helicase-initiation enzyme inhibitor for at least one month after subcutaneous or intramuscular injection.

[0175] In one embodiment, the present invention provides a long-acting injectable depot pharmaceutical composition comprising:

[0176] a) Priligy free base form (suitably as a hemihydrate, dihydrate or sesquihydrate, such as a sesquihydrate); and

[0177] b) one or more pharmaceutically acceptable excipients;

[0178] Wherein the long-acting injectable reservoir pharmaceutical composition comprises 300 to 900mg of praviridovir and the long-acting injectable reservoir pharmaceutical composition is administered with an injection volume of about 0.1 to about 3mL. Suitably, the long-acting injectable reservoir pharmaceutical composition continuously releases praviridovir at a certain rate within at least 3 months (preferably at least 6 months) after subcutaneous or intramuscular injection, and the rate obtains the therapeutic plasma concentration of praviridovir. Suitably, the long-acting injectable reservoir pharmaceutical composition includes a preparation based on a solution in an organic solvent comprising praviridovir and wherein praviridovir is dissolved in an organic solvent selected from the group consisting of N-methylpyrrolidone (NMP), dimethylacetamide (DMA), dimethyl sulfoxide (DMSO), benzyl alcohol, benzyl benzoate and mixtures thereof, preferably N-methylpyrrolidone (NMP), dimethylacetamide (DMA) and dimethyl sulfoxide (DMSO). The composition optionally further comprises an aqueous medium (such as an aqueous buffer or saline) and / or a surfactant (such as a polyethylene glycol fatty ester, such as PEG-15 hydroxystearate). Suitably, the long-acting injectable reservoir pharmaceutical composition comprises a water-based suspension. Suitably, the water-based suspension is a nanosuspension.

[0179] Suitably, the long-acting injectable depot pharmaceutical composition does not comprise a rate-controlling agent, such as a biodegradable polymer, or comprises less than 20% or less than 15, 10, 5, 3, 2 or 1% by weight of a rate-controlling agent of the total composition. Suitably, the priligy free base form is a sesquihydrate.

[0180] In one embodiment, the present invention provides a long-acting injectable depot pharmaceutical composition comprising:

[0181] a) Priligy free base form (suitably as a dihydrate or sesquihydrate); and

[0182] b) one or more pharmaceutically acceptable excipients;

[0183] Wherein the long-acting injectable depot pharmaceutical composition comprises 300 to 900 mg of a helicase-initiation enzyme inhibitor, the long-acting injectable depot pharmaceutical composition is administered with an injection volume of about 0.1 to about 2 mL, and wherein the long-acting injectable depot pharmaceutical composition comprises an aqueous suspension (such as an aqueous micron suspension). Suitably, the priligy free base form is a sesquihydrate.

[0184] In one embodiment, the present invention provides a long-acting injectable depot pharmaceutical composition comprising:

[0185] a) Compound 1 free form (suitably anhydrous Form A or Form C); and

[0186] b) one or more pharmaceutically acceptable excipients;

[0187] The long-acting injectable depot pharmaceutical composition continuously releases Compound 1 at a rate that provides a therapeutic plasma concentration of Compound 1 for at least 1 month after subcutaneous or intramuscular injection. Suitably, the long-acting injectable depot pharmaceutical composition is administered by intramuscular injection.

[0188] In one embodiment, the present invention provides a long-acting injectable depot pharmaceutical composition comprising:

[0189] a) Compound 1 free form (suitably anhydrous Form A or Form C); and

[0190] b) one or more pharmaceutically acceptable excipients;

[0191] Wherein the long-acting injectable reservoir pharmaceutical composition comprises 300 to 900mg compound 1 and the long-acting injectable reservoir pharmaceutical composition is administered with an injection volume of about 0.1 to about 3mL. Suitably, the long-acting injectable reservoir pharmaceutical composition continuously releases compound 1 at a certain rate within at least 3 months (preferably at least 6 months) after subcutaneous or intramuscular injection, and the rate obtains the therapeutic plasma concentration of compound 1. Suitably, the long-acting injectable reservoir pharmaceutical composition includes a formulation based on a solution in an organic solvent comprising compound 1, and wherein compound 1 is dissolved in an organic solvent selected from the following: N-methylpyrrolidone (NMP), dimethylacetamide (DMA), dimethyl sulfoxide (DMSO), benzyl alcohol, benzyl benzoate and a mixture thereof, preferably N-methylpyrrolidone (NMP), dimethylacetamide (DMA) and dimethyl sulfoxide (DMSO). The composition optionally further comprises an aqueous medium (such as an aqueous buffer or saline) and / or a surfactant (such as a polyethylene glycol fatty ester, such as PEG-15 hydroxystearate). Suitably, the long-acting injectable depot pharmaceutical composition comprises an aqueous suspension. Suitably, the aqueous suspension is a nanosuspension. Suitably, the long-acting injectable depot pharmaceutical composition does not contain a rate control agent, such as a biodegradable polymer, or contains less than 20% or less than 15, 10, 5, 3, 2 or 1% by weight of a rate control agent of the total composition. Suitably, the compound 1 form is Form C.

[0192] In one embodiment, the present invention provides a long-acting injectable depot pharmaceutical composition comprising:

[0193] a) Compound 1 free form (suitably anhydrous Form A or Form C); and

[0194] b) one or more pharmaceutically acceptable excipients;

[0195] Wherein the long-acting injectable reservoir pharmaceutical composition comprises 300 to 900 mg of compound 1, the long-acting injectable reservoir pharmaceutical composition is administered with an injection volume of about 0.1 to about 2 mL, and wherein the long-acting injectable reservoir pharmaceutical composition comprises an aqueous suspension, preferably an aqueous nanosuspension. Suitably, the compound 1 form is Form C.

[0196] In one embodiment, the present invention provides a long-acting injectable depot pharmaceutical composition comprising:

[0197] a) Compound 1 free form as anhydrous Form C; and

[0198] b) one or more pharmaceutically acceptable excipients;

[0199] Wherein the long-acting injectable reservoir-type pharmaceutical composition:

[0200] i. containing 300 to 900 mg of compound 1;

[0201] ii. comprising a water-based suspension;

[0202] iii. having an injection volume of about 0.1 to about 2 mL; and

[0203] iv. sustained release of Compound 1 for at least 6 months after subcutaneous administration at a rate that provides therapeutic plasma concentrations of Compound 1.

[0204] In one embodiment, the present invention provides a long-acting injectable depot pharmaceutical composition comprising:

[0205] a) Compound 1 free form as anhydrous Form C; and

[0206] b) one or more pharmaceutically acceptable excipients;

[0207] Wherein the long-acting injectable reservoir-type pharmaceutical composition:

[0208] i. containing 300 to 900 mg of compound 1;

[0209] ii. comprising a water-based suspension;

[0210] iii. having an injection volume of about 0.1 to about 2 mL; and

[0211] iv. sustained release of Compound 1 for at least 6 months after intramuscular administration at a rate that provides therapeutic plasma concentrations of Compound 1.

[0212] Suitably, the above-mentioned long-acting injectable reservoir pharmaceutical composition comprising compound 1 in an aqueous suspension may further comprise one or more isotonic agents (such as sodium chloride or mannitol), suspending agents (such as polyvinyl pyrrolidone or polysorbate, such as polysorbate 80) and surfactants (such as poloxamer, such as poloxamer 188). Suitably, the aqueous suspension is a micron suspension with a D10 of 0.3 to 2 μm, a D50 of 2 to 4 μm, and / or a D90 of 4 to 13 μm. Suitably, the aqueous suspension is a nanosuspension with a D10 of 1 to 100 nm, a D50 of 1 to 150 nm, and / or a D90 of 1 to 200 nm.

[0213] In one embodiment, the present invention provides a long-acting injectable depot pharmaceutical composition comprising:

[0214] a) Compound 1; and

[0215] b) one or more pharmaceutically acceptable excipients;

[0216] Wherein the long-acting injectable reservoir-type pharmaceutical composition:

[0217] i. containing 300 to 900 mg of compound 1;

[0218] ii. a solution comprising compound 1 in NMP;

[0219] iii. having an injection volume of about 0.1 to about 2 mL; and

[0220] iv. sustained release of Compound 1 for at least 6 months after subcutaneous administration at a rate that provides therapeutic plasma concentrations of Compound 1.

[0221] Suitably, the above-mentioned long-acting injectable reservoir pharmaceutical composition comprising compound 1 in NMP solution may also include one or more aqueous media (e.g., aqueous buffer or saline) and surfactants (e.g., polyethylene glycol fatty acid esters, e.g., PEG-15 hydroxystearate). Suitably, the composition comprises compound 1 at a concentration of 100-400 mg / mL and 5-95% v / v NMP.

[0222] Preparation of composition

[0223] In a second aspect, the present invention provides a method of forming a pharmaceutical composition according to the first aspect.

[0224] In one embodiment, the pharmaceutical composition of the present invention is a water-based suspension reservoir. In one embodiment, the method for preparing a water-based suspension reservoir comprises the following steps:

[0225] a) incorporating a helicase-primase inhibitor or a pharmaceutically acceptable salt thereof into a water-based carrier, wherein the water-based carrier optionally comprises a suspending agent (e.g., polysorbate 80 or poloxamer 188) and optionally an isotonic agent (e.g., sodium chloride); and

[0226] b) mixing the helicase-primase inhibitor or a pharmaceutically acceptable salt thereof (eg by stirring and / or vortexing and / or sonication) to provide a homogenous suspension suitable for subcutaneous or intramuscular administration.

[0227] In one embodiment, the pharmaceutical composition of the present invention is a water-based micron suspension reservoir. In one embodiment, the method for preparing a water-based micron suspension reservoir comprises the following steps:

[0228] a) reducing the particle size of the drug substance helicase-initase inhibitor or a pharmaceutically acceptable salt thereof, for example by wet or dry milling, to provide micron-sized particles (e.g. Dv90 < 10 μm);

[0229] b) incorporating micron-sized particles of a helicase-primase inhibitor or a pharmaceutically acceptable salt thereof in an aqueous carrier, wherein the aqueous carrier optionally comprises a suspending agent (e.g., polysorbate 80 or poloxamer 188) and optionally an isotonicity agent (e.g., sodium chloride) and optionally a viscosity modifier (e.g., hydroxypropyl methylcellulose); and

[0230] c) mixing micron-sized particles of a helicase-primase inhibitor or a pharmaceutically acceptable salt thereof in an aqueous carrier (eg by stirring and / or vortexing and / or sonication) to provide a homogenous suspension suitable for subcutaneous or intramuscular administration.

[0231] In one embodiment, the milling performed in step a) is jet milling.

[0232] In one embodiment, the pharmaceutical composition of the present invention is a water-based nanosuspension reservoir. In one embodiment, the method for preparing a water-based nanosuspension reservoir comprises the following steps:

[0233] a) reducing the particle size of the drug substance helicase-initase inhibitor or a pharmaceutically acceptable salt thereof, for example by wet milling, to provide nano-sized particles (e.g. Dv90 < 500 nm);

[0234] b) incorporating nanometer-sized particles of a helicase-primase inhibitor or a pharmaceutically acceptable salt thereof in an aqueous carrier, wherein the aqueous carrier optionally comprises a suspending agent (e.g., polysorbate 80 or poloxamer 188) and optionally an isotonic agent (e.g., mannitol) and optionally an isotonic agent (e.g., sodium carboxymethylcellulose); and

[0235] c) mixing nano-sized particles of a helicase-primase inhibitor or a pharmaceutically acceptable salt thereof in an aqueous carrier (eg by stirring and / or vortexing and / or sonication) to provide a homogenous nanosuspension suitable for subcutaneous or intramuscular administration.

[0236] In one embodiment, the grinding performed in step a) is wet grinding at 700 rpm for at least 300 minutes to reduce the particle size to nano-sized particles, for example Dv90 < 500 nm.

[0237] In one embodiment, reducing the particle size of the drug substance helicase-initase inhibitor or a pharmaceutically acceptable salt thereof to provide nanoscale particles (e.g., Dv90 <500 nm) for nanosuspension is performed in steps. For example, the particles of the drug substance helicase-initase inhibitor or a pharmaceutically acceptable salt thereof are first reduced to micrometer-sized material, for example, by a high pressure homogenizer until the size is about 1 μm. The method then includes the step of grinding the material, for example, wet grinding using zirconium oxide grinding beads, for example, stirring at a speed of 700 rpm for 500 minutes, to reduce the particle size to nanoscale particles (Dv90 <500 nm).

[0238] Therapeutic uses and applications

[0239] The present invention provides a method for treating herpes virus (suitably HSV) infection in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of a helicase-primase inhibitor or a pharmaceutically acceptable salt thereof, wherein the helicase-primase inhibitor or a pharmaceutically acceptable salt thereof is administered by an injectable route of administration. Suitably, the helicase-primase inhibitor or a pharmaceutically acceptable salt thereof is administered by subcutaneous or intramuscular injection.

[0240] The present invention provides a method for treating a herpes virus (suitably HSV) infection in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of a helicase-primase inhibitor or a pharmaceutically acceptable salt thereof, wherein the helicase-primase inhibitor or a pharmaceutically acceptable salt thereof is formulated as a pharmaceutical composition according to the first aspect of the present invention.

[0241] The present invention also provides a pharmaceutical composition according to the first aspect of the present invention, comprising a helicase-primase inhibitor or a pharmaceutically acceptable salt thereof, for use as a medicament.

[0242] The present invention also provides a pharmaceutical composition according to the first aspect of the invention, comprising a helicase-primase inhibitor or a pharmaceutically acceptable salt thereof, for use in treating a herpes virus (suitably HSV) infection in a subject in need thereof.

[0243] The present invention also provides the use of a pharmaceutical composition comprising a helicase-primase inhibitor or a pharmaceutically acceptable salt thereof according to the first aspect of the invention in the preparation of a medicament for treating a herpes virus (suitably HSV) infection in a subject in need thereof.

[0244] In a specific embodiment, the present invention provides a method for inhibiting HSV replication in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of a helicase-primase inhibitor or a pharmaceutically acceptable salt thereof, wherein the helicase-primase inhibitor or a pharmaceutically acceptable salt thereof is formulated as a pharmaceutical composition according to the first aspect of the present invention.

[0245] In one embodiment, the present invention also provides a method for reducing the likelihood or severity of symptoms of HSV infection in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of a helicase-initiase inhibitor or a pharmaceutically acceptable salt thereof, wherein the helicase-initiase inhibitor or a pharmaceutically acceptable salt thereof is formulated as a pharmaceutical composition according to the first aspect of the present invention.

[0246] In one embodiment, the present invention provides a method for inhibiting the development or progression of a disease or disorder caused by or associated with HSV infection in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of a helicase-primase inhibitor or a pharmaceutically acceptable salt thereof, wherein the helicase-primase inhibitor or a pharmaceutically acceptable salt thereof is formulated as a pharmaceutical composition according to the first aspect of the present invention.

[0247] In one embodiment, the present invention provides a method for inhibiting the recurrence or outbreak of HSV symptoms in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of a helicase-primase inhibitor or a pharmaceutically acceptable salt thereof, wherein the helicase-primase inhibitor or a pharmaceutically acceptable salt thereof is formulated as a pharmaceutical composition according to the first aspect of the present invention.

[0248] In one embodiment, the present invention provides a pharmaceutical composition according to the first aspect of the present invention, comprising a helicase-primase inhibitor or a pharmaceutically acceptable salt thereof, for inhibiting the recurrence or outbreak of HSV symptoms in a subject in need thereof.

[0249] In one embodiment, the present invention provides the use of a pharmaceutical composition comprising a helicase-initiase inhibitor or a pharmaceutically acceptable salt thereof according to the first aspect of the present invention in the preparation of a medicament for inhibiting the recurrence or outbreak of HSV symptoms in a subject in need thereof.

[0250] In one embodiment, the present invention provides a method for treating or preventing a disease or disorder caused by or associated with HSV infection in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of a helicase-primase inhibitor or a pharmaceutically acceptable salt thereof, wherein the helicase-primase inhibitor or a pharmaceutically acceptable salt thereof is formulated as a pharmaceutical composition according to the first aspect of the present invention. In a specific embodiment, the disease or disorder caused by or associated with HSV infection is selected from herpes labialis (e.g., oro-labial cold sores or Whitlow's), genital herpes, HSV-related keratitis, HSV-related encephalitis, pneumonia, herpes gladiatorum, primary HSV gingivostomatitis, Moralet's meningitis, and Bell's palsy.

[0251] In certain embodiments, the disease or disorder caused by or associated with HSV infection is selected from herpes labialis (oral herpes or Whitlow's) or genital herpes. In one embodiment, the disease or disorder is recurrent herpes labialis or recurrent genital herpes. An individual with a history of multiple herpes labialis or recurrent genital herpes (e.g., HSV recurring six or more times per year) may be considered to have recurrent HSV.

[0252] In one embodiment, the herpes virus treated is HSV2. In a further embodiment, the herpes virus treated is HSV2 and the subject in need of treatment has recurrent genital herpes due to HSV2.

[0253] In one embodiment, the herpes virus treated is HSV 1. In yet another embodiment, the herpes viruses treated are both HSV-1 and HSV-2.

[0254] In one embodiment, the herpes virus being treated is resistant to a nucleoside antiviral therapy. In one embodiment, the nucleoside antiviral therapy is selected from acyclovir, penciclovir, famciclovir, ganciclovir and valacyclovir.

[0255] In one embodiment, the herpes virus infection treated is resistant to nucleoside antiviral therapy, for example, a mucocutaneous HSV infection resistant to acyclovir. In a further embodiment, the HSV infection treated is a mucocutaneous HSV infection resistant to antiviral therapy using a nucleoside analog (e.g., acyclovir, penciclovir, famciclovir, ganciclovir or valacyclovir).

[0256] In a specific embodiment, a subject in need of the methods disclosed herein is immunocompromised. The subject may be immunocompromised due to conditions including HIV infection, cancer, hematopoietic cell or solid organ transplantation, and long-term use of glucocorticoids or inherited immunodeficiency.

[0257] In specific embodiments, a subject in need of the methods disclosed herein is a neonate or infant.

[0258] In a specific embodiment, the subject is a herpes positive patient.

[0259] In a specific embodiment, a subject in need of the methods disclosed herein suffers from an acyclovir-resistant mucocutaneous HSV infection. The subject may have been diagnosed with the condition based on clinical failure, e.g., no improvement after at least 7 days of oral or intravenous administration of approved doses of acyclovir.

[0260] In a specific embodiment, the subject in need of the methods disclosed herein suffers from a primary genital HSV-related herpes infection. In one embodiment, the subject in need of the methods disclosed herein suffers from a severe or progressive genital HSV-related herpes infection.

[0261] In a specific embodiment, the pharmaceutical composition according to the first aspect of the invention can reduce the recurrence of HSV infection causing a disease or disorder (e.g., herpes labialis or genital herpes) (i.e., provide suppressive therapy). In one embodiment, the reduction in the number of lesion recurrences within one year can be reduced by 20%, 30%, 40%, 50%, 75%, 90% or 95% or more. Suitably, the incidence of lesions within one year can be reduced by 90% or more.

[0262] In a particular embodiment, the pharmaceutical composition according to the first aspect of the invention can reduce the duration of a recurrent episode of HSV infection, for example by one or more days, for example at least 2, 3, 4, 5, 14, 21 or 28 days.

[0263] In a specific embodiment, the pharmaceutical composition according to the first aspect of the invention can reduce the lesion healing time (e.g., the time when the lesion is completely healed) and the duration of symptoms caused by HSV infection in a disease or disorder (e.g., herpes labialis or genital herpes). The lesion healing time can be defined as the complete epithelialization of the mucocutaneous HSV lesions during treatment and the absence of new lesions, for example, as assessed by a physician.

[0264] In one embodiment, the pharmaceutical composition according to the first aspect of the invention can reduce pain or pain intensity (eg, at the site of the lesion) caused by HSV infection in a disease or disorder (eg, herpes labialis or genital herpes).

[0265] In a specific embodiment, the pharmaceutical composition according to the first aspect of the present invention can reduce viral shedding or reduce the viral shedding rate of individuals with frequent recurrence of HSV (e.g., genital HSV2). For example, the genital HSV mucocutaneous shedding rate in a subject can be measured by collecting skin and mucosal swabs and performing HSV testing (e.g., by using real-time quantitative fluorescent polymerase chain reaction (PCR) to measure the HSVDNA in the analyzed sample). The frequency of HSV2 detection (viral shedding rate) can be defined as the number of days on which the genital swab is HSV positive divided by the total number of days on which the genital swab is obtained. The reduction in the shedding rate of HSV in subjects receiving the composition of the present invention can be compared relative to the shedding rate of subjects receiving a placebo or other treatment. The number of HSV in positive swabs and the frequency of genital lesions and shedding events can also be monitored.

[0266] In a specific embodiment, the present invention provides a method for reducing (or substantially inhibiting or eliminating) breakthrough HSV shedding in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of a helicase-primase inhibitor or a pharmaceutically acceptable salt thereof, wherein the helicase-primase inhibitor or a pharmaceutically acceptable salt thereof is formulated as a pharmaceutical composition according to the first aspect of the present invention.

[0267] In one embodiment, the present invention provides a pharmaceutical composition according to the first aspect of the present invention, comprising a helicase-initiase inhibitor or a pharmaceutically acceptable salt thereof, for reducing (or substantially inhibiting or eliminating) breakthrough HSV shedding in a subject in need thereof.

[0268] In one embodiment, the present invention provides the use of a pharmaceutical composition according to the first aspect of the present invention, wherein the pharmaceutical composition comprises a helicase-initiase inhibitor or a pharmaceutically acceptable salt thereof, for the manufacture of a medicament for reducing (or substantially inhibiting or eliminating) breakthrough HSV shedding in a subject in need thereof.

[0269] In a specific embodiment, the present invention provides a method for preventing the spread of an infectious disease caused by HSV, the method comprising administering to a subject a therapeutically effective amount of a helicase-primase inhibitor or a pharmaceutically acceptable salt thereof, wherein the helicase-primase inhibitor or a pharmaceutically acceptable salt thereof is formulated as a pharmaceutical composition according to the first aspect of the present invention.

[0270] In a specific embodiment, the present invention provides a method for reducing the side effects observed when a helicase-primase inhibitor or a pharmaceutically acceptable salt thereof is administered orally to a subject suffering from an infectious disease caused by HSV, the method comprising administering to the subject a therapeutically effective amount of a helicase-primase inhibitor or a pharmaceutically acceptable salt thereof, wherein the helicase-primase inhibitor or a pharmaceutically acceptable salt thereof is formulated as a pharmaceutical composition according to the first aspect of the present invention.

[0271] The pharmaceutical composition of the present invention is an injectable depot composition and can therefore be administered intramuscularly or subcutaneously. In a convenient embodiment, the pharmaceutical composition of the present invention is administered subcutaneously. In a convenient embodiment, the pharmaceutical composition of the present invention is administered by intramuscular administration.

[0272] In terms of use according to this aspect, the appropriate dosage is expected to vary according to, for example, the nature and severity of the infection to be treated, and is within the purview of the treating physician. Typically, the indicated dosage may be in the range of about 0.1 to about 1000 μg / kg body weight. In some cases, the dosage of the compound may be less than 400 μg / kg body weight. In other cases, the dosage may be less than 200 μg / kg body weight. In some other cases, the dosage may be in the range of about 0.1 to about 100 μg / kg body weight. In one embodiment, the therapeutically effective amount of the helicase-priming enzyme inhibitor or a pharmaceutically acceptable salt thereof is about 5 mg to about 900 mg, for example, about 100 mg to about 600 mg.

[0273] Advantageously, Applicants have discovered that the extended release provided by the compositions of the present invention and the resulting stable plasma levels which remain at therapeutically effective plasma concentrations for extended periods of time following administration allow for the possibility of using lower than desired doses.

[0274] The dosage may conveniently be administered twice a month or less, once a month or less, once every two, three, four, five or six months or less. Suitably, the dosage may conveniently be administered once a month, once every two months or once every three months.

[0275] In a suitable embodiment, the unit dosage form of the pharmaceutical composition is a subcutaneous injection. In a suitable embodiment, the unit dosage form of the pharmaceutical composition comprises about 5 mg to about 900 mg (e.g., about 100 mg to about 900 mg) of a helicase-primase inhibitor or a pharmaceutically acceptable salt thereof. In the most suitable embodiment, the unit dosage form of the pharmaceutical composition is a subcutaneous injection, comprising about 100 mg to about 600 mg of a helicase-primase inhibitor, or a pharmaceutically acceptable salt thereof.

[0276] In one suitable embodiment, the unit dosage form of the pharmaceutical composition comprises about 100 mg to about 1200 mg (such as about 100 mg to about 900 mg) of compound 1, or a pharmaceutically acceptable salt thereof. In a most suitable embodiment, the unit dosage form of the pharmaceutical composition is a subcutaneous injection, comprising about 100 mg to about 900 mg of compound 1, or a pharmaceutically acceptable salt thereof. In another suitable embodiment, the unit dosage form of the pharmaceutical composition is a subcutaneous injection, comprising about 200 mg to about 900 mg, such as about 200 mg to about 600 mg, about 300 mg to about 900 mg, or about 300 mg to about 600 mg of compound 1, or a pharmaceutically acceptable salt thereof.

[0277] In one embodiment, the present invention provides a method for treating a herpes virus (suitably HSV) infection in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of a helicase-primase inhibitor or a pharmaceutically acceptable salt thereof, wherein the helicase-primase inhibitor or a pharmaceutically acceptable salt thereof is formulated as a pharmaceutical composition according to the first aspect of the present invention (e.g., certain long-acting injectable depot pharmaceutical compositions, including solutions in organic solvents or aqueous microsuspensions or nanosuspensions), wherein no loading dose is required prior to administration of the long-acting injectable depot according to the first aspect of the present invention, such as an oral loading dose.

[0278] combination

[0279] The pharmaceutical composition of the present invention may be administered alone as a monotherapy, or may be administered together with one or more other substances and / or treatments. Such combined treatment may be achieved by simultaneous, sequential or separate administration of the individual components of the treatment.

[0280] Also contemplated herein are methods comprising administering a second active agent. For example, in addition to being infected with HSV, the subject or patient may also suffer from complications associated with HSV infection, i.e., diseases and other adverse health conditions associated with, exacerbated or aggravated by HSV infection. Also contemplated herein are combinations of the disclosed pharmaceutical compositions with at least one other agent that has previously been shown to treat these HSV infection-related conditions. This combination therapy can be achieved independently (by administering the individual components of the treatment simultaneously, sequentially or separately) and / or by a pharmaceutical composition of the invention comprising a second active agent.

[0281] Therefore, the present invention provides a method for treating or preventing HSV infection in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a helicase-primase inhibitor or a pharmaceutically acceptable salt thereof, wherein the helicase-primase inhibitor or a pharmaceutically acceptable salt thereof is formulated as a pharmaceutical composition according to the first aspect of the present invention, and co-administering to the subject a therapeutically effective amount of another therapeutic agent.

[0282] In an embodiment, the additional therapeutic agent is selected from one or more of the following agents:

[0283] i. Nucleoside polymerase inhibitors, such as acyclovir, valacyclovir, famciclovir, penciclovir and ganciclovir;

[0284] ii. Pyrophosphate polymerase inhibitors, such as foscarnet;

[0285] iii. saturated fatty alcohols, such as behenyl alcohol;

[0286] iv. Drugs such as iodoxuridine, trifluridine and adenosine;

[0287] v. Corticosteroids; and

[0288] vi. Other helicase-primase inhibitors, such as amenamevir.

[0289] In some cases, the pharmaceutical composition disclosed according to the first aspect of the present invention can be administered as part of a combination therapy with one or more antiviral drugs, including nucleoside analogs, such as acyclovir, foscarnet, ganciclovir or penciclovir or the corresponding prodrugs valacyclovir or famciclovir.

[0290] In some embodiments, the first and second amounts together constitute a pharmaceutically effective amount. The first amount, the second amount, or both may be the same, greater than, or less than the effective amount of each compound administered as a monotherapy. A therapeutically effective amount of a disclosed compound and an antiviral drug may be co-administered to a subject, that is, administered to a subject simultaneously or separately in any given order and by the same or different routes of administration. In some cases, it may be advantageous to begin administering Compound 1 first, for example one or more days or weeks before beginning administration of an antiviral drug. In addition, other drugs may be used in conjunction with the above-described combination therapy.

[0291] Reagent test kit

[0292] In one embodiment, the pharmaceutical compositions and methods described herein provide kits for treating diseases, such as kits described herein. These kits include pharmaceutical compositions described herein in containers, and optional instructions, which teach the use of the kits according to various methods and approaches described herein. Such kits may also include information, such as scientific literature references, package insert materials, clinical trial results, and / or summaries of these contents, etc., which indicate or determine the activity and / or advantages of the composition, and / or describe dosage, administration, side effects, drug interactions, or other information useful to health care providers. Such information may be based on the results of various studies, such as studies using experimental animals involving in vivo models and studies based on human clinical trials. Kits described herein may be provided to health care providers, including doctors, nurses, pharmacists, prescription officials, etc., and marketed and / or promoted to them. In some embodiments, the kit may also be marketed directly to consumers.

[0293] The pharmaceutical compositions of the present invention are useful in diagnosis and as research tools.

[0294] In addition to being useful for human treatment, the pharmaceutical compositions of the present invention may also be used for veterinary treatment of companion animals, exotic animals, and farm animals, including mammals, rodents, etc. Conveniently, such animals include horses, dogs, and cats.

[0295] The present invention is now illustrated by the following non-limiting examples. Example

[0296] The following abbreviations are used in this manual:

[0297] ACN: Acetonitrile

[0298] DCM: dichloromethane

[0299] DMF: Dimethylformamide

[0300] DMSO: dimethyl sulfoxide

[0301] DSC: Differential Scanning Calorimetry

[0302] DVS: Dynamic Vapor Sorption

[0303] EDCI: 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide

[0304] eq: equivalent

[0305] EtOAc: Ethyl acetate

[0306] hr:hour

[0307] HPLC: High Performance Liquid Chromatography

[0308] K2CO3: Potassium carbonate

[0309] K3PO4: Tripotassium phosphate

[0310] KF: Karl Fischer titration

[0311] MeOH: Methanol

[0312] min:minutes

[0313] NaCMC: Sodium carboxymethyl cellulose

[0314] Na2SO4: Sodium sulfate

[0315] NMP: N-Methylpyrrolidone

[0316] NMT: no more than

[0317] PLM: Polarized Light Microscopy

[0318] PVP K30:Povidone K30

[0319] RH:Relative humidity

[0320] RT: Room temperature (~22°C)

[0321] TBAB:Tetrabutylammonium bromide

[0322] TGA: Thermogravimetric analysis

[0323] THF:Tetrahydrofuran

[0324] TLC: Thin layer chromatography

[0325] Vol: Volume

[0326] XRPD: X-ray powder diffraction

[0327] Materials and methods

[0328] Particle size distribution by light diffraction

[0329] The particle size distribution of the suspensions prepared in Examples 1 and 10 was determined by laser diffraction using a NANOTRAC FLEX particle size analyzer (Microtrac Instruments). Water was used as the medium. Samples were added to maintain the laser obscuration level at 0.1-10%. A measurement time of 20 seconds was used, and the measurement was performed with water as the background medium, with a collection time of 10 seconds. Determination of Dv 10 、Dv 50 and Dv 90 diameter and reported below.

[0330] The particle size distribution of the suspensions prepared in Examples 14 and 17 was determined using a Mastersizer 3000 particle size analyzer (Malvern, UK). The suspensions were freeze dried and the freeze dried powder was used to prepare a 0.1% suspension in water for testing. Water was used as a dispersant. The suspension was added until the final coverage range was 3-10%. Each sample was measured 3 times. Dv was determined 10 、Dv 50 and Dv 90 diameter and reported below.

[0331] Scanning electron microscopy

[0332] Scanning electron microscope (SEM) images of the suspensions prepared in Examples 14 and 17 below were taken using an Oxford X-MAS scanning electron microscope (Zeiss-Sigma-HD, Germany). Before measurement, the suspensions were diluted with distilled water (4X) and sonicated for about 5 minutes. The samples were then dipped in a conductive adhesive and air-dried at room temperature. Loose powder was removed using a rubber suction bulb. The samples were then sprayed with gold for testing.

[0333] LC-MS analysis of plasma samples

[0334] Table 1 provides the LC-MS method and conditions used to analyze priligy in plasma samples in the PK studies described in Examples 1-5.

[0335] Table 1. LC-MS parameters used for quantification of priligy in plasma samples

[0336]

[0337] Table 2 provides the LC-MS method and conditions used to analyze priligy in dog plasma samples from the PK study described in Example 14.

[0338] Table 2. LC-MS parameters used for quantification of priligy in dog plasma samples

[0339]

[0340] The LC-MS method and conditions used to analyze the helicase-initase inhibitor Compound 1 in plasma samples from subcutaneous PK studies (Examples 7 and 10-12) are described in Tables 3 and 4.

[0341] Table 3. LC-MS parameters used for quantification of compound 1 in rat plasma samples

[0342]

[0343]

[0344] Table 4. LC-MS parameters used for quantification of compound 1 in dog and monkey plasma samples

[0345]

[0346]

[0347] The LC-MS method and conditions used to analyze Compound 1 in plasma samples from the intravenous injection PK study (Example 13) are described in Tables 5 to 7.

[0348] Table 5. LC-MS parameters for quantification of Compound 1 in rat plasma samples after intravenous administration in the study described in Example 13

[0349]

[0350] Table 6. LC-MS parameters for quantification of Compound 1 in dog plasma samples after intravenous administration in the study described in Example 13

[0351]

[0352]

[0353] Table 7. LC-MS parameters for quantification of Compound 1 in monkey plasma samples after intravenous administration in the study described in Example 13

[0354]

[0355]

[0356] Table 8 provides the LC-MS method and conditions used to analyze the helicase-primase inhibitor Compound 1 in rat plasma samples from the subcutaneous PK study described in Example 15.

[0357] Table 8. LC-MS parameters used for quantification of compound 1 in rat plasma samples

[0358]

[0359] Table 9 provides the LC-MS method and conditions used to analyze the helicase-primase inhibitor Compound 1 in dog plasma samples from the dog PK study described in Example 16.

[0360] Table 9. LC-MS parameters used for quantification of compound 1 in dog plasma samples

[0361]

[0362]

[0363] Table 10 gives the LC-MS method and conditions used to analyze Compound 1 in rat plasma samples from the PK study described in Example 17.

[0364] Table 10. LC-MS parameters used for quantification of compound 1 in rat plasma samples

[0365]

[0366]

[0367] X-ray powder diffraction (XRPD) analysis

[0368] Instrument: Rigaku Smartlab SE

[0369] X-ray wavelength: Cu, Kα, 1.540598, 1.544426

[0370] Kα2:Kα1 intensity ratio: 0.50

[0371] X-ray tube settings: 40kV, 15mA, scanning mode 1D

[0372] Scanning range (2θ): 3°-40°, step size (2θ): 0.02 o , scanning speed (2θ) 10o / min

[0373] Priligy free base (dihydrate)

[0374] Priligy (100 mg) was dissolved in 88:12 THF / water (1 mL) at ~60°C. The solution was cooled and water (4 mL) was added. The resulting suspension was filtered and the solids were collected and dried.

[0375] The crystalline Privir free base obtained according to the present method was analyzed as described below and used in Examples 1 to 6 described herein.

[0376] Privir free base was analyzed by TGA and XRPD. By TGA, the material lost 7.9% by weight of water between 30°C and 90°C, corresponding to the dihydrate. The XRPD diffraction pattern is shown in Figure 1 The peak positions are listed in the table below. Characteristic peaks include peaks at 11.0, 11.3 and 21.1° 2θ.

[0377]

[0378]

[0379] Priligy free base (sesquihydrate)

[0380] 2-(4-(Pyridin-2-yl)phenyl)acetic acid (38.40 g, 80%, 180.074 mmol, 1.00 eq) was added to DMF (385 mL). HOBT (17.38 g, 128.624 mmol, 0.71 eq) was added at 20 ° C and stirred for 10 minutes. 2-Chloro-4-methylthiazole-5-sulfonamide (26.66 g, 128.624 mmol, 0.71 eq) and EDCI (27.12 g, 141.486 mmol, 0.79 eq) were added and the mixture was stirred at 20 ° C for 36 hours under N2. The mixture was added to water (780 mL) and stirred for 30 minutes. The solid was collected by filtration. The filter cake was washed twice with water (2 x 100 mL) and then dried under reduced pressure at 50 ° C for 12 hours to obtain priligy as a white solid (yield 78.4%).

[0381] The crystalline priligy free base obtained according to this procedure was analyzed as described below and used in Example 14.

[0382] The free base of Priligy was analyzed by TGA and XRPD. By TGA, the material lost 5.6% of its weight between 35 and 101°C. Characterization data indicated that this was the hemihydrate of Priligy (1.5 water molecules per 1 molecule of Priligy). The XRPD diffraction pattern is shown in Fig.16 The peak positions are listed in the table below. Characteristic peaks include peaks at 12.1, 14.6 and 18.3° 2θ.

[0383] Peak position °2θ Peak position °2θ Peak position °2θ Peak position °2θ Peak position °2θ Peak position °2θ 7.2 18.3 22.6 26.9 31.9 36.3 8.7 19.9 23.7 27.6 32.3 36.5 10.3 20.4 23.9 28.0 33.2 36.9 12.1 20.7 24.1 28.7 33.8 37.8 14.6 20.9 24.5 29.4 34.4 38.8 15.0 21.5 24.8 29.6 34.9 39.5 15.4 22.1 25.6 30.5 35.4 39.8 16.3 22.4 26.1 30.8 35.8

[0384] Compound 1 free base (anhydrous form A)

[0385] The crude compound 1 (36.5 g) prepared according to Example 8 was stirred in DMSO (300 mL, 8.2 relative volumes) at 15-27° C. for 20 to 30 minutes to form a solution. Water (1100 mL, 30.1 relative volumes) was slowly added over 30 minutes, and the mixture was stirred for 1-2 hours at 15-27° C. Additional water (1820 mL, 49.9 relative volumes) was slowly added over 60 minutes, and the mixture was stirred for 12 hours at 15-27° C. The mixture was then filtered, washed with water (3 x 365 mL, 3 x 10 volumes), and the filter cake was dried at 45 to 55° C. to give 26.4 g (72% yield) of compound 1 as an off-white solid.

[0386] The crystalline Compound 1 free base Form A obtained according to this procedure was analyzed as described below and used in Examples 7 and 10-12.

[0387] The crystalline form obtained has Fig.17 The XRPD diffraction pattern shown is designated as Form A. The following table lists the peak positions present in the XRPD diffraction pattern obtained for Form A.

[0388]

[0389] Form A was further solid-state characterized using DSC and TGA and was determined to be an anhydrous crystalline form of Compound 1.

[0390] Compound 1 free base (anhydrous form C)

[0391] The crude compound 1 (4.0 kg) prepared according to Example 8 was stirred in DMSO (12 L, 3 relative volumes) at 75-85 ° C in the first reactor to form a solution. The solution was then filtered through a 0.2 μm filter into a second preheated (75 to 85 ° C) reactor, followed by rinsing the first reactor with DMSO (2 L, 0.5 relative volumes) and transferring the rinse to the second reactor. The contents of the second reactor were stirred at 75-85 ° C for 30-60 minutes and then cooled to a batch temperature of 50-55 ° C at a cooling rate of 8-12 ° C per hour. The mixture was stirred at 50-55 ° C for 1-2 hours. Ethanol (4 L, 1 relative volume) was then slowly added over 60-90 minutes while the batch temperature was maintained at 50-55 ° C. The internal batch temperature was raised to 57 to 62 ° C, and ethanol (52 L, 13 relative volumes) was slowly added over 7.0-7.5 hours. The contents were stirred at 57 to 62° C. for 1 to 2 hours and then cooled at a cooling rate of 8-12° C. per hour to bring the final batch temperature to 18-22° C. The mixture was stirred at 18-22° C. for 8-12 hours and then filtered, washed with ethanol (8 L, 2 volumes) and the filter cake was dried at 45 to 55° C. for 20-24 hours to give 3.6 kg (90% yield) of compound 1 as an off-white to light yellow solid.

[0392] The crystalline Compound 1 free base obtained according to this procedure was analyzed as described below and used in Example 17.

[0393] The crystalline form obtained has Fig.18 The XRPD diffraction pattern shown and designated as Form C. The peak positions in the XRPD diffraction pattern obtained for Form C are shown in the following table:

[0394]

[0395] Further solid-state characterization of Form C was performed using DSC, TGA, and DVS. Form C was determined to be an anhydrous crystalline form of Compound 1.

[0396] Antiviral test

[0397] The in vitro EC values ​​of helicase primase inhibitors for HSV-1 and HSV-2 can be determined using the following assay: 50 .

[0398] Cytopathic effect (CPE) inhibition assay

[0399] Cell culture - Vero cells were cultured in Dulbecco's Modified Eagle's Medium (DMEM) supplemented with 10% fetal bovine serum and 100 units / mL penicillin and streptomycin. Cells were passaged 2-3 times per week to maintain subconfluent density.

[0400] HSV-1 Antiviral Test -Vero cells were plated at 2.5×10 per well. 3 Cells were seeded into 96-well plates at a density of 10 cells / mL and allowed to attach overnight. After attachment, the medium was replaced with 50 μL infection medium (DMEM supplemented with 2% fetal bovine serum and 100 units / mL penicillin and streptomycin). Compounds were then added to the cultures in an 8-point 3-fold serial dilution format using a Tecan D300e digital dispenser. DMSO concentrations were normalized to 0.5% for all treatments. After compound addition, 50 μL containing 80 TCID 50 Infection medium for HSV-1 was added to the cells and incubated at 37°C for 4 days. After incubation, the plates were equilibrated to room temperature, the medium was removed, and 60 μL of a 1:1 dilution of Cell titer glow and phosphate buffered saline was added to the cells. After a 5-minute incubation, the brightness was measured using a Tecan Infinite M1000Pro plate reader to quantify cell viability.

[0401] HSV-2 Antiviral Test -Vero cells were plated at 1.0×10 per well. 4 Cells were seeded into 96-well plates at a density of 100 cells / mL and allowed to attach overnight. After attachment, the medium was replaced with 50 μL of infection medium (DMEM supplemented with 2% fetal bovine serum and 100 units / mL of penicillin and streptomycin). Compounds were then added to the cultures in an 8-point 3-fold serial dilution format using a Tecan D300e digital dispenser. DMSO concentrations were normalized to 0.5% for all treatments. After compound addition, 50 μL of 160 TCID 50 Infection medium of HSV-2G strain was added to the cells and incubated at 37°C for 5 days. After incubation, 10 μL / well WST-8 colorimetric reagent was added and the plate was incubated at 37°C for 3 hours. After incubation, cell viability was quantified by measuring absorbance at 460 nm and 620 nm using a Tecan Infinite M1000 Pro plate reader.

[0402] The mean EC values ​​of priligy for HSV-1 and HSV-2 using this CPE assay 50 The values ​​were 0.047 μM and 0.055 μM, respectively.

[0403] Example 1: Priligy PK Study in Rats

[0404] Preparation

[0405] Aqueous suspensions of crystalline priligy free base (dihydrate) in 0.9% NaCl + 0.5% polysorbate 80 vehicle at concentrations of 10.0 mg / mL and 50.0 mg / mL were prepared as follows:

[0406] 1) A vehicle (0.9% w / w NaCl and 0.5% w / w polysorbate 80) was prepared by dissolving NaCl and polysorbate 80 in water.

[0407] 2) Weigh an appropriate amount of crystalline priligy free base (dihydrate) and add to the carrier.

[0408] 3) The mixture was vortexed for 1 minute, stirred for 10 minutes, and then sonicated for 15 minutes.

[0409] 4) The final dosage form is a uniform white turbid suspension.

[0410] Formulation Characterization

[0411] The particle size distribution (PSD) of the suspension was determined to be Dv 10 =1.45μm, Dv 50 =4.23μm and Dv 90 =10.35μm.

[0412] Animal dosing

[0413] Male Sprague Dawley (SD) rats (6-8 weeks old) weighing about 300 g were used for pharmacokinetic studies. Animals were free to eat and drink. The above-mentioned priligy suspension was administered to each animal group (n=3) by subcutaneous injection, and the administration volume was 3.33 mL / kg to achieve a dose of 33.33 mg / kg and 166.67 mg / kg.

[0414] At each time point, approximately 200 μL of whole blood was collected via the jugular vein into a K2EDTA tube. The blood samples were placed on ice and centrifuged at 2000 g for 5 minutes, and plasma samples were obtained within 15 minutes. The plasma samples were stored at approximately -70°C and then analyzed by LC-MS.

[0415] result

[0416] Figure 2 Mean plasma concentration profiles following subcutaneous (SC) administration of Priligy suspension at two different dose levels (33.33 mg / kg and 166.67 mg / kg) are shown (n=3). Prolonged exposure of at least 720 hours (study duration) was achieved at both dose levels.

[0417] Example 2: Lower dose PK study of priligy in rats

[0418] Preparation

[0419] An aqueous suspension of crystalline priligy free base (dihydrate) in 0.9% NaCl + 0.5% polysorbate 80 vehicle at a concentration of 3.0 mg / mL was prepared as follows:

[0420] 1) The vehicle (0.9% w / w NaCl and 0.5% w / w polysorbate 80) was prepared by dissolving NaCl and polysorbate 80 in water.

[0421] 2) Weigh an appropriate amount of crystalline priligy free base (dihydrate) and add to the carrier.

[0422] 3) The mixture was stirred for 15 minutes, then vortexed for 1 minute, and then sonicated for 10 minutes.

[0423] 4) The final dosage form is a uniform white turbid suspension.

[0424] Animal dosing

[0425] The rat PK study used male SD rats weighing 230-250 g and aged 6-8 weeks. The animals were allowed to eat and drink freely. The prepared suspension was subcutaneously injected into each group of animals (n=3) at a volume of 3.33 mL / kg to achieve a dose of 10 mg / kg.

[0426] At each time point, approximately 200 μL of whole blood was collected via the jugular vein into a K2EDTA tube. The blood samples were placed on ice and centrifuged at 2000 g for 5 minutes, and plasma samples were obtained within 15 minutes. The plasma samples were stored at approximately -70°C until analysis by LC-MS.

[0427] result

[0428] Figure 3 Mean plasma concentration profiles following subcutaneous (SC) administration of priligy suspension are shown (n=3). Prolonged exposure of at least 144 hours (study duration) was achieved at the lower dose level of 10 mg / kg.

[0429] Example 3: Priligy PK Study in Monkeys

[0430] Preparation

[0431] An aqueous suspension of crystalline priligy free base (dihydrate) in 0.9% NaCl + 0.5% polysorbate 80 vehicle at a concentration of 3.0 mg / mL was prepared as follows:

[0432] 1) The vehicle (0.9% w / w NaCl and 0.5% w / w polysorbate 80) was prepared by dissolving NaCl and polysorbate 80 in water.

[0433] 2) Weigh an appropriate amount of crystalline priligy free base (dihydrate) and add to the carrier.

[0434] 3) The mixture was stirred for 15 minutes, then vortexed for 1 minute, and then sonicated for 10 minutes.

[0435] 4) The final dosage form is a uniform white turbid suspension.

[0436] Animal dosing

[0437] The monkey PK study was conducted using male non-naive cynomolgus monkeys weighing ~5 kg. The animals had free access to food and water. The prepared suspension was injected subcutaneously to each group of animals (n=3) at a volume of 3.33 mL / kg to achieve a dose of 10 mg / kg.

[0438] At each time point, approximately 0.5 mL of whole blood was collected into K2EDTA tubes via the cephalic vein. Blood samples were placed on ice and centrifuged at 2000 g for 5 minutes, and plasma samples were obtained within 15 minutes. Plasma samples were stored at approximately -70°C until analysis by LC-MS.

[0439] result

[0440] Figure 4 Mean plasma concentration profiles following subcutaneous (SC) administration of priligy suspension are shown (n=3). Prolonged exposure of at least 456 hours (study duration) was achieved at the 10 mg / kg administration dose level.

[0441] Example 4: Priligy PK Study in Dogs

[0442] Preparation

[0443] An aqueous suspension of crystalline priligy free base (dihydrate) in 0.9% NaCl + 0.5% poloxamer 188 vehicle at a concentration of 5.0 mg / mL was prepared as follows:

[0444] 1) The vehicle (0.9% w / w NaCl and 0.5% w / w Poloxamer 188) was prepared by dissolving NaCl and Poloxamer 188 in water.

[0445] 2) Weigh an appropriate amount of crystalline priligy free base (dihydrate) and add to the carrier.

[0446] 3) The mixture was stirred for 15 minutes, then vortexed for 1 minute, and then sonicated for 10 minutes.

[0447] 4) The final dosage form is a uniform white turbid suspension.

[0448] Animal dosing

[0449] Male non-naive beagles were used for the PK study in dogs. The animals had free access to food and water. The prepared suspension was injected subcutaneously to each group of animals (n=3) at a volume of 2 mL / kg to achieve a dose of 10 mg / kg.

[0450] At each time point, approximately 0.5 mL of whole blood was collected into K2EDTA tubes. Blood samples were placed on ice and centrifuged at 2000 g for 5 minutes, and plasma samples were obtained within 15 minutes. Plasma samples were stored at approximately -70°C until analysis by LC-MS.

[0451] result

[0452] Figure 5 Mean plasma concentration profiles following subcutaneous (SC) administration of priligy suspension are shown (n=3).After an initial drug burst, prolonged exposure was achieved at the administered dose level.

[0453] PK Study Summary

[0454] In three different species, an injectable depot formulation containing priligy was found to provide sustained and stable drug plasma levels for significantly prolonged periods of time after a single subcutaneous administration. In addition, extended release was achieved at different dose levels.

[0455] Furthermore, no adverse reactions were observed at the injection site in the study animals.

[0456] Example 5: Comparative PK study of priligy in rats after IV / SC administration

[0457] Preparation

[0458] A solution of crystalline priligyr free base (dihydrate) in 5% NMP, 5% Solutol HS15 and 90% saline was prepared at a concentration of 0.5 mg / mL as follows:

[0459] 1) Prepare a 10 mg / mL stock solution of Priligy (dihydrate) in NMP.

[0460] 2) Combine equal volumes of Priligy stock solution and Solutol HS15 and mix.

[0461] 3) Dilute the above solution with normal saline to achieve a target 0.5 mg / mL priligy concentration. The preparation is a transparent solution.

[0462] Animal dosing

[0463] Male Sprague Dawley (SD) rats (age 6-8 weeks) weighing 300 g were used for pharmacokinetic studies. Animals were allowed to eat and drink freely. The above-mentioned priligy solution was administered to animals (n=3) by intravenous injection at a dose of 2 mL / kg to achieve a dose of 1 mg / kg. For subcutaneous injection, the dose was 10 mL / kg to achieve a dose of 5 mg / kg.

[0464] At each time point, approximately 200 μL of whole blood was collected via the jugular vein into a K2EDTA tube. The blood samples were placed on ice and centrifuged at 2000 g for 5 minutes to obtain plasma samples within 15 minutes. The plasma samples were stored at approximately -70°C until analysis by LC-MS according to the above method.

[0465] result

[0466] Figure 6 Mean plasma concentration curves following intravenous (IV) administration of 1 mg / kg priligy solution and subcutaneous (SC) administration of 5 mg / kg priligy solution are shown (n=3). The calculated F for the SC route was approximately 117% relative to the IV route (Table below).

[0467]

[0468] This rat PK study demonstrated that the SC route of administration provided good exposure to priligy and that SC bioavailability was surprisingly comparable to IV bioavailability.

[0469] Example 6: Study on stability and solubility of Priligy suspension (dihydrate)

[0470] Preparation

[0471] An aqueous suspension of crystalline prilivir free base (dihydrate) in 0.9% NaCl + 0.5% polysorbate 80 vehicle was prepared as follows:

[0472] 1) Prepare the vehicle (0.9% w / w NaCl and 0.5% w / w Polysorbate 80) by dissolving NaCl and Polysorbate 80 in water.

[0473] 2) Weigh an excess of Priligy (dihydrate) and add to the vehicle to obtain a ~3 mg / mL suspension.

[0474] 3) The suspension was sonicated for 15 minutes and then stirred at 500 rpm at room temperature.

[0475] 4) After 24 hours, the suspension was filtered, the filter cake was dried and analyzed by XRPD. The filtrate was analyzed by HPLC to determine solubility.

[0476] HPLC analysis

[0477] The HPLC method and conditions used to quantify Priligy in the filtrate are shown in Table 11.

[0478] Table 11. HPLC parameters for quantification of priligy in filtrate

[0479]

[0480] result

[0481]

[0482] XRPD analysis of the solid remaining after the solubility experiment confirmed that it was the dihydrate (ie, no polymorphic transition occurred during the experiment).

[0483] Example 7: PK study using helicase-primase inhibitor Compound 1

[0484] Another helicase-primase inhibitor (HPI), Compound 1 (described in Example 8), was also subjected to animal PK testing when formulated into a long-acting injectable depot composition according to the present invention.

[0485] Compound 1 was isolated as a crystalline solid with an average in vitro EC of 50 The values ​​were 0.015 μM and 0.013 μM, respectively.

[0486] PK studies in rats

[0487] Preparation

[0488] An aqueous suspension of crystalline Compound 1 (Form A) in a 0.9% NaCl + 0.5% polysorbate 80 vehicle was prepared as described in Example 2 at a concentration of 3.0 mg / mL.

[0489] Then, according to the animal administration method of Example 2, the suspension was administered to male SD rats at 10 mg / kg.

[0490] result

[0491] Figure 7 Shown are mean plasma concentration curves (n=3) following subcutaneous (SC) administration of a 10 mg / kg suspension of Compound 1. When formulated as an aqueous suspension, Compound 1 achieved extended exposure times of at least 144 hours (the duration of the study) at 10 mg / kg.

[0492] PK studies in monkeys

[0493] Preparation

[0494] An aqueous suspension of crystalline Compound 1 (Form A) in a 0.9% NaCl + 0.5% polysorbate 80 vehicle was prepared as described in Example 3 at a concentration of 3.0 mg / mL.

[0495] Then, according to the animal administration method of Example 3, the suspension was administered to male cynomolgus monkeys at 10 mg / kg.

[0496] result

[0497] The mean plasma concentration curve (n=3) after subcutaneous (SC) administration of Compound 1 suspension at 10 mg / kg is shown in Figure 8 When formulated as an aqueous suspension, Compound 1 at 10 mg / kg achieved prolonged exposure of at least 1392 hours (the duration of the study).

[0498] PK studies in dogs

[0499] Preparation

[0500] An aqueous suspension of crystalline Compound 1 (Form A) in a 0.9% NaCl + 0.5% Poloxamer 188 vehicle was prepared as described in Example 4 at a concentration of 5.0 mg / mL.

[0501] Then, according to the animal administration method of Example 4, the suspension was administered to male non-naive beagle dogs at 10 mg / kg.

[0502] result

[0503] The mean plasma concentration curve (n=3) after subcutaneous (SC) administration of Compound 1 suspension at 10 mg / kg is shown in Fig. 9 When formulated as an aqueous suspension, Compound 1 at 10 mg / kg achieved prolonged exposure of at least 1392 hours (the duration of the study).

[0504] Summary of PK studies from Example 7 for Compound 1

[0505] In three different species, an injectable depot formulation containing compound 1 was found to provide sustained and stable drug plasma levels for a significantly prolonged period of time after a single subcutaneous administration. In addition, extended release was achieved at different dose levels. In addition, no adverse reactions were observed at the injection site in the study animals.

[0506] Example 8: Synthesis of 2-(3-(2',5'-difluoro-[1,1'-biphenyl]-4-yl)-2-oxotetrahydropyrimidine-1(2H)- 4-Methylthiazole-5-sulfonamide (Compound 1)

[0507]

[0508] Synthesis of 1-(4-methylthiazol-2-yl)tetrahydropyrimidin-2(1H)-one (1-2)

[0509] A mixture of compound 1-1 (6 g, 52.632 mmol) and 1-chloro-3-isocyanatopropane (6.26 g, 52.632 mmol) in THF (60 mL) was heated at 70 ° C for 6 h. TBAB (1.7 g, 5.263 mmol) and K2CO3 (18.15 g, 131.58 mmol) were added to the resulting solution in batches, the same temperature was maintained and stirring was continued at 70 ° C for 16 h. After the reaction was completed (monitored by TLC), the reaction mixture was diluted with water and extracted with EtOAc. The combined organic layer was dried with anhydrous Na2SO4, filtered and concentrated under reduced pressure. The crude compound was purified by Combiflash chromatography (eluted with 60-70% EtOAc in heptane) to obtain the title compound 1-2 (5.1 g, 49.22%), which was an off-white solid. TLC: 70% EtOAc / heptane (R f :0.5). Chemical formula: C8H 11 MS calculated value for N3OS: 197.06; found value: 198.17 [M+1] + . 1 H NMR (400MHz, DMSO-d6) δ7.30 (s, 1H), 6.60 (s, 1H), 3.99 (t, J = 5.4Hz, 2H), 3.20-3.19 (m, 2H), 2.28 (s, 3H), 1.99-1.89 (m, 2H).

[0510] Synthesis of 1-(2',5'-difluoro-[1,1'-biphenyl]-4-yl)-3-(4-methylthiazol-2-yl)tetrahydropyrimidin-2(1H)-one (1-3)

[0511] Int.1A (8.16 g, 30.456 mmol), K2CO3 (8.75 g, 63.45 mmol) were added to a stirred solution of compound 1-2 (5 g, 25.380 mmol) in 1,4-dioxane (100 mL), and then CuI (0.96 g, 5.076 mmol) was added and the resulting reaction mixture was purged under nitrogen for 20 minutes. 1,2-dimethylethylenediamine (0.9 g, 10.152 mmol) was added to the resulting reaction mixture under a nitrogen atmosphere. The reaction mixture was heated at 120 ° C for 24 h in a closed tube. After the reaction was completed, the reaction mixture was filtered through a diatomaceous earth bed and washed with ethyl acetate. The filtrate was diluted with water, extracted with EtOAc, and then extracted with brine. The combined organic layer was dried with anhydrous Na2SO4, filtered and concentrated under reduced pressure. The crude compound was purified by Combiflash chromatography (eluting with 30-40% EtOAc in heptane) to afford the title compound 1-3 (4.1 g, 41.96%) as an off-white solid. TLC: 50% EtOAc / heptane (R f :0.5). Chemical formula: C 20 H 17 MS calculated value for F2N3OS: 385.11; found value: 385.90 [M+1] + . 1 H NMR (400MHz, DMSO-d6) δ7.61 (d, J=7.8Hz, 2H), 7.54-7.35 (m, 4H), 7.35-7.21 (m, 1H), 6. 70 (s, 1H), 4.17 (t, J=5.6Hz, 2H), 3.81 (t, J=4.9Hz, 2H), 2.26 (s, 3H), 2.24-2.21 (m, 2H).

[0512] Synthesis of 2-(3-(2',5'-difluoro-[1,1'-biphenyl]-4-yl)-2-oxotetrahydropyrimidin-1(2H)-yl)-4-methylthiazole-5-sulfonic acid (1-4)

[0513] To a stirred solution of compound 1-3 (4 g, 10.389 mmol) in anhydrous DCM (40 mL) was added chlorosulfuric acid (2.07 mL, 31.168 mmol) at 0 ° C in an inert atmosphere and the resulting reaction mixture was slowly warmed to room temperature and stirred for 12 h. After the reaction was completed, the reaction mixture was concentrated to dryness under reduced pressure. The resulting crude residue was purified by grinding with ether. The resulting solid was filtered off and dried in vacuo to give the title compound 1-4 (3.35 g, crude material), which was an off-white solid. TLC: 100% EtOAc (R f :0.2). Chemical formula: C 20 H 17MS calculated value for F2N3O4S2: 465.06; found value: 466[M+1] + .

[0514] Synthesis of 2-(3-(2',5'-difluoro-[1,1'-biphenyl]-4-yl)-2-oxotetrahydropyrimidin-1(2H)-yl)-4-methylthiazole-5-sulfonamide (Compound 1)

[0515] The stirring solution of compound 1-4 (3.3 g, 7.096 mmol) in POCl (33 mL) was stirred at 90 ° C for 5 h. The reaction mixture was concentrated to dryness under reduced pressure. The obtained residue was dissolved in THF (66 mL), and ammonia (33 mL) was added at -5 ° C and stirred at room temperature for another 12 h. After the reaction was completed, the reaction mixture was diluted with water and extracted with EtOAc. The combined organic layer was dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure. The crude compound was purified by Combiflash chromatography (eluted with 100% EtOAc) to obtain the desired product compound 1 (1.1 g, 44.64%), which was a white solid. 1 H NMR (400MHz, DMSO-d6) δ7.65-7.59(m, 2H), 7.55(br s, 2H), 7.53-7.48 (m, 2H), 7.48-7.36 (m, 2H), 7.31-7.25 (m, 1H), 4.17 (t, J=6.1Hz, 2H), 3.82 (t, J=5.6Hz, 2H), 2.45 (s, 3H), 2.29-2.18 (m, 2H).

[0516] Synthesis of 4'-bromo-2,5-difluoro-1,1'-biphenyl (Int.1A)

[0517] To a stirred solution of compound 1A (5 g, 17.674 mmol) in 1,4-dioxane:HO (50:5 mL) was added (2,5-difluorophenyl)boric acid (3.07 g, 19.441 mmol) and KPO (7.5 g, 35.348 mmol) and the reaction mixture was purged under nitrogen for 10 minutes. PdCl (dppf) (1.29 g, 1.767 mmol) was added to the resulting solution under a nitrogen atmosphere. The reaction mixture was heated at 80 ° C for 1 h. After the reaction was completed (monitored by TLC), the reaction mixture was cooled to room temperature, filtered through a diatomaceous earth pad and washed with ethyl acetate. The filtrate was diluted with water and extracted with EtOAc. The combined organic layer was dried over anhydrous NaSO, filtered and concentrated under reduced pressure. The crude compound was purified by Combiflash chromatography (eluting with 100% heptane) to give the title compound Int. 1A (2.3 g, 48.62%) as an off-white solid. TLC: 100% heptane (Rf :0.5). 1 H NMR (400MHz, CDCl3) δ7.58 (d, J=8.3Hz, 2H), 7.40 (d, J=7.3Hz, 2H), 7.15-7.06 (m, 2H), 7.05-6.97 (m, 1H).

[0518] Example 9: 2-(3-(2',5'-difluoro-[1,1'-biphenyl]-4-yl)-2-oxotetrahydropyrimidin-1(2H)-yl)- Biological test data of 4-methylthiazole-5-sulfonamide (Compound 1)

[0519] Cell culture

[0520] Vero cells were cultured in Dulbecco's modified Eagle's medium (DMEM) supplemented with 10% fetal bovine serum and 100 units / ml of penicillin and streptomycin. Cells were passaged 2-3 times per week to maintain subconfluent density.

[0521] HSV-1 Antiviral Test

[0522] Vero cells were grown at 2.5 × 10 3 Cells were seeded into 96-well plates at a density of 100 cells per well and allowed to adhere overnight. After adherence, the medium was replaced with 50uL infection medium (DMEM supplemented with 2% fetal bovine serum and 100 units / mL penicillin and streptomycin). Compounds were then added to the cultures in an 8-point 3-fold serial dilution format using a Tecan D300e digital dispenser. DMSO concentrations were normalized to 0.5% for all treatments. After compound addition, 80TCID 50 50uL infection medium of HSV-1 was added to the cells and incubated at 37°C for 4 days. After incubation, the plates were equilibrated to room temperature, the medium was removed, and a 1:1 dilution of 60 Cell titer glow and phosphate buffered saline was added to the cells. After 5 minutes of incubation, the brightness was measured using a Tecan Infinite M1000 Pro plate reader to quantify cell viability.

[0523] HSV-2 Antiviral Test

[0524] Vero cells were plated at 1.0 × 10 4 Cells were seeded into 96-well plates at a density of 100 cells / mL and allowed to adhere overnight. After adhesion, the medium was replaced with 50uL infection medium (DMEM supplemented with 2% fetal bovine serum and 100 units / mL penicillin and streptomycin). Compounds were then added to the cultures in an 8-point 3-fold serial dilution format using a Tecan D300e digital dispenser. DMSO concentrations for all treatments were normalized to 0.5%. After compound addition, a 160TCID 5050uL infection medium of HSV-2G strain was added to the cells and incubated at 37°C for 5 days. After incubation, 10μL / well WST-8 colorimetric reagent was added and the plate was incubated at 37°C for 3 hours. After incubation, cell viability was quantified by measuring absorbance at 460nm and 620nm using a Tecan Infinite M1000 Pro plate reader.

[0525] In the HSV-1 antiviral test, compound 1 had an EC 50 In the HSV-2 antiviral test, compound 1 had an EC of 0.019 mM (n=22). 50 It is 0.011mM (n=35).

[0526] Example 10: PK study of compound 1 in rats

[0527] Preparation

[0528] An aqueous suspension of Compound 1 free form (Form A) at a concentration of 3 mg / mL in a 0.9% NaCl + 0.5% polysorbate 80 vehicle was prepared as follows:

[0529] 1) The vehicle (0.9% w / w NaCl and 0.5% w / w polysorbate 80) was prepared by dissolving NaCl and polysorbate 80 in water.

[0530] 2) An appropriate amount of Compound 1 free form was weighed and added to the carrier.

[0531] 3) The mixture was vortexed for 1 minute, stirred for 10 minutes, and then sonicated for 15 minutes.

[0532] 4) The final dosage form is a homogeneous white turbid suspension.

[0533] Formulation Characterization

[0534] The particle size distribution (PSD) of the suspension was determined as Dv 10 =0.57μm, Dv 50 =2.41μm and Dv 90 =14.54μm.

[0535] Animal dosing

[0536] Male Sprague Dawley (SD) rats (6-8 weeks old) with a body weight of 230-250 g were used for pharmacokinetic studies. Animals had free access to food and water. The above compound 1 suspension was administered to each animal group (n=3) by subcutaneous injection, and the administration volume was 3.33 mL / kg to achieve a 10 mg / kg dose.

[0537] At each time point, approximately 200 μL of whole blood was collected via the jugular vein in a K2EDTA tube. The blood samples were placed on ice and centrifuged at 2000 g for 5 minutes to obtain plasma samples within 15 minutes. The plasma samples were stored at approximately -70°C until analysis by LC-MS.

[0538] result

[0539] Fig.10 Mean plasma concentration curves (n=3) following subcutaneous (SC) administration of a 10 mg / kg suspension of the compound are shown. The exposure period was extended to at least 58 days. The variability of plasma concentrations was low.

[0540] Example 11: PK study of compound 1 in monkeys

[0541] Preparation

[0542] An aqueous suspension of Compound 1 free form (Form A) at a concentration of 3.0 mg / mL in a 0.9% NaCl + 0.5% polysorbate 80 vehicle was prepared as follows:

[0543] 1) The vehicle (0.9% w / w NaCl and 0.5% w / w polysorbate 80) was prepared by dissolving NaCl and polysorbate 80 in water.

[0544] 2) An appropriate amount of Compound 1 was weighed and added to the carrier.

[0545] 3) The mixture was stirred for 15 minutes, then vortexed for 1 minute, and then sonicated for 10 minutes.

[0546] 4) The final dosage form is a homogeneous white turbid suspension.

[0547] Animal dosing

[0548] Male non-naive cynomolgus monkeys with a body weight of 4-6 kg were used for the monkey PK study. Animals had free access to food and water. The prepared suspension was administered by subcutaneous injection to each animal group (n=3) at a volume of 3.33 mL / kg to achieve a 10 mg / kg dose.

[0549] At each time point, approximately 0.5 mL of whole blood was collected into K2EDTA tubes via the cephalic and saphenous veins. Blood samples were placed on ice and centrifuged at 2000 g for 5 minutes to obtain plasma samples within 15 minutes. Plasma samples were stored at approximately -70°C until analyzed by LC-MS.

[0550] result

[0551] The mean plasma concentration curves after subcutaneous (SC) administration of Compound 1 suspension (n=3) are shown in Fig.11 Notably, after the initial drug burst, extended exposure of at least 220 days was achieved at the 10 mg / kg administered dose level.

[0552] Example 12: PK study of compound 1 in dogs

[0553] Preparation

[0554] An aqueous suspension of Compound 1 free form (Form A) at a concentration of 5.0 mg / mL in a 0.9% NaCl + 0.5% Poloxamer 188 vehicle was prepared as follows:

[0555] 1) Vehicle (0.9% w / w NaCl and 0.5% w / w Poloxamer 188) was prepared by dissolving NaCl and Poloxamer 188 in water.

[0556] 2) Weigh an appropriate amount of Compound 1 free form and add to the carrier.

[0557] 3) The mixture was stirred for 15 minutes, then vortexed for 1 minute, and then sonicated for 10 minutes.

[0558] 4) The final dosage form is a homogeneous white turbid suspension.

[0559] Animal dosing

[0560] Male non-naive beagles with a body weight of ~10 kg were used for the PK study in dogs. Animals had free access to food and water. The prepared suspension was administered by subcutaneous injection to each animal group (n=3) at a volume of 2 mL / kg to achieve a dose of 10 mg / kg.

[0561] At each time point, approximately 0.5 mL of whole blood was collected into K2EDTA tubes. The blood samples were placed on ice and centrifuged at 2000 g for 5 minutes to obtain plasma samples within 15 minutes. The plasma samples were stored at approximately -70°C until analysis by LC-MS.

[0562] result

[0563] Fig.12 Mean plasma concentration curves (n=3) following subcutaneous (SC) administration of Compound 1 suspension are shown. After the initial drug burst, prolonged exposure of at least 220 days was achieved at the 10 mg / kg administration dose level.

[0564] Summary of PK studies of compounds 10-12

[0565] Injectable long-acting depot formulations containing Compound 1 have been found to provide sustained and stable plasma levels of the drug for significantly prolonged periods of time following a single subcutaneous administration in 3 different species.

[0566] Furthermore, no adverse reactions were observed at the injection site in the study animals.

[0567] Example 13: PK studies in rats, monkeys and dogs after intravenous injection

[0568] Preparation - Rat

[0569] A 0.2 mg / mL solution of Compound 1 in 10% NMP, 10% Solutol HS15, and 80% saline was prepared as follows:

[0570] 1) Prepare a stock solution of compound 1 in NMP.

[0571] 2) Combine equal volumes of Compound 1 stock solution and Solutol HS15 and mix.

[0572] 3) Dilute the above solution with saline to reach the target concentration. The final preparation is a clear solution.

[0573] Animal Dosing - Rats

[0574] Male Sprague Dawley (SD) rats weighing 230-250 g and aged 6-8 weeks were used for rat pharmacokinetic studies. Animals had free access to food and water. The above-mentioned Compound 1 solution was administered to animals (n=3) by intravenous injection to achieve a dose of 1.0 mg / kg.

[0575] At each time point, approximately 200 μL of whole blood was collected in K2EDTA tubes. The blood samples were placed on ice and centrifuged at 2000 g for 5 minutes to obtain plasma samples within 15 minutes. The plasma samples were stored at approximately -70°C until analyzed by LC-MS according to the above method.

[0576] result

[0577] The mean plasma concentration curves after intravenous (IV) administration of Compound 1 solution at 1.0 mg / kg to rats (n=3) can be found in Fig.13 .

[0578] Preparation-Monkey

[0579] A 0.25 mg / mL solution of Compound 1 in 10% DMSO, 10% Solutol HS15, and 80% saline was prepared as follows:

[0580] 1) Prepare a stock solution of Compound A 1 in DMSO.

[0581] 2) Combine equal volumes of Compound 1 stock solution and Solutol HS15 and mix.

[0582] 3) Dilute the above solution with saline to reach the target concentration. The final preparation is a clear solution.

[0583] Animal administration - monkey

[0584] Male non-naive cynomolgus monkeys were used for monkey PK study. Animals had free access to food and water. The prepared solution was administered to each animal group (n=3) by intravenous injection to achieve a dose of 0.25 mg / kg.

[0585] At each time point, approximately 0.5 mL of whole blood was collected into K2EDTA tubes. Blood samples were placed on ice and centrifuged at 2000 g for 5 minutes to obtain plasma samples within 15 minutes. Plasma samples were stored at approximately -70°C until analysis by LC-MS.

[0586] result

[0587] The mean plasma concentration curves (n=3) after intravenous (IV) administration of 0.25 mg / kg Compound 1 solution can be found in Fig.14 .

[0588] Preparation - Dog

[0589] A 0.15 mg / mL solution of Compound 1 in 10% DMSO, 10% Solutol HS15, and 80% saline was prepared as follows:

[0590] 1) Prepare a stock solution of Compound A 1 in DMSO.

[0591] 2) Combine equal volumes of Compound 1 stock solution and Solutol HS15 and mix.

[0592] 3) Dilute the above solution with saline to reach the target concentration. The final preparation is a clear solution.

[0593] Animal Drug Administration - Dogs

[0594] Male non-naive beagles were used for the PK study in dogs. The animals had free access to food and water. The prepared solution was administered by subcutaneous injection to each animal group (n=3) to achieve a dose of 0.15 mg / kg.

[0595] At each time point, approximately 0.5 mL of whole blood was collected into K2EDTA tubes. The blood samples were placed on ice and centrifuged at 2000 g for 5 minutes to obtain plasma samples within 15 minutes. The plasma samples were stored at approximately -70°C until analysis by LC-MS.

[0596] result

[0597] The mean plasma concentration curves (n=3) after intravenous (IV) administration of 0.15 mg / kg Compound 1 solution can be found in Fig.15 .

[0598] Summarize

[0599] IV administration of Compound 1 to rats, monkeys and dogs at doses of 1 mg / kg, 0.25 mg / kg and 0.15 mg / kg, respectively, resulted in long terminal half-lives, as shown in the following table:

[0600]

[0601] Example 13A: Further PK studies of Compound 1 after intravenous injection in rats, monkeys, dogs and mini-pigs

[0602] Preparation - Rat

[0603] A 0.2 mg / mL solution of Compound 1 in 10% NMP, 10% Solutol HS15, and 80% saline was prepared as follows:

[0604] 1) Prepare a stock solution of compound 1 in NMP.

[0605] 2) Combine equal volumes of Compound 1 stock solution and Solutol HS15 and mix.

[0606] 3) Dilute the above solution with saline to reach the target concentration. The final preparation is a clear solution.

[0607] Animal Dosing - Rats

[0608] Male Sprague Dawley (SD) rats weighing 180-184 g and aged 6-8 weeks were used for rat pharmacokinetic studies. Animals had free access to food and water. The above-mentioned Compound 1 solution was administered to each animal (n=3) by intravenous injection (1 mL / kg) to achieve a dose of 0.2 mg / kg.

[0609] At each time point, approximately 200 μL of whole blood was collected in a K2EDTA tube. The blood samples were placed on ice and centrifuged at 2000 g for 5 minutes to obtain plasma samples within 15 minutes. The plasma samples were stored at approximately -70°C until analyzed by LC-MS according to the above method.

[0610] Preparation-Monkey

[0611] A 0.2 mg / mL solution of Compound 1 in 10% DMSO, 10% Solutol HS15, and 80% saline was prepared as follows:

[0612] 1) Prepare a stock solution of Compound A 1 in DMSO.

[0613] 2) Combine equal volumes of Compound 1 stock solution and Solutol HS15 and mix.

[0614] 3) Dilute the above solution with saline to reach the target concentration. The final preparation is a clear solution.

[0615] Animal administration - monkey

[0616] Male non-naive cynomolgus monkeys were used for the monkey PK study. The animals had free access to food and water. The above Compound 1 solution was administered (1 mL / kg) to each animal (n=3) by intravenous injection to achieve a dose of 0.2 mg / kg.

[0617] At each time point, approximately 0.5 mL of whole blood was collected into K2EDTA tubes. Blood samples were placed on ice and centrifuged at 2000 g for 5 minutes to obtain plasma samples within 15 minutes. Plasma samples were stored at approximately -70°C until analysis by LC-MS.

[0618] Preparation - Dog

[0619] A 0.15 mg / mL solution of Compound 1 in 10% DMSO, 10% Solutol HS15, and 80% saline was prepared as follows:

[0620] 1) Prepare a stock solution of Compound A 1 in DMSO.

[0621] 2) Combine equal volumes of Compound 1 stock solution and Solutol HS15 and mix.

[0622] 3) Dilute the above solution with saline to reach the target concentration. The final preparation is a clear solution.

[0623] Animal Drug Administration - Dogs

[0624] Male non-naive beagles were used for the PK study in dogs. Animals had free access to food and water. The above Compound 1 solution was administered (1 mL / kg) to each animal (n=3) by intravenous injection to achieve a dose of 0.15 mg / kg.

[0625] At each time point, approximately 0.5 mL of whole blood was collected into K2EDTA tubes. The blood samples were placed on ice and centrifuged at 2000 g for 5 minutes to obtain plasma samples within 15 minutes. The plasma samples were stored at approximately -70°C until analysis by LC-MS.

[0626] Preparation-Mini Pig

[0627] A 0.25 mg / mL solution of Compound 1 in 10% DMSO, 10% Solutol HS15, and 80% saline was prepared as follows:

[0628] 4) Prepare a stock solution of Compound A 1 in DMSO.

[0629] 5) Combine equal volumes of Compound 1 stock solution and Solutol HS15 and mix.

[0630] 6) Dilute the above solution with saline to reach the target concentration. The final preparation is a clear solution.

[0631] Animal Drug Delivery - Mini Pigs

[0632] Naive Bama pigs (15-16 kg) were used for minipig PK study. Animals had free access to food and water. The above Compound 1 solution was administered to each animal (n=3) by intravenous injection (1 mL / kg) to achieve a dose of 0.25 mg / kg.

[0633] At each time point, approximately 0.5 mL of whole blood was collected into K2EDTA tubes. The blood samples were placed on ice and centrifuged at 2000 g for 5 minutes to obtain plasma samples within 15 minutes. The plasma samples were stored at approximately -70°C until analysis by LC-MS.

[0634] result

[0635] The mean plasma concentration curves after intravenous (IV) injection of Compound 1 solution in rats, dogs, monkeys and mini-pigs (n=3) can be found in Fig.19 (A)-(D).

[0636] Summarize

[0637] The terminal half-life and clearance rates were shown in the following table after IV administration of 0.2 mg / kg, 0.15 mg / kg, 0.2 mg / kg and 0.25 mg / kg to rats, dogs, monkeys and mini pigs, respectively:

[0638]

[0639]

[0640] Based on the PK data in multiple species above and using allometric scaling, the terminal biological half-life of compound 1 in humans is predicted to be 7.6 days (182 hours) and the clearance rate is 0.06 L / hour.

[0641] Example 14: Subcutaneous PK study of priligy in dogs

[0642] (A) Preparation

[0643] Solution preparation:

[0644] A solution of 100 mg / mL of priligyvir in 90% NMP (Sigma) and 10% Solutol HS15 (BASF) was prepared as follows:

[0645] 1) Weigh about 460 mg of crystalline Priligy free base sesquihydrate (based on assay) and add to about 3.9 mL of NMP. Stir the mixture for 10 minutes.

[0646] 2) After the Priligy was dissolved, Solutol HS15 (about 0.4 mL) was added to the solution and mixed for another 2 minutes.

[0647] 3) The final dosage form is a clear solution.

[0648] Aqueous micron suspension:

[0649] An aqueous microsuspension of crystalline priligyvir free base (sesquihydrate) at a concentration of 100 mg / mL in 2 mg / mL polysorbate 80 (Tween 80) and 5 mg / mL Methocel A4M vehicle was prepared as follows:

[0650] 1) Crystalline priligy free base (sesquihydrate) was jet milled [milling pressure 0.8 mPa] to obtain microparticles (Dv90 < 10 um).

[0651] 2) About 460 mg of the jet milled material (based on the assay) was then dispersed in 2 mg / mL Polysorbate 80 (Tween 80) and 5 mg / ml Methocel A4M solution (~4.4 mL) to obtain a 100 mg / ml suspension.

[0652] 3) The mixture was stirred for 10 minutes.

[0653] 4) The final dosage form is a homogeneous white turbid suspension.

[0654] Formulation Characterization of Microsuspensions:

[0655] The particle size distribution (PSD) of the micron suspension is determined as Dv 10 =1.96μm, Dv 50 =4.17μm and Dv 90 =8.51μm.

[0656] SEM images of the micron suspension can be found in Fig. 20 .

[0657] Aqueous Nanosuspension:

[0658] An aqueous nanosuspension of crystalline priligyr free base (sesquihydrate) at a concentration of 100 mg / mL in a vehicle of 10 mg / mL NaCMC, 2 mg / mL polysorbate 80 (Tween 80), and 50 mg / mL mannitol was prepared as follows:

[0659] 1) Disperse 10 mg / mL crystalline priligyvir free base (sesquihydrate) into a solution of 1 mg / mL NaCMC + 0.2 mg / mL polysorbate 80 (Tween 80) and 5 mg / mL mannitol.

[0660] 2) The suspension was placed in a high pressure homogenizer (about 20 cycles at 600 bar, followed by 130 cycles at 900 bar) until the particle size was about 1 μm.

[0661] 3) After reducing the particle size, the suspension is freeze-dried to obtain a freeze-dried powder.

[0662] 4) Reconstitute the lyophilized powder with water to obtain a 100 mg / ml suspension.

[0663] 5) The suspension was wet-milled (10:1 0.1 mm zirconium oxide grinding beads: priligy) at 700 rpm for 500 minutes to reduce the particle size to nano-sized particles (Dv90 < 500 nm).

[0664] 6) freeze-drying the nano suspension to obtain freeze-dried powder.

[0665] 7) Weigh an appropriate amount of lyophilized powder (based on the assay) and add to water to obtain a 100 mg / ml suspension.

[0666] 8) The mixture was stirred for 10 minutes.

[0667] 9) The final dosage form is a homogeneous white turbid suspension.

[0668] The SEM images of the nanosuspension can be found in Fig.21 .

[0669] (B) Animal drug administration

[0670] Male non-naive beagles were used for the PK study in dogs. Animals had free access to food and water. The prepared formulation was injected subcutaneously into each animal group (n=3) at a dosing volume of 0.1 mL / kg to achieve a dose of 10 mg / kg.

[0671] At each time point, approximately 0.5 mL of whole blood was collected into K2EDTA tubes. The blood samples were placed on ice and centrifuged at 2000 g for 5 minutes to obtain plasma samples within 15 minutes. The plasma samples were stored at approximately -70°C until analysis by LC-MS.

[0672] result

[0673] Fig. 22 The mean plasma concentration curves (n=3) after subcutaneous (SC) administration of pralivir preparations are shown. Using these three preparations, extended exposure is achieved at the dosage levels applied. The solution shows the highest plasma concentration levels at early time points. Surprisingly, for the aqueous suspension preparations at early time points (up to 234 hours), higher pralivir plasma concentrations are obtained by micron suspensions, rather than nanosuspension preparations. The preparation can achieve extended release using high concentrations of pralivir, so it can be administered using a small administration volume of only 0.1 mL / kg. In addition, no adverse reactions were observed at the injection site of the study animals.

[0674] Example 15: Further PK studies of compound 1 in rats

[0675] Preparation

[0676] A solution of Compound 1 at a concentration of 50 mg / mL in 90% NMP and 10% Solutol HS15 was prepared as follows:

[0677] 1) An appropriate amount of crystalline Compound 1 (Form C) was weighed and added to NMP.

[0678] 2) After Compound 1 is dissolved, add Solutol HS15 to the solution and mix [specify mixing conditions and time].

[0679] 3) The final dosage form is a clear solution.

[0680] Animal dosing

[0681] Male Sprague Dawley (SD) rats (6-8 weeks old) with a body weight of 230-250 g were used for pharmacokinetic studies. Animals had free access to food and water. The above-mentioned Compound 1 solution was administered to each animal group (n=3) by subcutaneous injection, and the administration volume was 2 mL / kg to achieve a 100 mg / kg dose.

[0682] At each time point, approximately 150 μL of whole blood was collected in a K2EDTA tube via the jugular vein. The blood samples were placed on ice and centrifuged at 2000 g for 5 minutes to obtain plasma samples within 15 minutes. The plasma samples were stored at approximately -70°C until analyzed by LC-MS.

[0683] result

[0684] Fig.23 The mean plasma concentration curves after subcutaneous administration of Compound 1 solution at a dose of 50 mg / kg are shown (n=3). After the initial drug burst, an extended exposure of 960 hours (study duration) was achieved at the administered dose level. The variability of plasma concentrations was low. In addition, no adverse reactions were observed at the injection site of the study animals.

[0685] Example 16: Further PK studies of Compound 1 in dogs

[0686] Preparation

[0687] A solution of compound 1 at a concentration of 50 mg / mL in 90% NMP and 10% Solutol HS15 was prepared as follows:

[0688] 1) An appropriate amount of crystalline Compound 1 (Form C) was weighed and added to NMP.

[0689] 2) After Compound 1 was dissolved, Solutol HS15 was added to the solution and mixed.

[0690] 3) The final dosage form is a clear solution.

[0691] A solution of Compound 1 at a concentration of 100 mg / mL in 90% NMP and 10% Solutol HS15 was prepared as follows:

[0692] 1) An appropriate amount of crystalline Compound 1 (Form C) was weighed and added to NMP.

[0693] 2) After Compound 1 was dissolved, Solutol HS15 was added to the solution and mixed.

[0694] 3) The final dosage form is a clear solution.

[0695] Animal dosing

[0696] Male non-naive beagles were used for the PK study in dogs. Animals had free access to food and water. The prepared 50 mg / mL solution was administered to each animal group (n=3) by subcutaneous injection, and the administration volume was 0.2 mL / kg to achieve a 10 mg / kg dose. The prepared 100 mg / mL solution was administered to each animal group (n=3) by subcutaneous injection, and the administration volume was 0.1 mL / kg to achieve a 10 mg / kg dose.

[0697] At each time point, approximately 0.5 mL of whole blood was collected into K2EDTA tubes. The blood samples were placed on ice and centrifuged at 2000 g for 5 minutes to obtain plasma samples within 15 minutes. The plasma samples were stored at approximately -70°C until analysis by LC-MS.

[0698] result

[0699] Fig.24 The mean plasma concentration curves after subcutaneous administration of Compound 1 solution in two dosing volumes are shown (n=3). Using two formulations, extended exposure of at least 60 days (study duration) was achieved at the administered dose level. The formulations were able to achieve extended release using small dosing volumes. In addition, no adverse reactions were observed at the injection site of the study animals.

[0700] Example 17: Subcutaneous and intramuscular administration of Compound 1 in rats using microsuspension and nanosuspension formulations PK study of internal administration

[0701] A) Preparation

[0702] Aqueous micron suspension:

[0703] An aqueous microsuspension of crystalline Compound 1 free form (crystalline Form C) at a concentration of 200 mg / mL in a vehicle of 20 mg / mL PVP K30, 10 mg / mL poloxamer 188, and 50 mg / mL mannitol was prepared as follows:

[0704] 1) Crystalline Compound 1 (Form C) was jet milled to obtain microparticles (Dv90 < 10 μm).

[0705] 2) The jet milled material was dispersed in 20 mg / ml PVP K30 + 10 mg / ml Poloxamer 188 + 50 mg / ml mannitol to obtain a homogeneous suspension.

[0706] 3) Freeze-drying the suspension to obtain freeze-dried powder.

[0707] 4) Weigh an appropriate amount of lyophilized powder (according to the assay) and add it to water to obtain a 200 mg / ml suspension.

[0708] 5) Stir the mixture for 5 minutes.

[0709] 6) The final dosage form is a homogeneous white turbid suspension.

[0710] Formulation Characterization of Microsuspensions:

[0711] The particle size distribution (PSD) of the micron suspension is determined as Dv 10 =1.19μm, Dv 50 =2.76μm and Dv90 =5.65μm.

[0712] SEM images of micron suspensions can be found in Fig.25 .

[0713] Aqueous Nanosuspension:

[0714] An aqueous nanosuspension of crystalline Compound 1 free form (Form C) at a concentration of 200 mg / mL in a vehicle of 20 mg / mL PVP K30, 10 mg / mL Poloxamer 188, and 50 mg / mL mannitol was prepared as follows:

[0715] 1) Disperse 20 mg / mL crystalline Compound 1 (Form C) into a solution of 2 mg / mL PVP K30, 1.0 mg / mL Poloxamer 188, and 5 mg / mL mannitol

[0716] 2) The suspension was placed in a high pressure homogenizer until the size was about 1 μm.

[0717] 3) After reducing the particle size, the suspension is freeze-dried to obtain a lyophilized powder.

[0718] 4) The lyophilized powder was reconstituted with water to obtain a 100 mg / mL suspension.

[0719] 5) The suspension was wet milled (10:1 0.1 mm zirconium oxide grinding beads: Compound 1) at 900 rpm for 600 minutes to reduce the particle size to nano-scale particles (Dv90 < 200 nm).

[0720] 6) The nanosuspension is freeze-dried to obtain freeze-dried powder.

[0721] 7) Weigh an appropriate amount of lyophilized powder (based on the assay) and add to water to obtain a 200 mg / ml suspension.

[0722] 8) The mixture was stirred for 10 minutes.

[0723] 9) The final dosage form is a uniform white turbid suspension.

[0724] Formulation Characterization of Nanosuspensions:

[0725] The particle size distribution (PSD) of the nanosuspension is determined as Dv 10 =69.1nm, Dv50=77.7nm, and Dv90=88.5nm.

[0726] The SEM images of the nanosuspensions can be seen in Fig.26 .

[0727] (B) Animal administration

[0728] Male Sprague Dawley (SD) rats (6-8 weeks old) with a body weight of 230-250 g were used for pharmacokinetic studies. Animals had free access to food and water. The above-mentioned Compound 1 solution was administered subcutaneously or intramuscularly to each animal group (n=3) in a volume of 0.1 mL / animal (~0.4 mL / kg) to achieve a 20 mg / animal dose (~80 mg / kg).

[0729] At each time point, approximately 150 μL of whole blood was collected in a K2EDTA tube via the jugular vein. The blood samples were placed on ice and centrifuged at 2000 g for 5 minutes to obtain plasma samples within 15 minutes. The plasma samples were stored at approximately -70°C until analyzed by LC-MS.

[0730] result

[0731] The mean plasma concentration curves (n=3) after subcutaneous administration of Compound 1 micron and nanosuspensions can be found in Fig. 27 Both formulations were able to provide extended exposure times of at least 600 hours (the length of the study). The nanosuspension provided higher plasma levels than the microsuspension throughout the testing period. These formulations were able to achieve extended release using high drug loads and small dosing volumes. In addition, no adverse reactions were observed at the injection site in the study animals.

[0732] The mean plasma concentration curves (n=3) after intramuscular administration of compound 1 micron suspension and nanosuspension can be found in Fig.28 Both formulations were able to provide extended exposure for at least 600 hours (the length of the study) and plasma levels were higher than those achieved when these formulations were administered by subcutaneous injection. The nanosuspension provided higher plasma levels than the microsuspension. These formulations were able to achieve extended release using high drug loads and small dosing volumes. In addition, no adverse reactions were observed at the injection site in the study animals.

Claims

1. A long-acting injectable reservoir-type pharmaceutical composition comprising: a. a helicase-primase inhibitor or a pharmaceutically acceptable salt thereof; and b. one or more pharmaceutically acceptable excipients; The helicase-primase inhibitor has a terminal half-life in humans of 10 hours or more.

2. The composition according to claim 1, wherein the composition is a subcutaneous or intramuscular long-acting injectable depot pharmaceutical composition.

3. The composition according to claim 2, wherein the composition is for administration twice a month, once a month, once every two months, once every three months, once every six months or once a year.

4. A composition according to any one of claims 1 to 3, wherein the helicase-primase inhibitor has an in vitro EC of 1.5 for HSV-1 and / or HSV-2. 50 The value was less than 0.1 μM.

5. The composition according to any one of claims 1 to 4, wherein the helicase-primase inhibitor is N-methyl-N-(4-methyl-5-sulfamoyl-1,3-thiazol-2-yl)-2-[4-(pyridin-2-yl)phenyl]acetamide: or a pharmaceutically acceptable salt thereof.

6. A composition according to any one of claims 1 to 5, wherein the helicase-primase inhibitor is N-methyl-N-(4-methyl-5-sulfamoyl-1,3-thiazol-2-yl)-2-[4-(pyridin-2-yl)phenyl]acetamide free base.

7. A composition according to claim 6, wherein the free base is present as a crystalline solid.

8. The composition according to claim 6, wherein the long-acting injectable depot pharmaceutical composition is an aqueous suspension and the free base is suspended as a crystalline solid.

9. The composition according to claim 8, wherein the long-acting injectable depot pharmaceutical composition is an aqueous nanosuspension and the free base is suspended as a crystalline solid.

10. The composition according to any one of claims 1 to 6, wherein the long-acting injectable depot pharmaceutical composition comprises a formulation based on a solution in an organic solvent.

11. The composition according to claim 10, wherein the organic solvent is selected from the group consisting of benzyl alcohol, benzyl benzoate, N-methyl-pyrrolidone (NMP), 2-pyrrolidone, ethanol, propylene glycol, acetone, methyl acetate, ethyl acetate, methyl ethyl ketone, dimethylacetamide (DMA), dimethylformamide, dimethyl sulfoxide (DMSO), tetrahydrofuran, caprolactam, oleic acid, 1-dodecylazacycloheptan-2-one, and mixtures thereof.

12. The composition according to claim 10, wherein the helicase-primase inhibitor is N-methyl-N-(4-methyl-5-sulfamoyl-1,3-thiazol-2-yl)-2-[4-(pyridin-2-yl)phenyl]-acetamide and the organic solvent is selected from NMP, DMA and DMSO.

13. A composition according to any one of claims 1 to 4, wherein the helicase-primase inhibitor is 2-(3-(2',5'-difluoro-[1,1'-biphenyl]-4-yl)-2-oxotetrahydropyrimidin-1(2H)-yl)-4-methyl-thiazole-5-sulfonamide (Compound 1): or a pharmaceutically acceptable salt thereof.

14. The composition according to any one of claims 1 to 4, wherein the helicase-primase inhibitor is 2-(3-(2',5'-difluoro-[1,1'-biphenyl]-4-yl)-2-oxotetrahydropyrimidin-1(2H)-yl)-4-methylthiazole-5-sulfonamide free form.

15. A composition according to claim 14, wherein the free form is present as a crystalline solid.

16. The composition according to claim 14, wherein the long-acting injectable depot pharmaceutical composition is an aqueous suspension and the free form is suspended as a crystalline solid.

17. The composition according to claim 16, wherein the long-acting injectable depot pharmaceutical composition is an aqueous microsuspension or nanosuspension and the free form is suspended as a crystalline solid.

18. The composition according to claim 13 or 14, wherein the long-acting injectable depot pharmaceutical composition comprises a formulation based on a solution in an organic solvent.

19. The composition according to claim 18, wherein the organic solvent is selected from the group consisting of N-methylpyrrolidone (NMP), dimethylacetamide (DMA), dimethyl sulfoxide (DMSO), benzyl alcohol, benzyl benzoate and mixtures thereof.

20. The composition according to claim 19, wherein the organic solvent is selected from the group consisting of NMP, DMA and DMSO.

21. The composition according to any one of claims 1 to 20, wherein the long-acting injectable depot pharmaceutical composition comprises from about 2 wt% to about 40 wt% of the helicase-primase inhibitor or a pharmaceutically acceptable salt thereof.

22. A composition according to any one of claims 1 to 21, wherein the long-acting injectable depot pharmaceutical composition comprises about 100 mg to about 1200 mg, about 10 mg to about 900 mg, or about 100 mg to about 600 mg of the helicase-primase inhibitor or a pharmaceutically acceptable salt thereof.

23. The composition of any one of claims 1 to 22, wherein the long-acting injectable depot pharmaceutical composition comprises N-methyl-N-(4-methyl-5-sulfamoyl-1,3-thiazol-2-yl)-2-[4-(pyridin-2-yl)phenyl]acetamide free base as a helicase-primase inhibitor and the composition upon administration produces in a subject a plasma concentration of N-methyl-N-(4-methyl-5-sulfamoyl-1,3-thiazol-2-yl)-2-[4-(pyridin-2-yl)phenyl]acetamide of at least 25 ng / mL for at least 80% of a dosing interval, wherein the dosing interval is at least 10 days.

24. The composition of any one of claims 1 to 22, wherein the long-acting injectable depot pharmaceutical composition comprises 2-(3-(2',5'-difluoro-[1,1'-biphenyl]-4-yl)-2-oxotetrahydropyrimidin-1(2H)-yl)-4-methylthiazole-5-sulfonamide free form as a helicase-initase inhibitor and the composition upon administration produces in a subject a plasma concentration of 2-(3-(2',5'-difluoro-[1,1'-biphenyl]-4-yl)-2-oxotetrahydropyrimidin-1(2H)-yl)-4-methylthiazole-5-sulfonamide of at least 25 ng / mL for at least 80% of a dosing interval, wherein the dosing interval is at least 10 days.

25. A composition according to any one of claims 1 to 24, wherein the composition is stable for at least 4 weeks.

26. A composition according to any one of claims 1 to 25 for use as a medicament.

27. A method for treating a herpes virus (such as HSV) infection in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of a helicase-primase inhibitor or a pharmaceutically acceptable salt thereof, wherein the helicase-primase inhibitor or a pharmaceutically acceptable salt thereof is formulated as a pharmaceutical composition according to any one of claims 1 to 25.

28. The method of claim 27, wherein the pharmaceutical composition is administered twice a month, once a month, once every two months, once every three months, once every six months, or once a year.

29. The method of claim 27, wherein the pharmaceutical composition is administered once a month or once every three months.

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