WO2021038480A1

WO2021038480A1 - Combinations for treating influenza virus

Info

Publication number: WO2021038480A1
Application number: PCT/IB2020/058000
Authority: WO
Inventors: Jin Wu; Johan Hendrika Jozef Vingerhoets; Dirk André Emmy ROYMANS
Original assignee: Janssen Pharmaceuticals, Inc.
Priority date: 2019-08-27
Filing date: 2020-08-27
Publication date: 2021-03-04

Abstract

The present invention relates to combinations comprising Compound (1) and baloxavir or a prodrug thereof. The present invention also relates to pharmaceutical compositions comprising a combination of Compound (1) and baloxavir, as well as processes for producing the pharmaceutical compositions. The invention further relates to methods of treating influenza virus infection in a subject or biological sample infected with influenza viruses, as well as methods of inhibiting the replication of, or reducing the number of influenza viruses in an in vitro biological sample.

Description

COMBINATIONS FOR TREATING INFLUENZA VIRUS

FIELD OF THE INVENTION

[0001] The present invention relates to combinations comprising Compound (1) and baloxavir. The present invention also relates to pharmaceutical compositions comprising a combination of Compound (1) and baloxavir, as well as processes for producing the pharmaceutical compositions. The invention further relates to methods of treating influenza virus infection in a subject or biological sample infected with influenza viruses, as well as methods of inhibiting the replication of, or reducing the number of influenza viruses in an in vitro biological sample.

BACKGROUND OF THE INVENTION

[0002] Influenza is primarily transmitted from person to person via large virus-laden droplets that are generated when infected persons cough or sneeze; these large droplets can then settle on the mucosal surfaces of the upper respiratory tracts of susceptible individuals who are near (e.g. within 6 feet) infected persons. Transmission might also occur through direct contact or indirect contact with respiratory secretions, such as touching surfaces contaminated with influenza virus and then touching the eyes, nose or mouth. Adults might be able to spread influenza to others from 1 day before getting symptoms to approximately 5 days after symptoms start. Young children and persons with weakened immune systems might be infectious for 10 or more days after onset of symptoms.

[0003] Influenza viruses are RNA viruses of the family Orthomyxoviridae, which comprises seven genera: Influenza A virus, Influenza B virus, Influenza C virus, Influenza D virus, ISA virus, Thogoto virus, and Quaranja virus.

[0004] The Influenza A virus genus has one species, influenza A virus. Wild aquatic birds are the natural hosts for a large variety of influenza A. Occasionally, viruses are transmitted to other species and may then cause devastating outbreaks in domestic poultry or give rise to human influenza pandemics. The type A viruses are the most virulent human pathogens among the three influenza types and cause the most severe disease. The influenza A virus can be subdivided into different serotypes based on the antibody response to these viruses. The serotypes that have been confirmed in humans, ordered by the number of known human pandemic deaths, are: H1N1 (which caused Spanish influenza in 1918), H2N2 (which caused Asian Influenza in 1957), H3N2 (which caused Hong Kong Flu in 1968), H5N1 (a pandemic threat in the 2007-08 influenza season), H7N7 (which has unusual zoonotic potential), H1N2 (endemic in humans and pigs), H9N2, H7N2, H7N3 and H10N7.

[0005] The Influenza B virus genus has one species, influenza B virus. Influenza B almost exclusively infects humans and is less common than influenza A. The only other animal known to be susceptible to influenza B infection is the seal. This type of influenza mutates at a rate 2-3 times slower than type A and consequently is less genetically diverse, with only one influenza B serotype. As a result of this lack of antigenic diversity, a degree of immunity to influenza B is usually acquired at an early age. However, influenza B mutates enough that lasting immunity is not possible. This reduced rate of antigenic change, combined with its limited host range (inhibiting cross species antigenic shift), ensures that pandemics of influenza B do not occur.

[0006] The Influenza C virus genus has one species, influenza C virus, which infects humans and pigs and can cause severe illness and local epidemics. However, influenza C is less common than the other types and usually seems to cause mild disease in children. [0007] Influenza A, B and C viruses are very similar in structure. The virus particle is

80-120 nanometers in diameter and usually roughly spherical, although filamentous forms can occur. Unusually for a virus, its genome is not a single piece of nucleic acid; instead, it contains seven or eight pieces of segmented negative-sense RNA. The Influenza A genome encodes 11 proteins: hemagglutinin (HA), neuraminidase (NA), nucleoprotein (NP), Ml, M2, NS1, NS2(NEP), PA, PB1, PB1-F2 and PB2.

[0008] HA and NA are large glycoproteins on the outside of the viral particles. HA is a lectin that mediates binding of the virus to target cells and entry of the viral genome into the target cell, while NA is involved in the release of progeny virus from infected cells, by cleaving sugars that bind the mature viral particles. Thus, these proteins have been targets for antiviral drugs. Furthermore, they are antigens to which antibodies can be raised. Influenza A viruses are classified into subtypes based on antibody responses to HA and NA, forming the basis of the H and N distinctions (vide supra ) in, for example, H5N1. [0009] Influenza produces direct costs due to lost productivity and associated medical treatment, as well as indirect costs of preventative measures. In the United States, influenza is responsible for a total cost of over $10 billion per year, while it has been estimated that a future pandemic could cause hundreds of billions of dollars in direct and indirect costs. Preventative costs are also high. Governments worldwide have spent billions ofU.S. dollars preparing and planning for a potential H5N1 avian influenza pandemic, with costs associated with purchasing drugs and vaccines as well as developing disaster drills and strategies for improved border controls.

[0010] Current treatment options for influenza include vaccination, and chemotherapy or chemoprophylaxis with anti-viral medications. Vaccination against influenza with an influenza vaccine is often recommended for high-risk groups, such as children and the elderly, or in people that have asthma, diabetes, or heart disease. However, it is possible to get vaccinated and still get influenza. The vaccine is reformulated each season for a few specific influenza strains but cannot possibly include all the strains actively infecting people in the world for that season. It may take six months for the manufacturers to formulate and produce the millions of doses required to deal with the seasonal epidemics; occasionally, a new or overlooked strain becomes prominent during that time and infects people although they have been vaccinated (as by the H3N2 Fujian flu in the 2003-2004 influenza season). It is also possible to get infected just before vaccination and get sick with the very strain that the vaccine is supposed to prevent, as the vaccine may take several weeks to become effective.

[0011] Further, the effectiveness of these influenza vaccines is variable. Due to the high mutation rate of the virus, a particular influenza vaccine usually confers protection for no more than a few years. A vaccine formulated for one year may be ineffective in the following year, since the influenza virus changes rapidly over time, and different strains become dominant.

[0012] Also, because of the absence of RNA proofreading enzymes, the RNA-dependent RNA polymerase of influenza vRNA makes a single nucleotide insertion error roughly every 10 thousand nucleotides, which is the approximate length of the influenza vRNA. Hence, nearly every newly-manufactured influenza virus is a mutant — antigenic drift. The separation of the genome into eight separate segments of vRNA allows mixing or reassortment of vRNAs if more than one viral line has infected a single cell. The resulting rapid change in viral genetics produces antigenic shifts and allows the virus to infect new host species and quickly overcome protective immunity.

[0013] Antiviral drugs can also be used to treat influenza, with neuraminidase inhibitors being particularly effective, but viruses can develop resistance to the standard antiviral drugs.

[0014] There is currently no approved therapy for the treatment of hospitalized patients with influenza infection and those infected with influenza that are at risk of influenza- related complications. However, these populations are often treated off-label with other influenza antivirals in which viral resistance has emerged, raising concerns.

[0015] Thus, there is still a need for drugs for treating influenza infections, such as for drugs with expanded treatment window, and/or reduced sensitivity to viral titer.

Compound (1) is a non-nucleotide inhibitor of the PB2 subunit of the influenza A virus polymerase complex and shows potent, antiviral activity against influenza A. Compound (1) has demonstrated antiviral activity in both in vitro and in vivo nonclinical models and has been shown to reduce peak viral load, area under the curve of viral shedding, and influenza-related clinical symptoms in a human challenge model and two Phase nb studies. Currently, Compound (1) is in Phase IP clinical development for the treatment of patients at risk of influenza-related complications, including hospitalized patients in the following studies: 63623872FLZ3001 (SAPPHIRE, NCT03376321) and 63623872FLZ3002 (DIAMOND, NCT03381196). BRIEF DESCRIPTION OF THE DRAWINGS [0016] FIG. 1 is an image of three different views of a surface graph plot of Synergy/Antagonism for the in-vitro Combination of Compound (1) and baloxavir. [0017] FIG. 2A is an image of heatmap plots presenting the raw values for 3 different plates.

[0018] FIG. 2B is an image of heatmap plots presenting the values for the 3 different plates normalized to % inhibition.

[0019] FIG. 3 is an image of graphs of the viral control (VC), cell control (CC), medium control (MC), and MC by plate and column.

[0020] FIG. 4A is an image of multiple plots presenting the raw data for the different doses of each compound.

[0021] FIG. 4B is an image of multiple plots presenting the raw data for the different doses of each compound. [0022] FIG. 5A is an image of multiple plots presenting the normalized data for the different doses of each compound.

[0023] FIG. 5B is an image of multiple plots presenting the normalized data for the different doses of each compound.

[0024] FIG. 6 is an image of two graphs showing the activity of Compound (1) and baloxavir for unrestricted parameters of monotherapy fits.

[0025] FIG. 7A is a contour plot of MaxR for unrestricted parameters of monotherapy fits.

[0026] FIG. 7B is a 3D plot of MaxR for unrestricted parameters of monotherapy fits. [0027] FIG. 8 is an image of two graphs showing the activity of Compound (1) and baloxavir for fixed maximal response and baseline of monotherapy fits.

[0028] FIG. 9A is a contour plot of MaxR for fixed maximal response and baseline of monotherapy fits.

[0029] FIG. 9B is a 3D plot of MaxR for fixed maximal response and baseline of monotherapy fits.

SUMMARY OF THE INVENTION

[0030] In one aspect, the invention includes a method for treating influenza virus infection in a subject or biological sample infected with influenza viruses comprising:

(0 administering to the subject or biological sample a therapeutically effective amount of Compound (1) or a pharmaceutically acceptable salt thereof wherein Compound

(1) has the structure:

(II) administering to the subject or biological sample a therapeutically effective amount of baloxavir or a pharmaceutically acceptable salt or prodrug thereof.

[0031] In some implementations, the administration of Compound (1) or a pharmaceutically acceptable salt thereof is prior to, concurrent with, or subsequent to the administration of baloxavir or a pharmaceutically acceptable salt or prodrug thereof.

[0032] Some implementations further comprise administering to the subject or biological sample a dosage of from about 100 mg to about 1600 mg of Compound (1) or a pharmaceutically acceptable salt thereof, wherein the dosage is administered 1 to 4 times per day during a treatment period.

[0033] In some implementations, the dosage is from about 200 mg to about 1200 mg of Compound (1) or a pharmaceutically acceptable salt thereof, and the dosage is administered 1 to 3 times per day during the treatment period. For example, the dosage is from about 550 mg to about 650 mg (e.g., about 600 mg), and the dosage is administered twice (i.e., BID) per day during the treatment period.

[0034] In some implementations, Compound (1) is administered to a subject in a unit dosage form, wherein the unit dosage form comprises from about 100 mg to about 800 mg of Compound (1) or a pharmaceutically acceptable salt thereof. In some implementations, the unit dosage form of Compound (1) is adapted for oral administration. In some implementations, Compound (1) in the unit dosage form is provided as a crystalline HC1 salt of Compound (1) · ½ H2O.

[0035] In some implementations, the unit dosage form is a tablet comprising from about 200 mg to about 700 mg of the crystalline HC1 salt of Compound (1) · ½ H2O. For example, the tablet comprises from about 250 mg to about 650 mg of the crystalline HC1 salt of Compound (1) · ½ H2O. In other examples, the tablet comprises from about

300 mg to about 375 mg of the crystalline HC1 salt of Compound (1) · ½ H2O. In some implementations, at least one tablet is administered to the subject 1 to 3 times per day during the treatment period.

[0036] In some implementations, the baloxavir or a pharmaceutically acceptable salt or prodrug thereof is administered to the sample or subject in a dosage of from about 10 mg to about

100 mg. For example, the dosage of baloxavir or a pharmaceutically acceptable salt or prodrug thereof is from about 30 mg to about 90 mg. In other examples, the dosage of baloxavir or a pharmaceutically acceptable salt or prodrug thereof is administered at least once during the treatment period.

[0037] In some implementations, baloxavir or a pharmaceutically acceptable salt or prodrug thereof is administered to a subject in a unit dosage form, wherein the unit dosage form comprises from about 10 mg to about 100 mg of baloxavir or a pharmaceutically acceptable salt or prodrug thereof. In some implementations, the unit dosage form of baloxavir is adapted for oral administration. And, in some implementations, baloxavir in the unit dosage form is provided as baloxavir marboxil. In some implementations, the unit dosage form is a tablet or capsule comprising from about 20 mg to about 90 mg of baloxavir marboxil. In some implementations, the tablet or capsule comprises from about 30 mg to about 50 mg of baloxavir marboxil. In some implementations, the tablet or capsule comprises from about 70 mg to about 90 mg of baloxavir marboxil. In some implementations, one tablet or capsule is administered to the subject at least once during the treatment period. [0038] In some implementations, the administration of Compound (1) or a pharmaceutically acceptable salt thereof in combination with the administration of baloxavir or a pharmaceutically acceptable salt or prodrug thereof results in a synergistic effect as compared with the administration of Compound (1) or a pharmaceutically acceptable salt thereof alone and the administration of baloxavir or a pharmaceutically acceptable salt or prodrug thereof alone.

[0039] In some implementations, the administration of Compound (1) or a pharmaceutically acceptable salt thereof in combination with the administration of baloxavir or a pharmaceutically acceptable salt or prodrug thereof results in a reduced or no development of virus resistance as compared with the administration of Compound (1) or a pharmaceutically acceptable salt thereof alone and the administration of baloxavir or a pharmaceutically acceptable salt or prodrug thereof alone.

[0040] In some implementations, the administration of Compound (1) or a pharmaceutically acceptable salt thereof in combination with the administration of oseltamivir or a pharmaceutically acceptable salt thereof results in a reduced or no development of virus resistance as compared with the administration of Compound (1) or a pharmaceutically acceptable salt thereof alone and the administration of oseltamivir or a pharmaceutically acceptable salt thereof alone.

[0041] In some implementations, the administration of oseltamivir or a pharmaceutically acceptable salt thereof in combination with the administration of baloxavir or a pharmaceutically acceptable salt or prodrug thereof results in a reduced or no development of virus resistance as compared with the administration of oseltamivir or a pharmaceutically acceptable salt thereof alone and the administration of baloxavir or a pharmaceutically acceptable salt or prodrug thereof alone. [0042] In some implementations, the administration of Compound (1) or a pharmaceutically acceptable salt thereof in combination with the administration of baloxavir or a pharmaceutically acceptable salt or prodrug thereof and in combination with the administration of oseltamivir or a pharmaceutically acceptable salt thereof results in a reduced or no development of virus resistance as compared with the administration of Compound (1) or a pharmaceutically acceptable salt thereof alone, the administration of baloxavir or a pharmaceutically acceptable salt or prodrug thereof alone and the administration of oseltamivir or a pharmaceutically acceptable salt thereof alone.

[0043] Another aspect of the present invention provides a pharmaceutical product comprising:

(a) a first unit dosage form comprising from about 200 mg to about 1600 mg of a crystalline HC1 salt of Compound (1) · ½ H2O, wherein Compound (1) has the structure:

(b) a second unit dosage form comprising from about 10 mg to about 100 mg of baloxavir marboxil, wherein the first unit dose and the second unit dose are adapted for oral administration.

[0044] In some embodiments, the second unit dosage form is provided as a single tablet or capsule.

[0045] In some implementations, the first unit dosage form is provided as one or more tablets, wherein each tablet comprises from about 200 mg to about 800 mg of the crystalline HC1 salt of Compound (1) · ½ H2O. For example, the first unit dosage form is provided as one or more tablets, wherein each tablet comprises from about 200 mg to about 400 mg of the crystalline HC1 salt of Compound (1) · ½ H2O.

[0046] Another aspect of the present invention provides a crystalline HC1 salt of Compound (1) · ½ H2O, wherein Compound (1) has the structure:

in combination with baloxavir or a pharmaceutically acceptable salt or prodrug thereof for use as a medicament for treating influenza virus infection.

[0047] In some embodiments, the crystalline HC1 salt of Compound (1) · ½ H2O is adapted for oral administration. For example, the crystalline HC1 salt of Compound (1) · ½ H2O is formulated as a tablet comprising from about 100 mg to about 700 mg of the crystalline HC1 salt. In other examples, the crystalline HC1 salt of Compound (1) · ½ H2O is formulated as a tablet comprising from about 200 mg to about 650 mg of the crystalline HC1 salt.

[0048] In some embodiments, baloxavir is adapted for oral administration. In some embodiments, baloxavir is baloxavir marboxil. In some embodiments, baloxavir is formulated as a tablet comprising from about 30 mg to about 50 mg or from about 70 mg to about 90 mg of baloxavir marboxil.

[0049] Another aspect of the present invention provides a method of inhibiting the replication of or reducing the number of influenza viruses in an in vitro biological sample, comprising:

(a) administering to the biological sample a therapeutically effective amount of Compound (1) or a pharmaceutically acceptable salt thereof wherein Compound (1) has the structure:

(b) administering to biological sample a therapeutically effective amount of baloxavir or a pharmaceutically acceptable salt or prodrug thereof, wherein Compound (1) is administered to give a concentration of from about 0.0018 mM to 0.4400 mM, and baloxavir is administered to give a concentration of from about 0.003 mM to 0.0750 mM. [0050] In some implementations, Compound (1) is administered prior to, concurrently with, or subsequent to the administration of baloxavir. [0051] Another aspect of the present invention provides a pharmaceutical composition comprising Compound (1) or a pharmaceutically acceptable salt thereof, wherein Compound (1) has the structure

baloxavir or a pharmaceutically acceptable salt or prodrug thereof.

[0052] In some embodiments, the composition comprises a single dosage form comprising Compound (1) or a pharmaceutically acceptable salt thereof and baloxavir or a pharmaceutically acceptable salt or prodrug thereof, or separate dosage forms, wherein the separate dosage forms comprise a dosage form of Compound (1) or a pharmaceutically acceptable salt thereof and a separate dosage form of baloxavir or a pharmaceutically acceptable salt or prodrug thereof.

[0053] In some embodiments, the composition comprises a single dosage form comprising Compound (1) or a pharmaceutically acceptable salt thereof and baloxavir or a pharmaceutically acceptable salt or prodrug thereof. For example, the single dosage form comprises a homogenous mixture of Compound (1) or a pharmaceutically acceptable salt thereof and baloxavir or a pharmaceutically acceptable salt or prodrug thereof. In other examples, the single dosage form comprises 2 or more distinct compositions, wherein each composition comprises Compound (1) or a pharmaceutically acceptable salt thereof or baloxavir or a pharmaceutically acceptable salt or prodrug thereof or a mixture of any combination thereof.

[0054] In some embodiments, the composition comprises separate dosage forms. In some embodiments, the separate dosage forms can be administered concurrently or consecutively. For example, the separate dosage forms are administered consecutively as a first dosage form and a second dosage form. In other examples, the second dosage form is administered from about one minute to about 48 hours after the first dose form.

[0055] In some embodiments, the second dosage form comprises Compound (1) or a pharmaceutically acceptable salt thereof, and the first dosage form comprises baloxavir or a pharmaceutically acceptable salt or prodrug thereof. [0056] In some embodiments, the second dosage form comprises baloxavir or a pharmaceutically acceptable salt or prodrug thereof, and the first dosage form comprises Compound (1) or a pharmaceutically acceptable salt thereof.

[0057] In some embodiments, Compound (1) or a pharmaceutically acceptable salt thereof is present in an amount sufficient to produce a concentration of from about 0.1 nM to about 0.1 mM of Compound (1) in the blood plasma of a patient after administration, and baloxavir or a pharmaceutically acceptable salt or prodrug thereof is present in an amount sufficient to produce a concentration of from about 0.1 nM to about 0.05 mM of baloxavir in the blood plasma of a patient after administration.

[0058] In some embodiments, the therapeutic effect is synergistic compared to the therapeutic effect observed for Compound (1) or a pharmaceutically acceptable salt thereof or baloxavir or a pharmaceutically acceptable salt or prodrug thereof alone.

[0059] In some embodiments, Compound (1) is provided in the composition as a crystalline HC1 salt of Compound (1) · ½ H2O. [0060] In some embodiments, baloxavir is provided in the composition as baloxavir marboxil.

[0061] In some embodiments, the dosage form of Compound (1) comprises from about 200 mg to about 800 mg of a crystalline HC1 salt of Compound (1) · ½ H2O. [0062] In some embodiments, the dosage form of baloxavir comprises from about 30 mg to about 90 mg of baloxavir marboxil.

[0063] In some embodiments, the dosage form of Compound (1) and the dosage form of baloxavir are adapted for oral administration.

[0064] In some embodiments, the dosage form of Compound (1) comprises one or more tablets, and the dosage form of baloxavir comprises a single tablet or capsule.

DETAILED DESCRIPTION OF THE INVENTION

[0065] I. DEFINITIONS

[0066] As used herein, “Compound (1)” refers to the compound having the structure

15

[0067] Without being bound by the following theory, it is believed that Compound (1) is a novel inhibitor of influenza virus replication that blocks the PB2 cap-snatching activity of the influenza viral polymerase complex. Compound (1) binds the cap-binding domain of the PB2 subunit with a dissociation constant of about 24 nM. In some embodiments, Compound (1) is present as the Compound (1) HC1 salt. In some other embodiments,

Compound (1) is present as the HC1 salt of Compound (1) hemihydrate. In still some other embodiments, Compound (1) is present as the HC1 salt of Compound (1) hemihydrate having Form A.

[0068] As used herein, the term “baloxavir” and “baloxavir acid” are used interchangeably and refer to the compound having the structure

[0069] As used herein, the term “baloxavir marboxyl” refers to a prodrug of baloxavir wherein the prodrug is a compound having the structure

[0070] As used herein, "oseltamivir" refers to an acetamido cyclohexene compound having the structure

Oseltamivir is a neuraminidase inhibitor that is sold (in phosphate salt form) under the trade name Tamiflu. [0071] As used herein, an “excipient” is an inactive ingredient in a pharmaceutical composition. Examples of excipients include fillers or diluents, wetting agents (e.g., surfactants), binders, glidants, lubricants, disintegrants, or the like.

[0072] As used herein, a “disintegrant agent” is an excipient that hydrates a pharmaceutical composition and aids in tablet dispersion. Examples of disintegrant agents include sodium croscarmellose, polyplasdone (i.e., cross-linked polyvinylpyrollidone), sodium starch glycolate, or any combination thereof.

[0073] As used herein, a “diluent” or “filler” is an excipient that adds bulkiness to a pharmaceutical composition. Examples of fillers include lactose, sorbitol, celluloses, calcium phosphates, starches, sugars (e.g., mannitol, sucrose, or the like) or any combination thereof.

[0074] As used herein, a “wetting agent” or a “surfactant” is an excipient that imparts pharmaceutical compositions with enhanced solubility and/or wetability. Examples of wetting agents include sodium lauryl sulfate (SLS), sodium stearyl fumarate (SSF), polyoxyethylene 20 sorbitan mono-oleate (e.g., Tween™), or any combination thereof. [0075] As used herein, a “binder” is an excipient that imparts a pharmaceutical composition with enhanced cohesion or tensile strength (e.g., hardness). Examples of binders include dibasic calcium phosphate, sucrose, com (maize) starch, microcrystalline cellulose, and modified cellulose (e.g., hydroxymethyl cellulose).

[0076] As used herein, a “glidant” is an excipient that imparts a pharmaceutical compositions with enhanced flow properties. Examples of glidants include colloidal silica and/or talc. [0077] As used herein, a “colorant” is an excipient that imparts a pharmaceutical composition with a desired color. Examples of colorants include commercially available pigments such as FD&C Blue # 1 Aluminum Lake, FD&C Blue #2, other FD&C Blue colors, titanium dioxide, iron oxide, and/or combinations thereof. Other colorants include commercially available pigments such as FD&C Green #3. [0078] As used herein, a “lubricant” is an excipient that is added to pharmaceutical compositions that are pressed into tablets. The lubricant aids in compaction of granules into tablets and ejection of a tablet of a pharmaceutical composition from a die press. Examples of lubricants include magnesium stearate, stearic acid (stearin), hydrogenated oil, sodium stearyl fumarate, or any combination thereof. [0079] For purposes of this invention, the chemical elements are identified in accordance with the Periodic Table of the Elements, CAS version, Handbook of Chemistry and Physics, 75th Ed. Additionally, general principles of organic chemistry are described in "Organic Chemistry", Thomas Sorrell, University Science Books, Sausolito: 1999, and "March’s Advanced Organic Chemistry", 5th Ed., Ed.: Smith, M.B. and March, J., John Wiley & Sons, New York: 2001, the entire contents of which are hereby incorporated by reference.

[0080] Unless otherwise indicated, structures depicted herein are also meant to include all isomeric (e.g., enantiomeric, diastereomeric, cis-trans, conformational, and rotational) forms of the structure. For example, the R and S configurations for each asymmetric center, (Z) and (E) double bond isomers, and (Z) and (E) conformational isomers are included in this invention, unless only one of the isomers is drawn specifically. As would be understood to one skilled in the art, a substituent can freely rotate around any rotatable bonds. [0081] Therefore, single stereochemical isomers as well as enantiomeric, diastereomeric, cis/trans, conformational, and rotational mixtures of the present compounds are within the scope of the invention.

[0082] Unless otherwise indicated, all tautomeric forms of the compounds of the invention are within the scope of the invention.

[0083] Additionally, unless otherwise indicated, structures depicted herein are also meant to include compounds that differ only in the presence of one or more isotopically enriched atoms. For example, compounds having the present structures except for the replacement of hydrogen by deuterium or tritium, or the replacement of a carbon by a ¹³C- or ¹⁴C-enriched carbon are within the scope of this invention. Such compounds are useful, for example, as analytical tools or probes in biological assays. Such compounds, especially deuterium (D) analogs, can also be therapeutically useful.

[0084] As used herein, the term “synergy” or “synergistic” refers to a combination of a compound of the invention and another therapy (e.g., a prophylactic or therapeutic agent), which is more effective than the additive effects of the therapies. A synergistic effect of a combination of therapies (e.g., a combination of prophylactic or therapeutic agents) can permit the use of lower dosages of one or more of the therapies and/or less frequent administration of said therapies to a subject. The ability to utilize lower dosages of a therapy (e.g., a prophylactic or therapeutic agent) and/or to administer said therapy less frequently can reduce the toxicity associated with the administration of said therapy to a subject without reducing the efficacy of said therapy in the prevention, management or treatment of a disorder. In addition, a synergistic effect can result in improved efficacy of agents in the prevention, management or treatment of a disorder. Finally, a synergistic effect of a combination of therapies (e.g., a combination of prophylactic or therapeutic agents) may avoid or reduce adverse or unwanted side effects associated with the use of either therapy alone.

[0085] As used herein, the phrase “therapeutically effective amount” refers to an amount sufficient to elicit the desired biological response. In the present invention the desired biological response is to inhibit the replication of influenza virus, to reduce the amount of influenza viruses or to reduce or ameliorate the severity, duration, progression, or onset of an influenza virus infection, prevent the advancement of an influenza viruses infection, prevent the recurrence, development, onset or progression of a symptom associated with an influenza virus infection, or enhance or improve the prophylactic or therapeutic effect(s) of another therapy used against influenza infections. The precise amount of compound administered to a subject will depend on the mode of administration, the type and severity of the infection and on the characteristics of the subject, such as general health, age, sex, body weight and tolerance to drugs. The skilled artisan will be able to determine appropriate dosages depending on these and other factors. In cases where no amount is expressly noted, an effective amount should be assumed. For example, the compounds disclosed herein can be administered to a subject in a dosage range from between approximately 0.01 to 100 mg/kg body weight/day for therapeutic or prophylactic treatment. [0086] As used herein, a “unit dosage form” refers to one or more pharmaceutical drug products in the form in which it is marketed for use, with a specific mixture of active ingredients and inactive components (excipients), in a particular configuration (such as a capsule shell, for example), and apportioned into a particular dose.

[0087] As used herein, a “subject” refers to a mammal (e.g., human, monkey, dog, mouse, rat, and the like).

[0088] P. EMBODIMENTS OF THE INVENTION

[0089] In one aspect, the invention includes a method for treating influenza virus infection in a subject or biological sample infected with influenza viruses comprising:

(0 administering to the subject or biological sample a therapeutically effective amount of Compound (1) or a pharmaceutically acceptable salt thereof wherein Compound (1) has the structure:

, and

[0090] In one embodiment, the administration of Compound (1) or a pharmaceutically acceptable salt thereof is prior to, concurrent with, or subsequent to the administration of baloxavir or a pharmaceutically acceptable salt or prodrug thereof.

[0091] In one embodiment, the method further comprises administering to the subject or biological sample a dosage of from about 100 mg to about 1600 mg (e.g., from about

200 mg to about 1200 mg, from about 250 mg to about 1000 mg, or from about 300 mg to about 900 mg) of Compound (1) or a pharmaceutically acceptable salt thereof, wherein the dosage is administered 1 to 4 times per day during a treatment period. In some examples, the dosage is from about 200 mg to about 1200 mg of Compound (1) or a pharmaceutically acceptable salt thereof, and the dosage is administered 1 to 3 times per day during the treatment period. And, in some embodiments, the dosage is from about 550 mg to about 650 mg (e.g., about 600 mg) of Compound (1) or a pharmaceutically acceptable salt thereof, and the dosage is administered twice (i.e., BID) per day during the treatment period. In other embodiments, the dosage is from about 200 mg to about 400 mg (e.g., from about 250 mg to about 375 mg or from about 275 mg to about 350 mg) of Compound (1) or a pharmaceutically acceptable salt thereof, and the dosage is administered 1-3 times per day during the treatment period.

[0092] In some embodiments, Compound (1) is administered to a subject in a unit dosage form, wherein the unit dosage form comprises from about 100 mg to about 800 mg (e.g., from about 100 mg to about 300 mg, from about 150 mg to about 300 mg, or from about 150 mg to about 250 mg) of Compound (1) or a pharmaceutically acceptable salt thereof. And, in some examples, the unit dosage form of Compound (1) is adapted for oral administration.

[0093] In some embodiments, Compound (1) in the unit dosage form is provided as a crystalline HC1 salt of Compound (1) · ½ H2O (i.e., the HC1 salt of Compound (1) hemihydrate). [0094] In some embodiments, the unit dosage form is a tablet comprising from about

100 mg to about 700 mg (e.g., from about 200 mg to about 700 mg, from about 100 mg to about 300 mg, from about 150 mg to about 300 mg, or from about 150 mg to about 250 mg) of the crystalline HC1 salt of Compound (1) · ½ H2O. For example, the tablet comprises from about 250 mg to about 650 mg of the crystalline HC1 salt of Compound (1) · ½ H2O. In other examples, the tablet comprises from about 300 mg to about 375 mg of the crystalline HC1 salt of Compound (1) · ½ H2O.

[0095] In some embodiments, at least one tablet is administered to the subject 1 to 3 times per day during the treatment period.

[0096] In some embodiment, the baloxavir or a pharmaceutically acceptable salt or prodrug thereof is administered to the sample or subject in a dosage of from about 10 mg to about 100 mg. For example, the dosage of baloxavir or a pharmaceutically acceptable salt or prodrug thereof is from about 20 mg to about 90 mg (e.g., from about 25 mg to about 75 mg, from about 30 mg to about 60 mg, or from about 30 mg to about 50 mg), and the dosage is administered 1-3 times per day during the treatment period. In other examples, the dosage of baloxavir or a pharmaceutically acceptable salt or prodrug thereof is from about 25 mg to about 50 mg (e.g., from about 35 mg to about 45 mg), and the dosage is administered 1-3 times per day during the treatment period.

[0097] In one embodiment, the dosage of baloxavir or a pharmaceutically acceptable salt or prodrug thereof is administered at least once (e.g., from 1 to 20 times, from 2 to 16 times, or from 2 to 10 times) during the treatment period.

[0098] In one embodiment, baloxavir or a pharmaceutically acceptable salt or prodrug thereof is administered to a subject in a unit dosage form, wherein the unit dosage form comprises from about 10 mg to about 100 mg of baloxavir or a pharmaceutically acceptable salt or prodrug thereof. In some examples, the unit dosage form of baloxavir is adapted for oral administration. And in some examples, baloxavir in the unit dosage form is provided as a prodrug (e.g., baloxavir marboxil).

[0099] In some embodiments, the unit dosage form is a tablet or capsule comprising from about 20 mg to about 90 mg of baloxavir marboxil. For example, the tablet or capsule comprises from about 30 mg to about SO mg of baloxavir marboxil. In other examples, the tablet or capsule comprises from about 70 mg to about 90 mg of baloxavir marboxil. And, in some examples, one tablet or capsule is administered to the subject at least once (e.g., 1-20 times, 2-18 times, or 4-12 times) during the treatment period. [00100] In some embodiment, oseltamivir or a pharmaceutically acceptable salt thereof is administered to the sample or subject in a dosage of from about 50 mg to about 100 mg. For example, the dosage of oseltamivir or a pharmaceutically acceptable salt thereof is from about 40 mg to about 90 mg (e.g., from about 50 mg to about 80 mg, from about 55 mg to about 75 mg, or from about 60 mg to about 70 mg), and the dosage is administered 1-3 times per day during the treatment period. In other examples, oseltamivir is administered in a dosage regimen according to its label. In some specific embodiments, it is administered 75 mg twice a day, or 150 mg once a day.

[00101] In one embodiment, the dosage of oseltamivir or a pharmaceutically acceptable salt thereof is administered at least once (e.g., from 1 to 20 times, from 2 to 16 times, or from 2 to 10 times) during the treatment period.

[00102] In one embodiment, oseltamivir or a pharmaceutically acceptable salt thereof is administered to a subject in a unit dosage form, wherein the unit dosage form comprises from about 50 mg to about 100 mg of oseltamivir or a pharmaceutically acceptable salt thereof. In some examples, the unit dosage form of oseltamivir is adapted for oral administration.

[00103] In some embodiments, the administration of Compound (1) or a pharmaceutically acceptable salt thereof in combination with the administration of baloxavir or a pharmaceutically acceptable salt or prodrug thereof results in a synergistic effect as compared with the administration of Compound (1) or a pharmaceutically acceptable salt thereof alone and the administration of baloxavir or a pharmaceutically acceptable salt or prodrug thereof alone.

[00104] In some embodiments, the administration of Compound (1) or a pharmaceutically acceptable salt thereof in combination with the administration of baloxavir or a pharmaceutically acceptable salt or prodrug thereof results in a reduced or no development of virus resistance as compared with the administration of Compound (1) or a pharmaceutically acceptable salt thereof alone and the administration of baloxavir or a pharmaceutically acceptable salt or prodrug thereof alone.

[00105] In some embodiments, the administration of Compound (1) or a pharmaceutically acceptable salt thereof in combination with the administration of oseltamivir or a pharmaceutically acceptable salt thereof results in a reduced or no development of virus resistance as compared with the administration of Compound (1) or a pharmaceutically acceptable salt thereof alone and the administration of oseltamivir or a pharmaceutically acceptable salt thereof alone.

[00106] In some embodiments, the administration of oseltamivir or a pharmaceutically acceptable salt thereof in combination with the administration of baloxavir or a pharmaceutically acceptable salt or prodrug thereof results in a reduced or no development of virus resistance as compared with the administration of oseltamivir or a pharmaceutically acceptable salt thereof alone and the administration of baloxavir or a pharmaceutically acceptable salt or prodrug thereof alone.

[00107] In some embodiments, the administration of Compound (1) or a pharmaceutically acceptable salt thereof in combination with the administration of baloxavir or a pharmaceutically acceptable salt or prodrug thereof and in combination with the administration of oseltamivir or a pharmaceutically acceptable salt thereof results in a reduced or no development of virus resistance as compared with the administration of Compound (1) or a pharmaceutically acceptable salt thereof alone, the administration of baloxavir or a pharmaceutically acceptable salt or prodrug thereof alone and the administration of oseltamivir or a pharmaceutically acceptable salt thereof alone. [00108] Another aspect of the present invention provides a pharmaceutical product comprising:

; and

(b) a second unit dosage form comprising from about 10 mg to about 100 mg of baloxavir marboxil, wherein the first unit dose and the second unit dose are adapted for oral administration. [00109] In some embodiments, the second unit dosage form is provided as a single tablet or capsule. In some embodiments, the second unit dosage form is provided as two tablets or capsules.

[00110] In some embodiments, the first unit dosage form is provided as one or more tablets, wherein each tablet comprises from about 200 mg to about 800 mg of the crystalline HC1 salt of Compound (1) · ½ H2O.

[00111] In one embodiment, the first unit dosage form is provided as one or more tablets, wherein each tablet comprises from about 200 mg to about 400 mg of the crystalline HC1 salt of Compound (1) · ½ H2O. [0100] In one aspect, the invention includes a crystalline HC1 salt of Compound (1) · ½

H2O, wherein Compound (1) has the structure:

in combination with baloxavir or a pharmaceutically acceptable salt or prodrug thereof for use as a medicament for treating influenza virus infection. [0101] In some embodiments, the crystalline HC1 salt is adapted for oral administration.

[0102] In some embodiments, the crystalline HC1 salt is formulated as a tablet comprising from about 100 mg to about 700 mg of the crystalline HC1 salt. For example, the crystalline HC1 salt is formulated as a tablet comprising from about 200 mg to about 650 mg of the crystalline HC1 salt. [0103] In some embodiments, baloxavir is adapted for oral administration.

[0104] In some embodiments, baloxavir is provided as baloxavir marboxil.

[0105] In some embodiments, baloxavir is formulated as a tablet comprising from about 30 mg to about 50 mg or from about 70 mg to about 90 mg of baloxavir marboxil.

[0106] Another aspect of the present invention provides a method of inhibiting the replication of or reducing the number of influenza viruses in an in vitro biological sample, comprising:

, and

(b) administering to biological sample a therapeutically effective amount of baloxavir or a pharmaceutically acceptable salt or prodrug thereof, wherein Compound (1) is to give a concentration of from about 0.0018 mM to 0.4400 mM, and baloxavir is administered to give a concentration of from about 0.003 mM to 0.0750 mM.

[0107] In some embodiments, Compoumd (1) is administered prior to, concurrently with, or subsequent to the administration of baloxavir.

[0108] Another aspect of the invention provides a pharmaceutical composition comprising Compound (1) or a pharmaceutically acceptable salt thereof, wherein Compound (1) has the structure

baloxavir or a pharmaceutically acceptable salt or prodrug thereof.

[0109] In some embodiments, the composition comprises

(a) a single dosage form comprising Compound (1) or a pharmaceutically acceptable salt thereof and baloxavir or a pharmaceutically acceptable salt or prodrug thereof, or

(b) separate dosage forms, wherein the separate dosage forms comprise a dosage form of Compound (1) or a pharmaceutically acceptable salt thereof and a separate dosage form of baloxavir or a pharmaceutically acceptable salt or prodrug thereof.

[0110] In some embodiments, the composition comprises a single dosage form comprising Compound (1) or a pharmaceutically acceptable salt thereof and baloxavir or a pharmaceutically acceptable salt or prodrug thereof.

[0111] In some embodiments, the single dosage form comprises a homogenous mixture of Compound (1) or a pharmaceutically acceptable salt thereof and baloxavir or a pharmaceutically acceptable salt or prodrug thereof.

[0112] In some embodiments, the single dosage form comprises 2 or more distinct compositions, wherein each composition comprises Compound (1) or a pharmaceutically acceptable salt thereof or baloxavir or a pharmaceutically acceptable salt or prodrug thereof or a mixture of any combination thereof.

[0113] In some embodiments, the composition comprises separate dosage forms. In these embodiments, the separate dosage forms can be administered concurrently or consecutively.

[0114] In some embodiments, the separate dosage forms are administered consecutively as a first dosage form and a second dosage form.

[0115] In some embodiments, the second dosage form is administered from about one minute to about 48 hours after the first dose form.

[0116] In some embodiments, the second dosage form comprises Compound (1) or a pharmaceutically acceptable salt thereof, and the first dosage form comprises baloxavir or a pharmaceutically acceptable salt or prodrug thereof.

[0117] In other embodiments, the second dosage form comprises baloxavir or a pharmaceutically acceptable salt or prodrug thereof, and the first dosage form comprises Compound (1) or a pharmaceutically acceptable salt thereof. [0118] In some embodiments, Compound (1) or a pharmaceutically acceptable salt thereof is present in an amount sufficient to produce a concentration of from about 0.1 nM to about 0.1 mM of Compound (1) in the blood plasma of a patient after administration, and baloxavir or a pharmaceutically acceptable salt or prodrug thereof is present in an amount sufficient to produce a concentration of from about 0.1 nM to about 0.05 mM of baloxavir in the blood plasma of a patient after administration.

[0119] In some embodiments, the therapeutic effect is synergistic compared to the therapeutic effect observed for Compound (1) or a pharmaceutically acceptable salt thereof or baloxavir or a pharmaceutically acceptable salt or prodrug thereof alone.

[0120] In some embodiments, Compound (1) is provided in the composition as a crystalline HC1 salt of Compound (1) · ½ H2O.

[0121] In some embodiments, baloxavir is provided in the composition as baloxavir marboxil.

[0122] In some embodiments, the dosage form of Compound (1) comprises from about 200 mg to about 800 mg of a crystalline HC1 salt of Compound (1) · ½ H2O. [0123] In some embodiments, the dosage form of baloxavir comprises from about 30 mg to about 90 mg of baloxavir marboxil.

[0124] In some embodiments, the dosage form of Compound (1) and the dosage form of baloxavir are adapted for oral administration.

[0125] In some embodiments, the dosage form of Compound (1) comprises one or more tablets, and the dosage form of baloxavir comprises a single tablet or capsule.

[0126] III. PHARMACEUTICAL COMPOSITIONS [0127] The compounds described herein can be formulated into pharmaceutical compositions that further comprise a pharmaceutically acceptable carrier, diluent, adjuvant or vehicle. In one embodiment, the present invention relates to a pharmaceutical composition comprising a compound of the invention described above, and a pharmaceutically acceptable carrier, diluent, adjuvant or vehicle. In one embodiment, the present invention is a pharmaceutical composition comprising an effective amount of a compound of the present invention or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier, diluent, adjuvant or vehicle. Pharmaceutically acceptable carriers include, for example, pharmaceutical diluents, excipients or carriers suitably selected with respect to the intended form of administration, and consistent with conventional pharmaceutical practices. [0128] Further to the definition above, an “effective amount” includes a “therapeutically effective amount” and a “prophylactically effective amount”. The term “therapeutically effective amount” refers to an amount effective in treating and/or ameliorating an influenza virus infection in a patient infected with influenza. The term “prophylactically effective amount” refers to an amount effective in preventing and/or substantially lessening the chances or the size of influenza virus infection outbreak.

[0129] A pharmaceutically acceptable carrier may contain inert ingredients which do not unduly inhibit the biological activity of the compounds. The pharmaceutically acceptable carriers should be biocompatible, e.g., non-toxic, non-inflammatory, non-immunogenic or devoid of other undesired reactions or side-effects upon the administration to a subject. Standard pharmaceutical formulation techniques can be employed.

[0130] The pharmaceutically acceptable carrier, adjuvant, or vehicle, as used herein, includes any and all solvents, diluents, or other liquid vehicle, dispersion or suspension aids, surface active agents, isotonic agents, thickening or emulsifying agents, preservatives, solid binders, lubricants and the like, as suited to the particular dosage form desired. Remington's Pharmaceutical Sciences, Sixteenth Edition, E. W. Martin (Mack Publishing Co., Easton, Pa., 1980) discloses various carriers used in formulating pharmaceutically acceptable compositions and known techniques for the preparation thereof. Except insofar as any conventional carrier medium is incompatible with the compounds described herein, such as by producing any undesirable biological effect or otherwise interacting in a deleterious manner with any other component(s) of the pharmaceutically acceptable composition, its use is contemplated to be within the scope of this invention. As used herein, the phrase “side effects” encompasses unwanted and adverse effects of a therapy (e.g., a prophylactic or therapeutic agent). Side effects are always unwanted, but unwanted effects are not necessarily adverse. An adverse effect from a therapy (e.g., prophylactic or therapeutic agent) might be harmful or uncomfortable or risky. Side effects include, but are not limited to fever, chills, lethargy, gastrointestinal toxicities (including gastric and intestinal ulcerations and erosions), nausea, vomiting, neurotoxicities, nephrotoxicities, renal toxicities (including such conditions as papillary necrosis and chronic interstitial nephritis), hepatic toxicities (including elevated serum liver enzyme levels), myelotoxicities (including leukopenia, myelosuppression, thrombocytopenia and anemia), dry mouth, metallic taste, prolongation of gestation, weakness, somnolence, pain (including muscle pain, bone pain and headache), hair loss, asthenia, dizziness, extra-pyramidal symptoms, akathisia, cardiovascular disturbances and sexual dysfunction.

[0131] Some examples of materials which can serve as pharmaceutically acceptable carriers include, but are not limited to, ion exchangers, alumina, aluminum stearate, lecithin, serum proteins (such as human serum albumin), buffer substances (such as twin 80, phosphates, glycine, sorbic acid, or potassium sorbate), partial glyceride mixtures of saturated vegetable fatty acids, water, salts or electrolytes (such as protamine sulfate, disodium hydrogen phosphate, potassium hydrogen phosphate, sodium chloride, or zinc salts), colloidal silica, magnesium trisilicate, polyvinyl pyrrolidone, polyacrylates, waxes, polyethylene-polyoxypropylene-block polymers, methylcellulose, hydroxypropyl methylcellulose, wool fat, sugars such as lactose, glucose and sucrose; starches such as com starch and potato starch; cellulose and its derivatives such as sodium carboxymethyl cellulose, ethyl cellulose and cellulose acetate; powdered tragacanth; malt; gelatin; talc; excipients such as cocoa butter and suppository waxes; oils such as peanut oil, cottonseed oil; safflower oil; sesame oil; olive oil; com oil and soybean oil; glycols; such a propylene glycol or polyethylene glycol; esters such as ethyl oleate and ethyl laurate; agar; buffering agents such as magnesium hydroxide and aluminum hydroxide; alginic acid; pyrogen-free water; isotonic saline; Ringer's solution; ethyl alcohol, and phosphate buffer solutions, as well as other non-toxic compatible lubricants such as sodium lauryl sulfate and magnesium stearate, as well as coloring agents, releasing agents, coating agents, sweetening, flavoring and perfuming agents, preservatives and antioxidants can also be present in the composition, according to the judgment of the formulator.

[0132] IV. ADMINISTRATION METHODS [0133] The compounds and pharmaceutically acceptable compositions described above can be administered to humans and other animals orally, rectally, parenterally, intracistemally, intravaginally, intraperitoneally, topically (as by powders, ointments, or drops), buccally, as an oral or nasal spray, or the like, depending on the severity of the infection being treated. [0134] Liquid dosage forms for oral administration include, but are not limited to, pharmaceutically acceptable emulsions, microemulsions, solutions, suspensions, symps and elixirs. In addition to the active compounds, the liquid dosage forms may contain inert diluents commonly used in the art such as, for example, water or other solvents, solubilizing agents and emulsifiers such as ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butylene glycol, dimethylformamide, oils (in particular, cottonseed, groundnut, com, germ, olive, castor, and sesame oils), glycerol, tetrahydrofurfuryl alcohol, polyethylene glycols and fatty acid esters of sorbitan, and mixtures thereof. Besides inert diluents, the oral compositions can also include adjuvants such as wetting agents, emulsifying and suspending agents, sweetening, flavoring, and perfuming agents.

[0135] Injectable preparations, for example, sterile injectable aqueous or oleaginous suspensions may be formulated according to the known art using suitable dispersing or wetting agents and suspending agents. The sterile injectable preparation may also be a sterile injectable solution, suspension or emulsion in a nontoxic parenterally acceptable diluent or solvent, for example, as a solution in 1,3-butanediol. Among the acceptable vehicles and solvents that may be employed are water, Ringer's solution, U.S.P. and isotonic sodium chloride solution. In addition, sterile, fixed oils are conventionally employed as a solvent or suspending medium. For this purpose any bland fixed oil can be employed including synthetic mono- or diglycerides. In addition, fatty acids such as oleic acid are used in the preparation of injectables.

[0136] The injectable formulations can be sterilized, for example, by filtration through a bacterial-retaining filter, or by incorporating sterilizing agents in the form of sterile solid compositions which can be dissolved or dispersed in sterile water or other sterile injectable medium prior to use.

[0137] In order to prolong the effect of a compound described herein, it is often desirable to slow the absorption of the compound from subcutaneous or intramuscular injection.

This may be accomplished by the use of a liquid suspension of crystalline or amorphous material with poor water solubility. The rate of absorption of the compound then depends upon its rate of dissolution that, in turn, may depend upon crystal size and crystalline form. Alternatively, delayed absorption of a parenterally administered compound form is accomplished by dissolving or suspending the compound in an oil vehicle. Injectable depot forms are made by forming microencapsule matrices of the compound in biodegradable polymers such as polylactide-polyglycolide. Depending upon the ratio of compound to polymer and the nature of the particular polymer employed, the rate of compound release can be controlled. Examples of other biodegradable polymers include poly(orthoesters) and poly(anhydrides). Depot injectable formulations are also prepared by entrapping the compound in liposomes or microemulsions that are compatible with body tissues.

[0138] Compositions for rectal or vaginal administration are specifically suppositories which can be prepared by mixing the compounds described herein with suitable non- irritating excipients or carriers such as cocoa butter, polyethylene glycol or a suppository wax which are solid at ambient temperature but liquid at body temperature and therefore melt in the rectum or vaginal cavity and release the active compound.

[0139] Solid dosage forms for oral administration include capsules, tablets, pills, powders, and granules. In such solid dosage forms, the active compound is mixed with at least one inert, pharmaceutically acceptable excipient or carrier such as sodium citrate or dicalcium phosphate and/or a) fillers or extenders such as starches, lactose, sucrose, glucose, mannitol, and silicic acid, b) binders such as, for example, carboxymethylcellulose, alginates, gelatin, polyvinylpyrrolidinone, sucrose, and acacia, c) humectants such as glycerol, d) disintegrating agents such as agar-agar, calcium carbonate, potato or tapioca starch, alginic acid, certain silicates, and sodium carbonate, e) solution retarding agents such as paraffin, f) absorption accelerators such as quaternary ammonium compounds, g) wetting agents such as, for example, cetyl alcohol and glycerol monostearate, h) absorbents such as kaolin and bentonite clay, and i) lubricants such as talc, calcium stearate, magnesium stearate, solid polyethylene glycols, sodium lauryl sulfate, and mixtures thereof. In the case of capsules, tablets and pills, the dosage form may also comprise buffering agents.

[0140] Solid compositions of a similar type may also be employed as fillers in soft and hard-filled gelatin capsules using such excipients as lactose or milk sugar as well as high molecular weight polyethylene glycols and the like. The solid dosage forms of tablets, dragees, capsules, pills, and granules can be prepared with coatings and shells such as enteric coatings and other coatings well known in the pharmaceutical formulating art.

They may optionally contain opacifying agents and can also be of a composition that they release the active ingredient(s) only, or preferentially, in a certain part of the intestinal tract, optionally, in a delayed manner. Examples of embedding compositions that can be used include polymeric substances and waxes. Solid compositions of a similar type may also be employed as fillers in soft and hard-filled gelatin capsules using such excipients as lactose or milk sugar as well as high molecular weight polethylene glycols and the like. [0141] The active compounds can also be in microencapsulated form with one or more excipients as noted above. The solid dosage forms of tablets, dragees, capsules, pills, and granules can be prepared with coatings and shells such as enteric coatings, release controlling coatings and other coatings well known in the pharmaceutical formulating art. In such solid dosage forms the active compound may be admixed with at least one inert diluent such as sucrose, lactose or starch. Such dosage forms may also comprise, as is normal practice, additional substances other than inert diluents, e.g., tableting lubricants and other tableting aids such a magnesium stearate and microcrystalline cellulose. In the case of capsules, tablets and pills, the dosage forms may also comprise buffering agents. They may optionally contain opacifying agents and can also be of a composition that they release the active ingredient(s) only, or preferentially, in a certain part of the intestinal tract, optionally, in a delayed manner. Examples of embedding compositions that can be used include polymeric substances and waxes.

[0142] Dosage forms for topical or transdermal administration of a compound described herein include ointments, pastes, creams, lotions, gels, powders, solutions, sprays, inhalants or patches. The active component is admixed under sterile conditions with a pharmaceutically acceptable carrier and any needed preservatives or buffers as may be required. Ophthalmic formulation, eardrops, and eye drops are also contemplated as being within the scope of this invention. Additionally, the present invention contemplates the use of transdermal patches, which have the added advantage of providing controlled delivery of a compound to the body. Such dosage forms can be made by dissolving or dispensing the compound in the proper medium. Absorption enhancers can also be used to increase the flux of the compound across the skin. The rate can be controlled by either providing a rate controlling membrane or by dispersing the compound in a polymer matrix or gel. [0143] The compositions described herein may be administered orally, parenterally, by inhalation spray, topically, rectally, nasally, buccally, vaginally or via an implanted reservoir. The term "parenteral" as used herein includes, but is not limited to, subcutaneous, intravenous, intramuscular, intra-articular, intra-synovial, intrastemal, intrathecal, intrahepatic, intralesional and intracranial injection or infusion techniques. Specifically, the compositions are administered orally, intraperitoneally or intravenously. [0144] Sterile injectable forms of the compositions described herein may be aqueous or oleaginous suspension. These suspensions may be formulated according to techniques known in the art using suitable dispersing or wetting agents and suspending agents. The sterile injectable preparation may also be a sterile injectable solution or suspension in a non-toxic parenterally-acceptable diluent or solvent, for example as a solution in

1,3-butanediol. Among the acceptable vehicles and solvents that may be employed are water, Ringer's solution and isotonic sodium chloride solution. In addition, sterile, fixed oils are conventionally employed as a solvent or suspending medium. For this purpose, any bland fixed oil may be employed including synthetic mono- or di-glycerides. Fatty acids, such as oleic acid and its glyceride derivatives are useful in the preparation of injectables, as are natural pharmaceutically-acceptable oils, such as olive oil or castor oil, especially in their polyoxyethylated versions. These oil solutions or suspensions may also contain a long-chain alcohol diluent or dispersant, such as carboxymethyl cellulose or similar dispersing agents which are commonly used in the formulation of pharmaceutically acceptable dosage forms including emulsions and suspensions. Other commonly used surfactants, such as Tweens, Spans and other emulsifying agents or bioavailability enhancers which are commonly used in the manufacture of pharmaceutically acceptable solid, liquid, or other dosage forms may also be used for the purposes of formulation. [0145] The pharmaceutical compositions described herein may be orally administered in any orally acceptable dosage form including, but not limited to, capsules, tablets, aqueous suspensions or solutions. In the case of tablets for oral use, carriers commonly used include, but are not limited to, lactose and com starch. Lubricating agents, such as magnesium stearate, are also typically added. For oral administration in a capsule form, useful diluents include lactose and dried cornstarch. When aqueous suspensions are required for oral use, the active ingredient is combined with emulsifying and suspending agents. If desired, certain sweetening, flavoring or coloring agents may also be added. [0146] Alteratively, the pharmaceutical compositions described herein may be administered in the form of suppositories for rectal administration. These can be prepared by mixing the agent with a suitable non-irritating excipient which is solid at room temperature but liquid at rectal temperature and therefore will melt in the rectum to release the drug. Such materials include, but are not limited to, cocoa butter, beeswax and polyethylene glycols. [0147] The pharmaceutical compositions described herein may also be administered topically, especially when the target of treatment includes areas or organs readily accessible by topical application, including diseases of the eye, the skin, or the lower intestinal tract. Suitable topical formulations are readily prepared for each of these areas or organs. [0148] Topical application for the lower intestinal tract can be effected in a rectal suppository formulation (see above) or in a suitable enema formulation. Topically- transdermal patches may also be used.

[0149] For topical applications, the pharmaceutical compositions may be formulated in a suitable ointment containing the active component suspended or dissolved in one or more carriers. Carriers for topical administration of the compounds of this invention include, but are not limited to, mineral oil, liquid petrolatum, white petrolatum, propylene glycol, polyoxyethylene, polyoxypropylene compound, emulsifying wax and water.

Alternatively, the pharmaceutical compositions can be formulated in a suitable lotion or cream containing the active components suspended or dissolved in one or more pharmaceutically acceptable carriers. Suitable carriers include, but are not limited to, mineral oil, sorbitan monostearate, polysorbate 60, cetyl esters wax, cetearyl alcohol,

2 octyldodecanol, benzyl alcohol and water.

[0150] For ophthalmic use, the pharmaceutical compositions may be formulated as micronized suspensions in isotonic, pH adjusted sterile saline, or, specifically, as solutions in isotonic, pH adjusted sterile saline, either with or without a preservative such as benzylalkonium chloride. Alternatively, for ophthalmic uses, the pharmaceutical compositions may be formulated in an ointment such as petrolatum.

[0151] The pharmaceutical compositions may also be administered by nasal aerosol or inhalation. Such compositions are prepared according to techniques well-known in the art of pharmaceutical formulation and may be prepared as solutions in saline, employing benzyl alcohol or other suitable preservatives, absorption promoters to enhance bioavailability, fluorocarbons, and/or other conventional solubilizing or dispersing agents. [0152] The compounds for use in the methods of the invention can be formulated in unit dosage form. The term “unit dosage form” refers to physically discrete units suitable as unitary dosage for subjects undergoing treatment, with each unit containing a predetermined quantity of active material calculated to produce the desired therapeutic effect, optionally in association with a suitable pharmaceutical carrier. The unit dosage form can be for a single daily dose or one of multiple daily doses (e.g., 1 to 4 or more times per day). When multiple daily doses are used, the unit dosage form can be the same or different for each dose.

[0153] V. EXAMPLES [0154] Procedures for the synthetic process for producing Compound (1) and the procedure for creating the polymorphic form Compound (1) HC1 salt Form A from Compound (1) are provided in U.S. Patent Nos. 10,023,569 and 9,771,361, respectively. The entire contents of both US patents are hereby incorporated by reference in their entirety. [0155] Example 1: In-vitro Combination Experiments with Compound (1) and Other

Influenza Antiviral Drugs

[0156] The in-vitro antiviral effects of the combination of Compound (1) with baloxavir are investigated.

[0157] Methods (In-vitro Analyses^') [0158] Antiviral activity (synergistic, additive, or antagonistic) of Compound (1) with other antivirals in vitro was evaluated in a 3-day cytopathic effect assay using Madin- Darby canine kidney cells infected with the H1N1 strain A/Puerto Rico/8/34 (or the H3N2 strain A/Port Chalmers/1/73 for combinations with AL794) at a multiplicity of infection of 0.01. [0159] Read-out of cell viability was performed using CellTiter-Glo® or ATPlite™ luminescence measurements.

[0160] Analysis of the in-vitro results was performed using the Biochemically Intuitive Generalized Loewe (BIGL) methodology was used for the combination of Compound (1) with baloxavir. [0161] The BIGL method is an extension of the Loewe additivity model, and it is based on four-parameter log-logistic curves (Hill equation) fitted to the monotherapy data. The predicted response surface (indicating additive effects) is calculated based on the Generalized Loewe equation, and statistical tests are performed to assess deviations from the predicted surface named ‘Residuals’ (R). Both the mean (overall) and point-by-point R was calculated using R-Package.

[0162] Results

[0163] In the analysis of the Compound (1) and baloxavir combination, the BIGL method resulted in an overall synergistic effect, with an overall R of 18.97 which was statistically significant (p<2-16) (FIG. 3). No cytotoxic effects were observed with this method. [0164] For FIG. 1, three representative views are shown. The grey plane represents additive effects, and individual dots represent measurements of antiviral activity (yellow = Compound (1) alone, red = baloxavir alone, black = combination of Compound (1) and baloxavir). Blue areas on the plane indicate synergy. [0165] Based on these results, it can be concluded that Compound (1) showed synergistic antiviral activity in vitro with baloxavir. Overall, these data will enable exploration of the combination of Compound (1) and baloxavir in patients with influenza.

[0166] Example 2: In vitro combination study of Compound (1) and baloxavir [0167] To determine synergy effects of Compound (1) and baloxavir, baloxavir was spotted horizontally and the Compound (1) was spotted vertically to create a matrix of combinations of two compounds at variable concentrations. Compound (1) was spotted 100 nl from 0.045 mM to 0.00019 mM with 3-fold dilution. Baloxavir was spotted 100 nl from 0.075 mM to 0.0003 mM with 2-fold dilution. Plates were stored at -20 °C. Before the experiment, plates were placed at room temperature. At the day of infection, the influenza strain A/PR/8/1934(HlNl) virus was dropped to the plates containing test compounds at a concentration to reach a multiplicity of infection (MOI) of 0.01 using infection medium: Ultra MDCK serum free medium containing 0.0002% trypsine. Madin-Darby Canine Kidney (MDCK) cells were plated in assay medium (cat no BE12-749Q, lot no 8MB122, without trypsin) at a density of 6000 cells/well in 384-well white microplates (from Greiner Bio-One Catalog number GREI781098) using Multidrop (from Thermo, equipment id B0046). The assay plates were incubated for three days at incubator with 5% CO2 at 37 °C. At day 3, 40 mΐ of reconstituted ATPlite 1 step reagent was added to all wells. Luminescence signal was measured on ViewLux imager (Perkin Elmer), 3x binning and 0.1 s measuring time.

[0168] Results

[0169] Drug interactions were calculated using the BIGL method as discussed above in Example 1.

[0170] Example 3: Synergistic effect of the combination of Compound (1) and baloxavir

[0171] Overview

[0172] In this Example 3, Compound (1) and baloxavir were assayed for synergistic effects using the following dose levels in mM:

[0173] Normalization

[0174] Prior to the analysis the following normalization was applied to account for the between-plate variability for the values of the activity readouts: i

where “CC” = cell controls, and “VC” = virus controls.

[0175] Data Exploration

[0176] Plate layout: The heatmap plots present the raw (FIG. 2A) and normalized to % inhibition (FIG. 2B) values for the 3 different plates. For the normalized values the CC,

VC and MC columns were excluded. Overall, the inhibition is increasing as the doses of the two compounds are increasing.

[0177] FIG. 3 provides graphs of the viral control (VC), cell control (CC), medium control (MC), and MC by plate and column.

[0178] Activity

[0179] The plots provided by FIGS. 4 A, 4B, 5 A, and 5B present the normalized and raw data for the different doses of each compound.

[0180] Analysis and Results

[0181] Synergy analysis was performed following the Biochemically Intuitive Generalized Loewe Model (BIGL) methodology using R package BIGL (httDs://cran.r- Droiect.org/package=BIGL)

[0182] With this method:

1. Four-parameter log-logistic curves (Hill equation) are fitted to the monotherapy data.

2. Predicted (additive) response surface is calculated based on the Generalized Loewe equation.

3. Statistical tests are performed to assess deviations from the predicted surface: a global test (meanR) and point-by-point test (maxR).

4. The values of the test-statistics calculated in step 3 are displayed graphically. [0183] Note that the method relies on the assumption that monotherapy dose-response curves follow log-logistic shape (Hill equation).

[0184] In the BIGL assessment the variance of each dose combination is modeled as a function of its mean effect. Based on the parameter estimates from the model, the predicted variance is obtained for the corresponding mean effect at each dose combination. The predicted variances are used in the calculation of the test statistics.

[0185] Results (Unrestricted parameters of monotherapy fits)

[0186] For the analysis the parameters of the log-logistic curves are unrestricted, assuming a common baseline for the two monotherapy curves.

[0187] The graphic results provided by FIG. 6 (2D activity plot) can be used to calculate the EC50 of baloxavir (0.0041569 mM) and Compound (1) (0.0021605 mM).

[0188] The results of the instructions above is provided in FIGS. 7A (contour plot) and 7B (3D plot).

[0189] Table 1 presents the dose combinations of baloxavir and Compound (1) that show a synergistic/antagonistic effect. [0190] Table 1.

[0191] In order to investigate the antagonistic results, the maximal response level of the monotherapy fits is fixed at 1 (100% inhibition) and the baseline is fixed at 0 (0% inhibition).

[0192] Results (Fixed maximal response and baseline of monotherapy fits)

[0193] In the monotherapy fitting the maximal response is fixed at 1 (100% inhibition) and the two curves are assumed to have a common baseline at 0 (0% inhibition).

[0194] The graphic results provided by FIG. 8 (2D activity plot) can be used to calculate the EC50 of baloxavir (0.0041569 mM) and Compound (1) (0.0021605 mM).

[0195] The results of the instructions above are provided in FIGS. 9A (contour plot) and

9B (3D plot). Overall, there is evidence for non-additivity in the data. A total of nine (9) combinations were called synergistic according to the maxR test.

[0196] Table 2 presents the dose combinations of baloxavir and Compound (1) that show a synergistic/antagonistic effect. [0197] Table 2.

[0198] Based on this experiment, there is evidence of present synergistic effect between Compound (1) and baloxavir for the lower doses of Compound (1). There is an indication of antagonism between the two compounds at certain dose combinations, however this is an artifact of the low variability observed for these doses and the unrestricted monotherapy fits. When the upper asymptote of the monotherapy curves is fixed at 100% inhibition and the baseline is fixed at 0% inhibition then the antagonistic effects are not present. [0199] Example 4: Process for producing Tablet Compositions 1 -6, having 600 mg

Compound (1) (molar equivalent of the HC1 salt).

[0200] Step 1 : Fluid Bed Granulation Process

[0201] Binder solution: Hydroxypropylmethyl cellulose (HPMC) 2910 15 mPa.s (21.00 mg per unit) and polysorbate 20 (3.00 mg per unit; common commercial brand names include Scattics, Alkest TW 20, and Tween 20) were added to purified water (700.00 mg per unit) and mixed until a clear solution was obtained.

[0202] Granulation : Compound (1) HC1 hemihydrate salt Form A (668.40 mg per unit) and crospovidone (20.00 mg per unit) were transferred to a fluid bed granulator, and the resulting mixture was warmed while fluidizing. The binder solution was then sprayed upon the ingredients using standard wet granulation techniques.

[0203] The granulate was dried while fluidizing, the dried granules collected, and then packed in aluminum bags for later use.

[0204] Step 2: Blending and Tableting

[0205] Granules comprising Compound (1) HC1 hemihydrate salt Form A from step 1 and colloidal anhydrous silica are passed through a sieve (0.950 mm sieve size; 0.4 mm wire diameter) and mixed until homogeneous using a high speed blender (10 rpm, 5 min).

[0206] To this mixture was added a second mixture of silicified microcrystalline cellulose, microcrystalline cellulose (if present), pregelatinized starch (if present), and crospovidone that was previously also passed through a sieve (0.950 mm sieve size; 0.4 mm wire) diameter. The resulting mixture was then also blended until homogenous (10 rpm,

10 min).

[0207] To this mixture was added sodium stearyl fumarate that was previously also passed through a sieve (0.950 mm sieve size; 0.4 mm wire diameter). The resulting mixture was then also blended until homogenous (10 rpm, 10 min).

[0208] The resulting blended mixture was then compressed into tablets using a tableting press. The resulting tablets were then collected into aluminum laminated bags in suitable containers.

[0209] Step 3: Film Coating [0210] Coating powder, for example Opadry P White 85F 18422, was mixed with purified water (amounts specified in a per unit basis in Tables 3 A and 3B below) to create a coating suspension. The core tablets from steps 1 and 2 were transferred to a coating pan and sprayed with the coating suspension using the film coating technique, which comprises 1) loading the tablets into the coating pan and allowing them to pre-warm to the required temperature; 2) spraying the tablets with the coating suspension based on the parameters as set in Table 10 until the required weight film coating layer (weight gain) is achieved on the tablets (“the spraying phase”); and 3) drying the tablets at the set inlet and exhaust air temperature for 5 minutes. The dried, film coated tablets were then transferred to aluminum laminated bags in suitable containers. [0211] The component compositions of Tablet Compositions 1 - 6 are presented in Tables

3a and 3b below.

[0212] Table 3a: Tablet Compositions 1-3.

[0214] The parameters and results of the tablet compression process for Compositions 1-6 are provided in Table 4 below. [0215] Table 4: Tablet compression parameters for Compositions 1-6.

[0216] The parameters and results of the film coating process are provided in Table 5 below.

[0217] Table 5: Parameters and results of the film coating process for Compositions 1-6.

[0218] Example 5: Process for producing Tablet Composition 7 having 300 mg Compound (1) (molar equivalent of the HC1 salt).

[0219] Additional process and formula development directed 300 mg tablet compositions by reducing the amounts of excipients from those present in the 600 mg tablet compositions. Composition 8 was developed by dose-proportionally reducing the excipients present in the equivalent 600 mg Composition 3 tablet by 50%. Composition 8 is a formulation that is in between Composition 3 and Composition 7, and serves as a bridging composition between the two. [0220] It was demonstrated that the influence of the API properties was significant in the

300 mg formulations. Low-density APIs caused higher volumetric load in the fluid bed granulator resulting in flow challenges and this yielded granules with a significantly finer particle size, which was associated with flow problems during compression. API lots with higher bulk density and lower Specific Surface Area (SSA) were associated with better granulation outcomes. To overcome these issues, the composition of the intragranular phase was adjusted, increasing the binder and wetting agent concentration to improve binding properties of the granules and wetting of the API during granulation. The compositions of the eq. 300 mg Compound (1) final film coated tablets, is shown in Table 6 below.

[0221] Table 6: Tablet Composition 7 and 8.

[0222] Parameters and results of the tablet compression process for tablet composition 7 are provided in Tables 7 and 8 below. The tables present results for a single batch of tablet composition 7 at 1) four different sampling intervals during compression (Table 7) and 2) five different sampling intervals during compression (Table 8).

[0225] The conditions for granulation of Tablet Composition 8 are provided in Table 9 below.

[0226] Table 9: Granulation conditions for tablet composition 8.

[0227] Compression parameters and physical attributes of the tablet cores of Tablet Composition 8 are provided in Table 10 below.

[0228] Table 10: Compression parameters and physical attributes of tablet cores for Tablet Composition 8.

[0229] The coating parameters (provided as ranges) for the spraying phase of the Opadry P coating are summarized in Table 11 below. [0230] Table 11 : Spray coating parameters for Opadry P coating.

[0231] The experimental trials in small and scale up (30 Kg batch/45,000 tablets) batch sizes also confirm, robustness of the process. Generally, the physical attributes of the compressed tablets were acceptable, with no filming on the punches and acceptable final blend flowability, no rat-holding at larger scale, minimal batch to batch variability during dissolution.

[0232] Example 6: Process for producing Tablet Composition 9 having 300 mg Compound (1) (molar equivalent of the HC1 salt). [0233] It was demonstrated that a tablet formulation having added external lubrication as well as less lubricant compound in the formulation itself would not hinder tablet manufacturing and production. Sodium Stearyl Fumarate (SSF) was used both in the external phase (see Table 12a below) and as a process aid. As a process aid SSF is sprayed externally to the punches and dies via an External Lubrication System that is connected to the tablet press for lubrication during compression and ejection. The overage of SSF applied with the External Lubrication System is removed by vacuum and thus does not affect formula composition.

[0234] Table 12a: Tablet Composition 9.

[0235] Compression parameters and physical attributes of the tablet cores of Tablet Composition 9 are provided in Table 12b below.

[0236] Table 12b: Compression parameters and physical attributes of Composition 9.

OTHER EMBODIMENTS

[0237] It is to be understood that while the invention has been described in conjunction with the detailed description thereof, the foregoing description is intended to illustrate and not limit the scope of the invention, which is defined by the scope of the appended claims. Other aspects, advantages, and modifications are within the scope of the following claims.

Claims

WHAT IS CLAIMED IS:

1. A method for treating influenza virus infection in a subject or biological sample infected with influenza viruses comprising:

, and

2. The method of claim 1, wherein the administration of Compound (1) or a pharmaceutically acceptable salt thereof is prior to, concurrent with, or subsequent to the administration of baloxavir or a pharmaceutically acceptable salt or prodrug thereof.

3. The method of claim 1 or claim 2, further comprising administering to the subject or biological sample a dosage of from about 100 mg to about 1600 mg of Compound (1) or a pharmaceutically acceptable salt thereof, wherein the dosage is administered 1 to 4 times per day during a treatment period.

4. The method of any one of claims 1-3, wherein the dosage is from about 200 mg to about 1200 mg of Compound (1) or a pharmaceutically acceptable salt thereof, and the dosage is administered 1 to 3 times per day during the treatment period.

5. The method of any one of claims 1-4, wherein Compound (1) is administered to a subject in a unit dosage form, wherein the unit dosage form comprises from about 100 mg to about

800 mg of Compound (1) or a pharmaceutically acceptable salt thereof.

6. The method of claim 5, wherein the unit dosage form of Compound (1) is adapted for oral administration.

7. The method of claim 5 or claim 6, wherein Compound (1) in the unit dosage form is provided as a crystalline HC1 salt of Compound (1) · ½ H2O.

8. The method of claim 7, wherein the unit dosage form is a tablet comprising from about 200 mg to about 700 mg of the crystalline HC1 salt of Compound (1) · ½ H2O.

9. The method of claim 8, wherein the tablet comprises from about 250 mg to about 650 mg of the crystalline HC1 salt of Compound (1) · ½ H2O.

10. The method of claim 9, wherein the tablet comprises from about 300 mg to about

375 mg of the crystalline HC1 salt of Compound (1) · ½ H2O.

11. The method of any one of claims 8-10, wherein at least one tablet is administered to the subject 1 to 3 times per day during the treatment period.

12. The method of any one of claims 1-11, wherein the baloxavir or a pharmaceutically acceptable salt or prodrug thereof is administered to the sample or subject in a dosage of from about 10 mg to about 100 mg.

13. The method of claim 12, wherein the dosage of baloxavir or a pharmaceutically acceptable salt or prodrug thereof is from about 30 mg to about 90 mg.

14. The method of claim 12 or claim 13, wherein the dosage of baloxavir or a pharmaceutically acceptable salt or prodrug thereof is administered at least once during the treatment period.

15. The method of any one of claims 12-14, wherein baloxavir or a pharmaceutically acceptable salt or prodrug thereof is administered to a subject in a unit dosage form, wherein the unit dosage form comprises from about 10 mg to about 100 mg of baloxavir or a pharmaceutically acceptable salt or prodrug thereof.

16. The method of claim 15, wherein the unit dosage form of baloxavir is adapted for oral administration.

17. The method of claim 15 or claim 16, wherein baloxavir in the unit dosage form is provided as baloxavir marboxil.

18. The method of claim 17, wherein the unit dosage form is a tablet or capsule comprising from about 20 mg to about 90 mg of baloxavir marboxil.

19. The method of claim 18, wherein the tablet or capsule comprises from about 30 mg to about 50 mg of baloxavir marboxil.

20. The method of claim 18, wherein the tablet or capsule comprises from about 70 mg to about 90 mg of baloxavir marboxil.

21. The method of any one of claims 18-20, wherein one tablet or capsule is administered to the subject at least once during the treatment period.

22. The method of any one of claims 1-3, wherein the administration of Compound (1) or a pharmaceutically acceptable salt thereof in combination with the administration of baloxavir or a pharmaceutically acceptable salt or prodrug thereof results in a synergistic effect as compared with the administration of Compound (1) or a pharmaceutically acceptable salt thereof alone and the administration of baloxavir or a pharmaceutically acceptable salt or prodrug thereof alone.

23. A pharmaceutical product comprising:

; and

24. The pharmaceutical product of claim 23, wherein the second unit dosage form is provided as a single tablet or capsule.

25. The pharmaceutical product of claim 23 or claim 24, wherein the first unit dosage form is provided as one or more tablets, wherein each tablet comprises from about 200 mg to about

800 mg of the crystalline HC1 salt of Compound (1) · ½ H2O.

26. The pharmaceutical product of claim 25, wherein the first unit dosage form is provided as one or more tablets, wherein each tablet comprises from about 200 mg to about 400 mg of the crystalline HC1 salt of Compound (1) · ½ H2O.

27. A crystalline HC1 salt of Compovmd (1) · ½ H2O, wherein Compovmd (1) has the structure:

28. The crystalline HC1 salt of claim 27, wherein the crystalline HC1 salt is adapted for oral administration.

29. The crystalline HC1 salt of claim 28, wherein the crystalline HC1 salt is formulated as a tablet comprising from about 100 mg to about 700 mg of the crystalline HC1 salt.

30. The crystalline HC1 salt of claim 29, wherein the crystalline HC1 salt is formulated as a tablet comprising from about 200 mg to about 650 mg of the crystalline HC1 salt.

31. The crystalline HC1 salt of claim any one of claims 27-30, wherein baloxavir is adapted for oral administration.

32. The crystalline HC1 salt of claim 31, wherein baloxavir is baloxavir marboxil.

33. The crystalline HC1 salt of claim 32, wherein baloxavir is formulated as a tablet comprising from about 30 mg to about 50 mg or from about 70 mg to about 90 mg of baloxavir marboxil.

34. A method of inhibiting the replication of or reducing the number of influenza viruses in an in vitro biological sample, comprising: (a) administering to the biological sample a therapeutically effective amount of Compound (1) or a pharmaceutically acceptable salt thereof wherein Compound (1) has the structure:

, and

(b) administering to biological sample a therapeutically effective amount of baloxavir or a pharmaceutically acceptable salt or prodrug thereof, wherein

Compound (1) is administered to give a concentration of from about 0.0018 mM to 0.4400 mM, and baloxavir is administered to give a concentration of from about 0.003 mM to0.0750 mM.

35. The method of claim 34, wherein Compound (1) is administered prior to, concurrently with, or subsequent to the administration of baloxavir.

36. A pharmaceutical composition comprising Compound (1) or a pharmaceutically acceptable salt thereof, wherein Compound (1) has the structure

; and baloxavir or a pharmaceutically acceptable salt or prodrug thereof.

37. The pharmaceutical composition of claim 36, wherein the composition comprises

(b) a single dosage form comprising Compound (1) or a pharmaceutically acceptable salt thereof and baloxavir or a pharmaceutically acceptable salt or prodrug thereof, or

(c) separate dosage forms, wherein the separate dosage forms comprise a dosage form of Compound (1) or a pharmaceutically acceptable salt thereof and a separate dosage form of baloxavir or a pharmaceutically acceptable salt or prodrug thereof.

38. The pharmaceutical composition of claim 36 or claim 37, wherein the composition comprises a single dosage form comprising Compound (1) or a pharmaceutically acceptable salt thereof and baloxavir or a pharmaceutically acceptable salt or prodrug thereof.

39. The pharmaceutical composition of claim 38, wherein the single dosage form comprises a homogenous mixture of Compound (1) or a pharmaceutically acceptable salt thereof and baloxavir or a pharmaceutically acceptable salt or prodrug thereof.

40. The pharmaceutical composition of claim 38, wherein the single dosage form comprises 2 or more distinct compositions, wherein each composition comprises Compound (1) or a pharmaceutically acceptable salt thereof or baloxavir or a pharmaceutically acceptable salt or prodrug thereof or a mixture of any combination thereof.

41. The pharmaceutical composition of claim 36 or claim 37, wherein the composition comprises separate dosage forms.

42. The pharmaceutical composition of claim 41, wherein the separate dosage forms can be administered concurrently or consecutively.

43. The pharmaceutical composition of claim 42, wherein the separate dosage forms are administered consecutively as a first dosage form and a second dosage form.

44. The pharmaceutical composition of claim 43, wherein the second dosage form is administered from about one minute to about 48 hours after the first dose form.

45. The pharmaceutical composition of claim 44, wherein the second dosage form comprises Compound (1) or a pharmaceutically acceptable salt thereof, and the first dosage form comprises baloxavir or a pharmaceutically acceptable salt or prodrug thereof.

46. The pharmaceutical composition of claim 44, wherein the second dosage form comprises baloxavir or a pharmaceutically acceptable salt or prodrug thereof, and the first dosage form comprises Compound (1) or a pharmaceutically acceptable salt thereof.

47. The pharmaceutical composition of any one of claims 36-46, wherein Compound (1) or a pharmaceutically acceptable salt thereof is present in an amount sufficient to produce a concentration of from about 0.1 nM to about 0.1 mM of Compound (1) in the blood plasma of a patient after administration, and baloxavir or a pharmaceutically acceptable salt or prodrug thereof is present in an amount sufficient to produce a concentration of from about 0.1 nM to about 0.05 mM of baloxavir in the blood plasma of a patient after administration.

48. The pharmaceutical composition of any one of claims 36-47, wherein the therapeutic effect is synergistic compared to the therapeutic effect observed for Compound (1) or a pharmaceutically acceptable salt thereof or baloxavir or a pharmaceutically acceptable salt or prodrug thereof alone.

49. The pharmaceutical composition of any one of claims 36-48, wherein Compound (1) is provided in the composition as a crystalline HC1 salt of Compound (1) · ½ H2O.

50. The pharmaceutical composition of any one of claims 36-49, wherein baloxavir is provided in the composition as baloxavir marboxil.

51. The pharmaceutical composition of claim 41 , wherein the dosage form of

Compound (1) comprises from about 200 mg to about 800 mg of a crystalline HC1 salt of Compound (1) · ½ H2O.

52. The pharmaceutical composition of claim 51, wherein the dosage form of baloxavir comprises from about 30 mg to about 90 mg of baloxavir marboxil.

53. The pharmaceutical composition of claim 52, wherein the dosage form of Compound (1) and the dosage form of baloxavir are adapted for oral administration.

54. The pharmaceutical composition of claim 53, wherein the dosage form of

Compound (1) comprises one or more tablets, and the dosage form of baloxavir comprises a single tablet or capsule.