Combinations of influenza virus replication inhibitors

Abstract

This article provides a combination of compounds that can inhibit influenza virus replication, reduce influenza virus load, and / or treat influenza.

Description

Technical Field

[0001] This disclosure generally relates to combinations of influenza virus replication inhibitors, and methods for treating or preventing influenza infection or replication by administering said combination to patients in need of such treatment. Background Technology

[0002] Influenza spreads globally as a seasonal epidemic, causing hundreds of thousands of deaths each year, and millions in pandemic years. For example, the 20th century saw three influenza pandemics, killing tens of millions of people, each caused by the emergence of novel viral strains in humans. Typically, these novel strains are caused by existing influenza viruses that have spread from other animal species to humans.

[0003] Influenza is primarily transmitted from person to person through numerous virus-laden droplets produced when an infected person coughs or sneezes. These droplets can then remain on the mucous membranes of the upper respiratory tract of susceptible individuals in the vicinity of the infected person (e.g., within about 6 feet). Transmission can also occur through direct or indirect contact with respiratory secretions, such as touching surfaces contaminated with the influenza virus and subsequently touching the eyes, nose, or mouth. Adults can transmit the flu to others from one day before the onset of symptoms until about five days after the onset of symptoms. Young children and people with weakened immune systems may remain infectious for 10 days or more after the onset of symptoms.

[0004] Influenza viruses are RNA viruses belonging to the Orthomyxoviridae family, which includes five genera: influenza A virus, influenza B virus, influenza C virus, infectious salmon anemia virus (Isavirus), and Thogotovirus.

[0005] Influenza A viruses are the cause of both seasonal and pandemic influenza. It comprises one species, influenza A virus, and wild waterfowl are the natural hosts for many types of influenza A. Occasionally, the virus can spread to other species and subsequently cause devastating outbreaks in poultry or lead to human influenza pandemics. Influenza A viruses are the most virulent human pathogens of the three influenza types and cause the most severe illness. Influenza A viruses can be further classified into different serotypes based on antibody responses to these viruses. The confirmed human serotypes, ranked by the number of known human pandemic deaths, are: H1N1 (which caused the Spanish flu in 1918), H2N2 (which caused the Asian flu in 1957), H3N2 (which caused the Hong Kong flu in 1968), H5N1 (a pandemic threat during the 2007-2008 flu season), H7N7 (a potential pandemic threat), H1N2 (an endemic disease present in humans and pigs), H9N2, H7N2, H7N3, and H10N7.

[0006] Influenza B viruses belong to the genus *Influenza b*, which causes seasonal influenza and comprise a single species, *Influenza b* virus. Influenza b infects almost exclusively humans and is less common than influenza A. The only other animal known to be susceptible to influenza b is the seal. This type of influenza mutates at a rate 2 to 3 times slower than influenza A, resulting in lower genetic diversity and only one influenza B serotype. Due to this lack of antigenic diversity, some degree of immunity to influenza b is usually acquired at a very young age. However, the robust mutation of influenza b makes sustained immunity impossible. This reduced rate of antigenic change, combined with its limited host range (suppressing cross-species antigenic transfer), ensures that pandemics of influenza b do not occur.

[0007] The genus *Influenza C* contains one species, *Influenza C* virus, which infects humans and pigs and can cause severe illness and localized epidemics. However, *Influenza C* is less common than other types and generally appears to cause mild illness in children.

[0008] Influenza viruses are structurally remarkably similar across serotypes and species. The influenza virus genome consists of eight single-stranded RNAs packed into rod-shaped structures of varying sizes, known as the ribonucleoprotein complex (RNP). Each RNP contains a unique viral RNA, multiple copies of the scaffold nucleoprotein, and a heterotrimeric viral polymerase composed of PA, PB1, and PB2 subunits, which catalyzes the transcription and replication of the viral genome. Recent biochemical and structural studies of the influenza polymerase complex have provided insights into the mechanisms of cap-snatching and RNA synthesis by the influenza polymerase. Simply put, the PB2 cap-binding domain first binds to its 5' cap, isolating the host pre-mRNA. Subsequently, the PA (endonuclease subunit) cleaves 10–13 nucleotides of the pre-mRNA captured downstream of the cap. The PB2 subunit then rotates approximately 70° to guide the capping initiator to the PB1 polymerase active site. The PB1 subunit interacts directly with the PB2 and PA subunits. These subunits contain highly conserved domains across different influenza virus strains and have attracted attention as attractive targets for anti-influenza drugs. In addition to the polymerase complex, the influenza genome encodes its own neuraminidase (NA), hemagglutinin (HA), nucleoprotein (NP), matrix proteins M1 and M2, and non-structural proteins NS1 and NS2. NA is the target of the antiviral drug oseltamivir. Laninamivir Peramivir and Zanamivir These drugs inhibit the enzyme activity of NA, thereby slowing the release of progeny viruses from infected cells.

[0009] Influenza incurs direct costs due to lost productivity and related medical treatment, as well as indirect costs from preventative measures. In the United States, influenza costs more than $10 billion annually, and future pandemics are estimated to potentially cost hundreds of billions of dollars in direct and indirect costs. Prevention and control costs are also high. Governments worldwide have spent billions of dollars preparing for and planning for a potential H5N1 avian influenza pandemic, with expenses linked to the purchase of medicines and vaccines, disaster drills, and the development of strategies to improve border control.

[0010] Current treatment options for influenza include vaccination and chemotherapy or chemoprevention using antiviral drugs. Influenza vaccination is generally recommended for high-risk groups, such as children and the elderly, or people with asthma, diabetes, or heart disease. However, it is possible to contract influenza even after vaccination. The vaccine is reconfigured each season against several specific influenza virus strains, but may not include all strains that are effective in infecting the global population during that season. Manufacturers spend approximately six months reconfiguring and producing the millions of doses needed to combat seasonal epidemics; occasionally, new or overlooked strains become prominent during that period and infect vaccinated individuals (such as the H3N2 Fujian influenza in the 2003-2004 flu season). It is also possible to contract the specific strain that the vaccine is supposed to prevent, just before vaccination, as the vaccine takes about two weeks to become effective.

[0011] Furthermore, the effectiveness of these flu vaccines is variable. Due to the high mutation rate of the virus, a particular flu vaccine typically provides protection for no more than a few years. A vaccine formulated for one year may be ineffective the following year because the flu virus mutates rapidly over time, and different strains can become dominant.

[0012] Because it lacks an RNA proofreading enzyme, the RNA-dependent RNA polymerase of influenza vRNA produces a nucleotide insertion error approximately once every 10,000 nucleotides (which is the approximate length of influenza vRNA). Therefore, almost every newly manufactured influenza virus is a mutant antigenic drift. If more than one viral strain infects a single cell, then separating the genome into eight separate vRNA fragments allows for vRNA mixing or reassortment. The resulting rapid changes in viral genetics produce antigenic shifts and allow the virus to infect new host species and quickly overcome protective immunity.

[0013] Antiviral drugs can also be used to treat influenza, with norepinephrine (NA) inhibitors being particularly effective; however, viruses can develop resistance to approved NA antiviral drugs. Similarly, the emergence of multidrug-resistant pandemic influenza A viruses is well documented. Drug-resistant pandemic influenza A has become a major public health threat. In addition to drug-resistant influenza A viruses, NA inhibitors are approved for the treatment of early influenza infections (within 48 hours of the onset of influenza symptoms).

[0014] Therefore, there is still a need for drugs to treat influenza infections, such as those that can prolong treatment duration and / or reduce sensitivity to viral titers. Summary of the Invention

[0015] This disclosure generally relates to methods for treating influenza, methods for inhibiting the replication of influenza viruses, methods for reducing the amount of influenza viruses, and compounds and compositions that can be used in these methods.

[0016] This article provides a method for treating or preventing influenza infection or replication in an individual in need, the method comprising administering to the individual a therapeutically effective amount of (1) 3-(2-(5-chloro-1H-pyrrolo[2,3-b]pyridin-3-yl)-5-fluoro-7H-pyrrolo[2,3-d]pyrimidin-7-yl)bicyclo[2.2.2]octane-2-carboxylic acid or a pharmaceutically acceptable salt or solvate thereof, and (2) a second influenza treatment agent selected from the group consisting of baloxavirmarboxil, baloxavir, neuraminidase inhibitors and favipiravir, or a pharmaceutically acceptable salt or solvate thereof.

[0017] Furthermore, this disclosure provides a combination comprising (1) 3-(2-(5-chloro-1H-pyrrolo[2,3-b]pyridin-3-yl)-5-fluoro-7H-pyrrolo[2,3-d]pyrimidin-7-yl)bicyclo[2.2.2]octane-2-carboxylic acid or a pharmaceutically acceptable salt or solvate thereof, and (2) baloxavir or baloxavir macozylate, or a pharmaceutically acceptable salt or solvate thereof; and (3) a neuraminidase inhibitor, such as oseltamivir, oseltamivir acid, zanamivir, lanamivir, peramivir, or a pharmaceutically acceptable salt or solvate thereof.

[0018] Further methods are provided for administering safe and effective amounts of combinations as disclosed herein to biological samples or patients.

[0019] This article also provides methods for reducing the amount of influenza virus in a biological sample or patient by administering a safe and effective amount of the combination as disclosed herein to the biological sample or patient.

[0020] Further methods for treating or preventing influenza A or influenza B infection in a patient include administering a safe and effective amount of the combination as disclosed herein to the patient.

[0021] The uses of the combinations described herein are also provided for inhibiting or reducing the replication of influenza virus in biological samples or patients, for reducing the amount of influenza virus in biological samples or patients, or for treating influenza in patients.

[0022] This article further provides the use of the combination described herein for manufacturing agents for treating influenza in patients, for reducing the amount of influenza virus in biological samples or patients, or for inhibiting the replication of influenza virus in biological samples or patients.

[0023] The combinations disclosed in this article can be administered together or individually to an individual, patient, or host. Attached Figure Description

[0024] Figure 1 MacSynergy™ II schema is used to illustrate the role of the combination of compound 1 and baloxavir in the analysis of influenza virus A / PR / 8 / 34 (H1N1) replication in MDCK cells. Synergistic / antagonistic effects are shown compared to the 95% confidence interval. Throughout the text, compound 1 refers to 3-(2-(5-chloro-1H-pyrrolo[2,3-b]pyridin-3-yl)-5-fluoro-7H-pyrrolo[2,3-d]pyrimidin-7-yl)bicyclo[2.2.2]octane-2-carboxylic acid.

[0025] Figure 2 MacSynergy aims to demonstrate the role of the combination of compound 1 and oseltamivir in the analysis of influenza virus A / PR / 8 / 34 (H1N1) replication in MDCK cells. TM II. Diagram. Shows synergistic / antagonistic effects compared to the 95% confidence interval.

[0026] Figure 3 MacSynergy aims to demonstrate the role of the combination of compound 1 and favipiravir in the analysis of influenza virus A / PR / 8 / 34 (H1N1) replication in MDCK cells. TM II. Diagram. Shows synergistic / antagonistic effects compared to the 95% confidence interval. Detailed Implementation

[0027] This document discloses combinations of anti-influenza compounds (antiviral agents) and their use in inhibiting the activity of influenza viruses. In some aspects, this disclosure generally relates to the use of the compositions described herein for inhibiting the replication of influenza virus in biological samples or patients, for reducing the amount of influenza virus in biological samples or patients (reducing viral titers), and for treating or preventing influenza in patients. The combinations disclosed herein can be co-formulated or administered alone to an individual, patient, or host.

[0028] The combination particularly comprises 3-(2-(5-chloro-1H-pyrrolo[2,3-b]pyridin-3-yl)-5-fluoro-7H-pyrrolo[2,3-d]pyrimidin-7-yl)bicyclo[2.2.2]octane-2-carboxylic acid or a pharmaceutically acceptable salt or solvate thereof as the active ingredient, or referred to herein as "Compound 1". This compound is considered to be an inhibitor of the PB2 domain of CAP.

[0029] In various embodiments, compound 1 is used in combination with a second antiviral agent as a preventative or therapeutic agent against influenza, such as against influenza virus replication or infection. Influenza may be a pandemic or drug-resistant pandemic / seasonal influenza virus. In some cases, the influenza may be influenza A or influenza B. The second antiviral agent may be a polymerase inhibitor, endonuclease inhibitor, or neuraminidase inhibitor, or an influenza vaccine. Further discussion regarding the timing of administration of compound 1 and the second antiviral agent is provided below.

[0030] In some aspects, a combination is provided comprising 3-(2-(5-chloro-1H-pyrrolo[2,3-b]pyridin-3-yl)-5-fluoro-7H-pyrrolo[2,3-d]pyrimidin-7-yl)bicyclo[2.2.2]octane-2-carboxylic acid or a pharmaceutically acceptable salt or solvate thereof and a second antiviral agent, and in some cases, the second antiviral agent is selected from the group consisting of baloxavir, baloxavir macozylate, neuraminidase inhibitors and favipiravir, or a pharmaceutically acceptable salt or solvate thereof.

[0031] When administered alone, Compound 1 and the second antiviral agent may be administered simultaneously (e.g., within about 5 to 10 minutes of each other) or at intervals of one or more hours (e.g., 1 hour, 2 hours, 3 hours, 4 hours, 5 hours, 6 hours, 7 hours, 8 hours, 9 hours, 10 hours, 11 hours, 12 hours, 24 hours, 26 hours, 48 ​​hours, or 72 hours). In some cases, Compound 1 may be administered before the second antiviral agent. In some other cases, Compound 1 may be administered after the second antiviral agent. Further discussion regarding the timing of administration of Compound 1 and the second antiviral agent is provided below.

[0032] In some aspects, a combination is provided comprising a therapeutically effective amount of a) 3-(2-(5-chloro-1H-pyrrolo[2,3-b]pyridin-3-yl)-5-fluoro-7H-pyrrolo[2,3-d]pyrimidin-7-yl)bicyclo[2.2.2]octane-2-carboxylic acid or a pharmaceutically acceptable salt or solvate thereof and b) a therapeutically effective amount of a second antiviral agent, and in some cases, the second antiviral agent is selected from the group consisting of baloxavir, baloxavir macozylate, neuraminidase inhibitors and favipiravir, or a pharmaceutically acceptable salt or solvate thereof.

[0033] In other respects, the use provides therapeutically effective amounts of the combinations disclosed herein for the treatment or prevention of influenza virus infection or replication in human patients. For example, the influenza virus may be a pandemic or drug-resistant pandemic / seasonal influenza virus. In some cases, the influenza virus may be influenza A or influenza B. Other influenza viruses are described below.

[0034] In other respects, the use of 3-(2-(5-chloro-1H-pyrrolo[2,3-b]pyridin-3-yl)-5-fluoro-7H-pyrrolo[2,3-d]pyrimidin-7-yl)bicyclo[2.2.2]octane-2-carboxylic acid or a pharmaceutically acceptable salt or solvate thereof in combination with a second antiviral agent is provided for the manufacture of an agent for the treatment or prevention of influenza virus infection or replication, and in some cases, the second antiviral agent is selected from the group consisting of baloxavir, baloxavir macozylate, neuraminidase inhibitors and favipiravir, or a pharmaceutically acceptable salt or solvate thereof.

[0035] In other respects, the use of 3-(2-(5-chloro-1H-pyrrolo[2,3-b]pyridin-3-yl)-5-fluoro-7H-pyrrolo[2,3-d]pyrimidin-7-yl)bicyclo[2.2.2]octane-2-carboxylic acid or a pharmaceutically acceptable salt or solvate thereof in combination with a second antiviral agent for the inhibition of influenza virus infection or replication is provided, and in some cases, the second antiviral agent is selected from the group consisting of baloxavir, baloxavir macozylate, neuraminidase inhibitors and favipiravir, or a pharmaceutically acceptable salt or solvate thereof.

[0036] In other aspects, a method for treating or preventing influenza virus infection or replication is provided, comprising administering to a human patient suffering from or at risk of influenza virus infection the following combination: a) a therapeutically effective amount of 3-(2-(5-chloro-1H-pyrrolo[2,3-b]pyridin-3-yl)-5-fluoro-7H-pyrrolo[2,3-d]pyrimidin-7-yl)bicyclo[2.2.2]octane-2-carboxylic acid or a pharmaceutically acceptable salt or solvate thereof, and b) a therapeutically effective amount of a second antiviral agent, wherein in some cases the second antiviral agent is selected from the group consisting of baloxavir, baloxavir macozide, neuraminidase inhibitors and favipiravir, or a pharmaceutically acceptable salt or solvate thereof.

[0037] In other respects, a method for treating or preventing influenza virus infection or replication is provided, comprising administering to a human patient suffering from or at risk of influenza infection a dose of about 10 to 1,000 mg / kg of 3-(2-(5-chloro-1H-pyrrolo[2,3-b]pyridin-3-yl)-5-fluoro-7H-pyrrolo[2,3-d]pyrimidin-7-yl)bicyclo[2.2.2]octane-2-carboxylic acid or a pharmaceutically acceptable salt or solvate thereof, and a therapeutically effective amount of a second antiviral agent, wherein in some cases the second antiviral agent is selected from the group consisting of baloxavir, baloxavir macozylate, neuraminidase inhibitors and favipiravir, or a pharmaceutically acceptable salt or solvate thereof.

[0038] In other respects, a pharmaceutical composition is provided for treating or preventing influenza virus infection or replication in a patient, comprising 3-(2-(5-chloro-1H-pyrrolo[2,3-b]pyridin-3-yl)-5-fluoro-7H-pyrrolo[2,3-d]pyrimidin-7-yl)bicyclo[2.2.2]octane-2-carboxylic acid or a pharmaceutically acceptable salt or solvate thereof as an active ingredient, wherein said composition is administered in combination with a second antiviral agent, in some cases, the second antiviral agent being selected from the group consisting of baloxavir, baloxavir macozide, neuraminidase inhibitors, and favipiravir, or a pharmaceutically acceptable salt or solvate thereof.

[0039] In other aspects, a method is provided for inhibiting the endonuclease activity of influenza polymerase in influenza A or influenza B, the method comprising contacting a virus with a combination of compound 1 as disclosed herein and a second antiviral agent.

[0040] In other aspects, a method for treating or preventing influenza A or influenza B infection in a host is provided, the method comprising administering to the host a therapeutic amount of a combination of compound 1 as disclosed herein and a second antiviral agent.

[0041] In other aspects, a method is provided for reducing the endonuclease activity of influenza polymerase in a host of influenza A or influenza B, the method comprising administering to the host a therapeutic amount of a combination of compound 1 as disclosed herein and a second antiviral agent.

[0042] In other respects, a method for reducing influenza virus replication in a host is provided, the method comprising administering to the host a therapeutic amount of a combination of compound 1 as disclosed herein and a second antiviral agent.

[0043] In some embodiments, a method is provided for using a combination of the disclosed compound 1 and a second antiviral agent, the method further comprising contacting the influenza virus with a therapeutically effective amount of a third antiviral agent or administering a therapeutically effective amount of the third antiviral agent to a host. For example, in some embodiments, the method may further include administering an influenza vaccine to the host before, after, or simultaneously with the combination. In some cases, the disclosed method comprises administering compound 1 and an influenza vaccine without administering another antiviral agent (i.e., the vaccine is the second antiviral agent). In some cases, compound 1 and the influenza vaccine are administered simultaneously. In some cases, compound 1 and the influenza vaccine are co-formulated.

[0044] In other respects, the use of a combination of compound 1 as disclosed herein with a second antiviral agent for the treatment of influenza A or influenza B virus infection is provided.

[0045] In other respects, there is provided the use of a combination of compound 1 as disclosed herein with a second antiviral agent for the manufacture of an agent for the treatment of influenza A or influenza B virus infection.

[0046] In other aspects, a combination is provided comprising a) 3-(2-(5-chloro-1H-pyrrolo[2,3-b]pyridin-3-yl)-5-fluoro-7H-pyrrolo[2,3-d]pyrimidin-7-yl)bicyclo[2.2.2]octane-2-carboxylic acid or a pharmaceutically acceptable salt or solvate thereof, b) baloxavir, baloxavir macozylate or a pharmaceutically acceptable salt or solvate thereof, and c) a neuraminidase inhibitor.

[0047] The neuraminidase inhibitor used in the methods disclosed herein may be oseltamivir, oseltamivir acid, or zanamivir. Lanamivir Or peramivir, or a pharmaceutically acceptable salt or solvate thereof. Endonuclease inhibitors may be used in the methods disclosed herein, and in some cases, endonuclease inhibitors are referred to as "PA" inhibitors. In various embodiments, the endonuclease inhibitor may be baloxavir or baloxavir masocytin, or a pharmaceutically acceptable salt or solvate thereof. Polymerase inhibitors may be used in the methods disclosed herein, and in some cases, polymerase inhibitors are referred to as "PB1" inhibitors. In various embodiments, the polymerase inhibitor may be favipiravir, or a pharmaceutically acceptable salt or solvate thereof. Influenza vaccines may be used in the methods disclosed herein.

[0048] In various embodiments of the aspects disclosed herein, the second antiviral agent is baloxavir, baloxavir masocytin, or a pharmaceutically acceptable salt or solvate thereof. In other embodiments of the aspects disclosed herein, the second antiviral agent is a neuraminidase inhibitor (e.g., more specifically, oseltamivir, oseltamivir acid, or a pharmaceutically acceptable salt or solvate thereof). In other embodiments of the aspects disclosed herein, the second antiviral agent is favipiravir or a pharmaceutically acceptable salt or solvate thereof.

[0049] How to use

[0050] The combinations described herein can be used to reduce viral titers in biological samples (e.g., infected cell cultures) or in humans (e.g., lung viral titers in patients).

[0051] As used in this article, the terms “influenza virus-mediated symptoms,” “influenza infection,” or “influenza” are used interchangeably to refer to illness caused by infection with the influenza virus.

[0052] Influenza is an infectious disease affecting birds and mammals caused by influenza viruses. Influenza viruses are RNA viruses belonging to the Orthomyxoviridae family, which includes five genera: influenza A virus, influenza B virus, influenza C virus, infectious salmon anemia virus, and togovirus. The influenza A virus genus contains one species, influenza A virus, which can be further divided into different serotypes based on antibody responses to these viruses: H1N1, H2N2, H3N2, H5N1, H7N7, H1N2, H9N2, H7N2, H7N3, H7N9, and H10N7. The influenza B virus genus contains one species, influenza B virus. Influenza B virus almost exclusively infects humans and is less prevalent than influenza A virus. The influenza C virus genus contains one species, influenza C virus, which infects humans and pigs and can cause severe illness and localized epidemics. However, influenza C virus is less prevalent than other types and generally appears to cause mild illness in children.

[0053] In some embodiments, the influenza or influenza virus is associated with influenza A or influenza B virus. In some embodiments, the influenza or influenza virus is associated with influenza A virus. In some specific embodiments, the influenza A virus is H1N1, H2N2, H3N2, H7N9, or H5N1. In some embodiments, the disclosed combination effectively inhibits the growth or replication of pandemic or drug-resistant pandemic / seasonal influenza viruses.

[0054] In humans, common symptoms of influenza include chills, fever, sore throat, muscle aches, severe headache, cough, weakness, and general malaise. In more severe cases, influenza can cause pneumonia, which can be fatal, especially in young children and the elderly. Although often confused with the common cold, influenza is a much more serious illness caused by different types of viruses. Influenza can cause nausea and vomiting, especially in children, but these symptoms are more like those of an unrelated gastroenteritis, sometimes called "stomach flu" or "24-hour flu."

[0055] Flu symptoms can begin quite suddenly, one to two days after infection. The first symptom is usually chills or cold shivering, but fever is also common in the early stages of infection, with temperatures ranging from 38°C to 39°C (approximately 100°F to 103°F). Many people become severely ill, requiring bed rest for several days, experiencing general malaise, which is more pronounced in the back and legs. Flu symptoms may include: general aches and pains (especially in the joints and throat), extreme chills and fever, fatigue, headache, irritated and watery eyes, red eyes, red and swollen skin (especially the face), red and swollen mouth, red and swollen throat and nose, and abdominal pain (in children with influenza B). Flu symptoms are not specific and overlap with many pathogens (“influenza-like illnesses”). Laboratory data are usually required to confirm the diagnosis.

[0056] The terms “disease,” “symptom,” and “symptom” are used interchangeably in this article to refer to medical or pathological symptoms mediated by the influenza virus.

[0057] As used herein, the terms “individual,” “host,” and “patient” are used interchangeably. The terms “individual,” “host,” and “patient” refer to an animal (e.g., a bird such as a chicken, quail, or turkey) or a mammal, specifically a mammal such as a non-primate (e.g., a cow, pig, horse, sheep, rabbit, guinea pig, rat, cat, dog, or mouse) and a primate (e.g., a monkey, chimpanzee, or human), and more specifically, a human. In some embodiments, the individual is a non-human animal, such as livestock (e.g., a horse, cow, pig, or sheep), or a pet (e.g., a dog, cat, guinea pig, or rabbit). In a preferred embodiment, the individual is a human.

[0058] As used herein, the term "biological sample" includes (but is not limited to): cell cultures or extracts thereof; biopsy material obtained from mammals or extracts thereof; blood, saliva, urine, feces, semen, tears or other bodily fluids or extracts thereof.

[0059] As used herein, the term "inhibition of influenza virus replication" includes both reducing the amount of viral replication (e.g., reducing it by at least 10%) and completely suppressing viral replication (i.e., reducing the amount of viral replication by 100%). In some embodiments, influenza virus replication is inhibited by at least 50%, at least 65%, at least 75%, at least 85%, at least 90%, or at least 95%.

[0060] Influenza virus replication can be measured by any suitable method known in the field. For example, the influenza virus titer in a biological sample (e.g., an infected cell culture) or in a human (e.g., a patient's lung virus titer) can be measured. More specifically, for cell-based analyses, in each case of in vitro cell culture, the virus is added to the culture with or without a test reagent, and the virus-dependent endpoint is assessed after an appropriate time period. For typical analyses, Madin-Darby canine kidney cells (MDCK) suitable for influenza virus strain A / Puerto Rico / 8 / 34 and standard tissue cultures can be used. The first class of cell analyses that can be used depends on the death of infected target cells (a process called cytopathic effect (CPE), in which viral infection causes depletion of cell-derived cells and eventual cell lysis. In the first type of cell assay, a small fraction of cells in the wells of a microtiter plate are infected (typically 1 / 10 to 1 / 1000), allowing the virus to undergo several rounds of replication over 48 to 72 hours. Cell death is then measured by the decrease in cellular ATP content compared to an uninfected control. The second type of cell assay, which can be used, depends on the proliferation of virus-specific RNA molecules in the infected cells, where RNA levels are directly measured using branched-strand DNA hybridization (bDNA). In this second type of cell assay, a smaller number of cells are initially infected in the wells of the microtiter plate, allowing the virus to replicate in the infected cells and spread to additional rounds of cells. Cells are then lysed, and viral RNA levels are measured. This assay is typically stopped early, 18 to 36 hours later, when all target cells are still viable. Viral RNA is quantified by hybridization to specific oligonucleotide probes immobilized in the wells of the assay plate, followed by amplification of the signal through hybridization to additional probes linked to a reporter enzyme.

[0061] As used herein, “viral titer” or “titer” is a measure of viral concentration. Titer testing can be performed using serial dilutions to obtain approximate quantitative information from an analytical procedure that essentially only assesses positive or negative results. The titer corresponds to the highest dilution factor that still produces a positive reading; for example, positive readings from the first eight consecutive 2-fold dilutions are converted to a titer of 1:256. A specific example is viral titer. To determine the titer, several dilutions will be prepared, such as 10... -1 10 -2 10 -3 、…、10-8 The lowest viral concentration that can still infect cells is the viral titer.

[0062] As used herein, the term "treat / treatment / treating" refers to both therapeutic and preventative treatment. For example, therapeutic treatment includes the reduction or slowing of the progression, severity, and / or duration of influenza virus-mediated illness, or the improvement of one or more symptoms (specifically, one or more identifiable symptoms) of influenza virus-mediated illness caused by administration of one or more therapies (e.g., one or more therapeutic agents, such as compounds or compositions described herein). In a particular embodiment, therapeutic treatment includes improving at least one measurable bodily parameter of influenza virus-mediated symptoms. In other embodiments, therapeutic treatment includes physically suppressing the development of influenza virus-mediated symptoms by, for example, stabilizing identifiable symptoms, or physiologically by, for example, stabilizing bodily parameters, or both. In other embodiments, therapeutic treatment includes reducing or stabilizing influenza virus-mediated infection. Antiviral drugs can be used in community settings to treat individuals already suffering from influenza to reduce the severity of symptoms and the number of days they are ill.

[0063] As used herein, the terms “prevention,” “preventative,” “preventative use,” and “preventative treatment” refer to any medical or public health procedure intended to prevent, rather than treat or cure, a disease. As used herein, the term “prevent / prevention / preventing” means reducing the risk of developing or producing a given symptom, or reducing or suppressing the recurrence of the symptom in an individual who is not currently ill but has become or is likely to become ill. The term “chemoprevention” refers to the use of a drug, such as a small molecule drug (rather than a vaccine), to prevent a symptom or disease.

[0064] Prophylactic use includes use in situations where an outbreak has been detected to prevent the spread of infection in places where many people live in close contact with each other at high risk of severe influenza complications (e.g., hospitals, daycare centers, prisons, nursing homes, etc.). It also includes use in individuals who need protection from influenza but have not received protection after vaccination (e.g., due to a weakened immune system), or when a vaccine is unavailable to them or when they cannot obtain a vaccine due to side effects. It also includes use within two weeks of vaccination or during any period after vaccination but before the vaccine becomes effective. Prophylactic use may also include treating individuals who do not have influenza or are not considered at high risk of complications to reduce the chance of them contracting influenza and transmitting it to high-risk individuals in close contact with them (e.g., healthcare workers, home care workers, etc.).

[0065] As used in this article and according to the United States Centers for Disease Control and Prevention (US CDC), an influenza “outbreak” is defined as an acute febrile respiratory illness (AFRI) occurring within a 48 to 72-hour period in close proximity to each other (e.g., in the same area of ​​an assisted living facility, in the same household, etc.) or when any individual in the analyzed population tests positive for influenza.

[0066] In some embodiments, the combination is applicable as a preventative or control measure for patients (specifically humans) susceptible to complications arising from influenza virus infection. The combination may be applicable in control measures in cases of confirmed index cases or outbreaks to prevent the spread of infection to the rest of the community or population.

[0067] As used herein, "effective amount" means an amount sufficient to elicit the desired biological response. In this disclosure, the desired biological response is the inhibition of influenza virus replication, reduction of the amount of influenza virus, or mitigation or improvement of the severity, duration, progression, or onset of influenza virus infection, prevention of the acceleration of influenza virus infection, prevention of recurrence, manifestation, onset, or progression of symptoms associated with influenza virus infection, or enhancement or improvement of the preventive or therapeutic effect of another therapy used against influenza infection. The precise amount of compound administered to an individual will depend on the administration pattern, the type and severity of the infection, and the individual's characteristics (e.g., general health condition, age, sex, weight, and tolerance to the drug). Those skilled in the art will be able to determine the appropriate dosage based on these and other factors.

[0068] Therefore, when 3-(2-(5-chloro-1H-pyrrolo[2,3-b]pyridin-3-yl)-5-fluoro-7H-pyrrolo[2,3-d]pyrimidin-7-yl)bicyclo[2.2.2]octane-2-carboxylic acid is administered co-administered with other antiviral agents, such as when co-administered with anti-influenza drugs, the effective amount of the second agent will depend on the type of drug used. A safe amount is one with minimal or acceptable number and severity of side effects, as readily determined by someone knowledgeable in the field. Suitable doses of approved agents are known and can be adjusted by someone knowledgeable in the field based on the individual's condition, the type of condition being treated, and the amount of the compound described herein used. Where the amount is not explicitly stated, a safe and effective amount should be assumed. For example, an individual may be given 3-(2-(5-chloro-1H-pyrrolo[2,3-b]pyridin-3-yl)-5-fluoro-7H-pyrrolo[2,3-d]pyrimidin-7-yl)bicyclo[2.2.2]octane-2-carboxylic acid or a pharmaceutically acceptable salt or solvate thereof in a dose range of about 0.01 to 100 mg / kg body weight / day.

[0069] As used herein, a “safe and effective amount” of a compound or composition described herein is an effective amount of a compound or composition that does not cause excessive or harmful side effects in patients.

[0070] Generally, dosing regimens can be selected based on a variety of factors, including: the condition being treated and its severity; the activity of the specific compound used; the specific combination used; the patient's age, weight, general health condition, sex, and diet; the timing, route of administration, and excretion rate of the specific compound used; the individual's renal and hepatic function; the specific compound used or its salts, the duration of treatment; drugs used in combination with or concurrently with the specific compound used; and similar factors well known in the medical field. A person of ordinary skill in the art can readily determine and prescribe a safe and effective amount of the compound described herein required for the treatment, prevention, inhibition (complete or partial) or cessation of disease progression.

[0071] The dosage of the compounds described herein may range from about 0.01 to about 100 mg / kg body weight / day, from about 0.01 to about 50 mg / kg body weight / day, from about 0.1 to about 50 mg / kg body weight / day, or from about 1 to about 25 mg / kg body weight / day. It should be understood that the total daily dose may be given as a single dose or may be given multiple times, such as twice a day (e.g., every 12 hours), three times a day (e.g., every 8 hours), or four times a day (e.g., every 6 hours).

[0072] For therapeutic treatment, the compound described herein may be administered to the patient within, for example, 48 hours (or 40 hours, or less than 2 days, or less than 1.5 days, or 24 hours) of the onset of symptoms (e.g., nasal congestion, sore throat, cough, pain, fatigue, headache, and chills / night sweats). Therapeutic treatment may continue for any appropriate duration, such as 5 days, 7 days, 10 days, 14 days, etc. For prophylactic treatment during community outbreaks, the compound described herein may be administered to the patient, for example, within 2 days of the onset of symptoms indicative cases, and may continue for any appropriate duration, such as 7 days, 10 days, 14 days, 20 days, 28 days, 35 days, 42 days, etc.

[0073] Combination therapy

[0074] The combinations described herein can be administered alone or in further combination with additional suitable therapeutic agents (e.g., a third antiviral agent or a vaccine). When using combination therapy, a safe and effective amount may be achieved using a first amount of compound 1, namely 3-(2-(5-chloro-1H-pyrrolo[2,3-b]pyridin-3-yl)-5-fluoro-7H-pyrrolo[2,3-d]pyrimidin-7-yl)bicyclo[2.2.2]octane-2-carboxylic acid or a pharmaceutically acceptable salt or solvate thereof, a second antiviral agent, and an amount of one or more third antiviral agents. In some cases, the third antiviral agent is a pyrazinamide antiviral compound, an influenza neuraminidase inhibitor, or an influenza PB1 polymerase domain inhibitor. Other treatment combinations may be achieved with additional suitable therapeutic agents (e.g., antiviral agents or vaccines).

[0075] The second antiviral agent suitable for these combinations is baloxavir (CAS No. 1985605-59-1), which is a prodrug of masocytin (CAS No. 1985606-14-1; trademark name). It is being developed by Shionogi Co. (a Japanese pharmaceutical company) for the treatment of influenza A and influenza B.

[0076] Another suitable second antiviral agent for these combinations is oseltamivir phosphate (CAS No. 204255-11-8; brand name). It is a neuroglucosidase inhibitor being developed by Roche pharmaceutical company as a drug for the treatment or prevention of influenza infection.

[0077] Another suitable second antiviral agent for these combinations is favipiravir (CAS No. 259793-96-9; T-705; trademark name). Favipiravir is a pyrazinamide derivative developed by Toyama Chemical Co., Ltd. (a Japanese pharmaceutical company) for the treatment of RNA viruses, including influenza A and influenza B. It is approved in Japan for use in 200 mg tablets to treat influenza. For adults, favipiravir can be administered orally at doses from 10 mg to 10,000 mg daily. In some embodiments, favipiravir can be administered orally at doses from 100 mg to 4,000 mg daily. In some embodiments, favipiravir can be administered orally at a dose of 1,600 mg twice daily on day 1, and 600 mg twice daily from day 2 to day 5, for a total dosing period of five days.

[0078] Kitano et al. reported that the combination of baloxavir and a neuraminidase inhibitor synergistically inhibited influenza A / H1N1 virus replication in MDCK cells (Open Forum Infectious Diseases, 4, Issue suppl_1, October 1, 2017, p. S371). Therefore, in some embodiments, a combination comprising 3-(2-(5-chloro-1H-pyrrolo[2,3-b]pyridin-3-yl)-5-fluoro-7H-pyrrolo[2,3-d]pyrimidin-7-yl)bicyclo[2.2.2]octane-2-carboxylic acid and baloxavir or baloxavir macozylate and a neuraminidase inhibitor. Neuraminidase inhibitors suitable for this combination include, for example, oseltamivir, oseltamivir acid, zanamivir, lanamivir, and peramivir, or more specifically, oseltamivir phosphate, oseltamivir acid, zanamivir hydrate, lanamivir, and peramivir trihydrate.

[0079] In some embodiments of this disclosure, compound 1 or a pharmaceutically acceptable salt thereof and a second antiviral agent are each administered in a safe and effective amount (i.e., each in an amount that would be therapeutically effective when administered alone). In some embodiments, compound 1 and the second antiviral agent are each administered in an amount that does not provide a therapeutic effect when administered alone (below a therapeutic amount). In some embodiments, compound 1 may be administered in a safe and effective amount, while the second antiviral agent is administered below a therapeutic amount. In some embodiments, compound 1 may be administered below a therapeutic amount, while the second antiviral agent is administered in a safe and effective amount.

[0080] As used herein, the terms “combination therapy,” “combination,” and “co-administered” or “co-administered” are used interchangeably to refer to the use of more than one therapy (e.g., one or more preventative and / or therapeutic agents). The use of these terms does not restrict the order in which therapies (e.g., preventative and / or therapeutic agents) are administered to an individual.

[0081] Co-administration can cover administering the first and second amounts of the compound in a substantially simultaneous manner, such as in the form of a single pharmaceutical composition, for example, capsules or tablets having a fixed ratio of first and second amounts, or in the form of multiple separate capsules or tablets. Furthermore, such co-administration can also cover administering each compound of the combination in any sequential order.

[0082] In some embodiments, this disclosure pertains to a combination therapy method for inhibiting influenza virus replication in a biological sample or patient, or for treating or preventing influenza virus infection in a patient, using compounds or pharmaceutical compositions of this disclosure. Therefore, the pharmaceutical compositions described herein also include those compositions exhibiting anti-influenza virus activity comprising 3-(2-(5-chloro-1H-pyrrolo[2,3-b]pyridin-3-yl)-5-fluoro-7H-pyrrolo[2,3-d]pyrimidin-7-yl)bicyclo[2.2.2]octane-2-carboxylic acid in combination with a second antiviral agent.

[0083] Methods of use also include combinations of compound 1 with a second antiviral agent, other combinations with another antiviral agent, and / or vaccination using an influenza vaccine.

[0084] When co-administering a first amount of 3-(2-(5-chloro-1H-pyrrolo[2,3-b]pyridin-3-yl)-5-fluoro-7H-pyrrolo[2,3-d]pyrimidin-7-yl)bicyclo[2.2.2]octane-2-carboxylic acid and a second amount of a second antiviral agent, the administration of 3-(2-(5-chloro-1H-pyrrolo[2,3-b]pyridin-3-yl)-5-fluoro-7H-pyrrolo[2,3-d]pyrimidin-7-yl)bicyclo[2.2.2]octane-2-carboxylic acid and the second antiviral agent is sufficiently close in timing to achieve the desired therapeutic effect. For example, the time interval between each administration can range from minutes to hours and can be selected by considering the characteristics of each compound (e.g., properties, solubility, bioavailability, plasma half-life, and kinetic profile) to produce the desired therapeutic effect. For example, 3-(2-(5-chloro-1H-pyrrolo[2,3-b]pyridin-3-yl)-5-fluoro-7H-pyrrolo[2,3-d]pyrimidin-7-yl)bicyclo[2.2.2]octane-2-carboxylic acid and the second antiviral agent may be administered in any order within approximately 24 hours, approximately 16 hours, approximately 8 hours, approximately 4 hours, approximately 1 hour, or approximately 30 minutes of each other.

[0085] More specifically, 3-(2-(5-chloro-1H-pyrrolo[2,3-b]pyridin-3-yl)-5-fluoro-7H-pyrrolo[2,3-d]pyrimidin-7-yl)bicyclo[2.2.2]octane-2-carboxylic acid may be administered to an individual before (e.g., 15 minutes, 30 minutes, 45 minutes, 1 hour, 2 hours, 4 hours, 6 hours, 12 hours, 24 hours, 48 ​​hours, 72 hours, 96 hours, 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 8 weeks, or 12 weeks prior to, simultaneously with, or after (e.g., 15 minutes, 30 minutes, 45 minutes, 1 hour, 2 hours, 4 hours, 6 hours, 12 hours, 24 hours, 48 ​​hours, 72 hours, 96 hours, 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 8 weeks, or 12 weeks after) the administration of a second antiviral agent.

[0086] As described herein, the co-administration of a certain amount of 3-(2-(5-chloro-1H-pyrrolo[2,3-b]pyridin-3-yl)-5-fluoro-7H-pyrrolo[2,3-d]pyrimidin-7-yl)bicyclo[2.2.2]octane-2-carboxylic acid and a certain amount of a second antiviral agent can produce enhanced or synergistic therapeutic effects, wherein the observed combined effect is greater than the expected additive effect produced by administering a certain amount of compound 1 and a certain amount of the second antiviral agent alone.

[0087] As used herein, the term "synergistic" refers to a combination of the present disclosure that is more effective than the additive effect of the component compounds. The synergistic effect of a combination of therapies (e.g., a combination of preventative or therapeutic agents) may allow for the use of one or more therapies at lower doses and / or for administration to an individual less frequently. The ability to utilize lower doses of therapies (e.g., preventative or therapeutic agents) and / or administer the therapy less frequently reduces the toxicity associated with administration of the therapy to an individual without diminishing the efficacy of the therapy in preventing, treating, or managing a condition. Furthermore, synergistic effects may improve the efficacy of the agents in preventing, treating, or managing a condition. Finally, the synergistic effect of a combination of therapies (e.g., a combination of preventative or therapeutic agents) may avoid or reduce adverse or unwanted side effects associated with the use of any single therapy.

[0088] Furthermore, when using the combination therapy of this disclosure, the component therapeutic agents can be administered to allow for longer intervals between each administration (e.g., days, weeks, or months).

[0089] The presence of synergistic effects can be determined using appropriate methods for assessing drug interactions. Appropriate methods include, for example, the Sigmoid-Emax equation (Holford, NHG and Scheiner, LB, *Clinical Pharmacokinetics*, 6:429-453 (1981)), the Loewe additivity equation (Loewe, S and Muischnek, H., *Archives of Experimental Pharmacology and Pathology*, 114:313-326 (1926)), and the median effect equation (Chou, TC and Talalay, P., *Advances in Enzyme Regulation*, 22:27-55 (1984)). Each of the equations mentioned above can be applied in conjunction with experimental data to generate corresponding graphs that help assess the effects of drug combinations. The corresponding graphs associated with the equations mentioned above are concentration-response curves, equivalence curves, and combination exponent curves. MacSynergy TM II is a recognized graphical software suitable for calculating composite indices (Prichard and Shipman, 1990).

[0090] Influenza vaccine

[0091] The compounds described herein can be prophylactically combined with anti-influenza vaccines. These vaccines can be administered, for example, subcutaneously or intranasally. Subcutaneous vaccination typically induces neutralizing IgG antibodies in the serum and is highly effective in preventing the progression of the disease to more severe conditions such as pneumonia and similar illnesses. However, in the upper respiratory tract mucosa, which is the site of infection, IgA is the main protective component. Since IgA is not induced by subcutaneous administration, it can also be advantageously administered via the intranasal route.

[0092] Definitions and General Terms

[0093] The compounds described herein are defined by their chemical structure and / or chemical name. When a compound is mentioned by both its chemical structure and chemical name, and the chemical structure contradicts the chemical name, the chemical structure determines the compound's identity.

[0094] Pharmaceutically acceptable salts and solvates

[0095] The compounds described herein may exist in free form or, where appropriate, in salt form. Pharmaceutically acceptable salts are of particular interest because they are suitable for administering the compounds, as components of the described combinations, for medical purposes. Non-pharmaceutically acceptable salts are suitable for manufacturing processes to achieve separation and purification purposes, and in some cases, for separating stereoisomers of the compounds described herein or their intermediates.

[0096] As used herein, the term “pharmaceutically acceptable salt” refers to a salt of a compound that, within the bounds of reasonable medical judgment, is suitable for contact with human and lower animal tissues without undue side effects (e.g., toxicity, irritation, allergic reactions, and similar effects) and is commensurate with a reasonable benefit / risk ratio.

[0097] Pharmaceutically acceptable salts are well known in the field. For example, SMBerge et al. described pharmaceutically acceptable salts in detail in the *Journal of Pharmaceutical Sciences*, 1977, 66, 1-19, which is incorporated herein by reference. Pharmaceutically acceptable salts of the compounds described herein include those derived from suitable inorganic acids and bases, as well as organic acids and bases. These salts can be prepared in situ during the final isolation and purification of the compounds.

[0098] In the case of compounds described herein containing a base or sufficiently basic bioelectron isotopes, acid addition salts can be prepared by 1) reacting the purified compound, in its free base form, with a suitable organic or inorganic acid and 2) separating the resulting salt. In practice, acid addition salts may be the more suitable form for use, and the use of said salt is equivalent to the use of the free base form.

[0099] Examples of pharmaceutically acceptable non-toxic acid addition salts are salts formed by amino groups with inorganic acids (e.g., hydrochloric acid, hydrobromic acid, phosphoric acid, sulfuric acid, and perchloric acid) or with organic acids (e.g., acetic acid, oxalic acid, maleic acid, tartaric acid, citric acid, succinic acid, or malonic acid), or salts formed by other methods used in the field (e.g., ion exchange). Other pharmaceutically acceptable salts include adipates, alginates, ascorbic acid salts, aspartate salts, benzenesulfonates, benzoates, hydrogen sulfates, borates, butyrates, camphorates, camphorsulfonates, citrates, cyclopentanepropionates, diglucuronates, dodecyl sulfates, ethanesulfonates, formates, transbutenedioic acid salts, glucohepanoates, glycerophosphates, glycolate salts, glucuronates, glycolate salts, hemisulfates, heptahydrates, hexanoates, hydrochlorides, hydrobromide salts, hydroiodates, 2-hydroxy- Ethyl sulfonate, lactobionate, lactate, lauryl sulfate, malate, maleate, malonate, methanesulfonate, 2-naphthalenesulfonate, nicotinate, nitrate, oleate, oxalate, palmitate, dihydroxynaphthalate, pectate, persulfate, 3-phenylpropionate, phosphate, picrate, pentanoate, propionate, salicylate, stearate, succinate, sulfate, tartrate, thiocyanate, p-toluenesulfonate, undecanoate, valerate and similar salts.

[0100] In cases where the compounds described herein contain a carboxylic acid group or sufficiently acidic bioelectron isotopes, base addition salts can be prepared by 1) reacting the purified compound in its acidic form with a suitable organic or inorganic base and 2) separating the resulting salt. In practice, the use of base addition salts may be in a more suitable form, and the use of the salt form itself is equivalent to the use of the free acid form. Salts derived from suitable bases include alkali metal (e.g., sodium, lithium, and potassium) salts, alkaline earth metal (e.g., magnesium and calcium) salts, ammonium salts, and N2 salts. + (C 1-4 Alkyl)4 salts. This disclosure also envisions quaternization of any basic nitrogen-containing group of the compounds disclosed herein. Such quaternization can yield water-soluble, oil-soluble, or dispersible products.

[0101] Base addition salts include pharmaceutically acceptable metal salts and amine salts. Suitable metal salts include sodium, potassium, calcium, barium, zinc, magnesium, and aluminum. Sodium and potassium salts are generally preferred. Where appropriate, other pharmaceutically acceptable salts include non-toxic ammonium, quaternary ammonium, and amine cations formed using the relative ions of, for example, halides, hydroxides, carboxylates, sulfates, phosphates, nitrates, low-carbon alkyl sulfonates, and aryl sulfonates. Suitable inorganic base addition salts are prepared from metal bases, including sodium hydride, sodium hydroxide, potassium hydroxide, calcium hydroxide, aluminum hydroxide, lithium hydroxide, magnesium hydroxide, zinc hydroxide, and the like. Suitable amine base addition salts are prepared from amines, which are frequently used in medicinal chemistry due to their low toxicity and acceptability for medical use. Ammonia, ethylenediamine, N-methylglucosamine, lysine, arginine, ornithine, choline, N,N'-diphenylmethylethylenediamine, chloroprocaine, diethanolamine, procaine, N-phenylmethylphenylethylamine, diethylamine, piperazine, tris(hydroxymethyl)aminomethane, tetramethylammonium hydroxide, triethylamine, diphenylmethylamine, diphenylhydroxymethylamine, dehydrorosinamine, N-ethylpiperidine, benzylamine, tetramethylammonium, tetraethylammonium, methylamine, dimethylamine, trimethylamine, ethylamine, basic amino acids, dicyclohexylamine and analogues.

[0102] Other acids and bases, when not pharmaceutically acceptable on their own, can be used to prepare salts suitable as intermediates in obtaining the compounds described herein and their pharmaceutically acceptable acid or base addition salts.

[0103] The components of this combination may include mixtures / combinations of different pharmaceutically acceptable salts and mixtures / combinations of compounds in free form with pharmaceutically acceptable salts.

[0104] The components of a compound can exist as a solvate. The term "solvate" refers to a molecular complex of a compound (including its salts) and one or more solvent molecules. These solvent molecules are those commonly used in pharmaceutical techniques and are known to be harmless to the recipient, such as water, ethanol, dimethyl sulfoxide, acetone, and other common organic solvents. The term "hydrate" refers to a molecular complex containing a compound and water.

[0105] Pharmaceutical Composition

[0106] The compounds described herein can be formulated into pharmaceutical compositions further comprising pharmaceutically acceptable carriers, diluents, adjuvants, or mediators. In some embodiments, this disclosure relates to a pharmaceutical composition comprising the compounds described herein and pharmaceutically acceptable carriers, diluents, adjuvants, or mediators. In some embodiments, this disclosure includes a pharmaceutical composition comprising a safe and effective amount of the compounds described herein or pharmaceutically acceptable salts thereof and pharmaceutically acceptable carriers, diluents, adjuvants, or mediators. Pharmaceutically acceptable carriers include, for example, pharmaceutical diluents, excipients, or carriers appropriately selected with respect to the intended form of administration and conforming to conventional pharmaceutical practices.

[0107] "Effective dose" includes "therapeutic effective dose" and "preventive effective dose." The term "therapeutic effective dose" refers to the dose that is effective in treating and / or reducing influenza virus infection in patients. The term "preventive effective dose" refers to the dose that is effective in preventing and / or substantially reducing the chance or size of an influenza virus outbreak.

[0108] Pharmaceutically acceptable carriers may contain inert components that do not excessively inhibit the biological activity of the compound. Pharmaceutically acceptable carriers should be biocompatible, for example, non-toxic, non-inflammatory, non-immunogenic, or have no other undesirable reactions or side effects after administration to an individual. Standard pharmaceutical compounding techniques may be used.

[0109] Pharmaceutically acceptable carriers, adjuvants, or mediators as used herein include any solvent, diluent or other liquid mediator, dispersant or suspending agent, surfactant, isotonic agent, thickener or emulsifier, preservative, solid binder, lubricant, and the like suitable for the particular dosage form desired. Remington's Pharmaceutical Sciences, 16th edition, EW Martin (Mack Publishing Co., Easton, PA, 1980) discloses various carriers used in the formulation of pharmaceutically acceptable compositions and known techniques for their preparation. Unless any conventional carrier medium is incompatible with the compounds described herein, for example by producing any undesirable biological effects or otherwise interacting in a harmful manner with any other component of the pharmaceutically acceptable composition, its use is contemplated within the scope of this disclosure.

[0110] As used herein, the term "side effect" encompasses an unwanted and adverse effect of a therapy (e.g., a preventative or therapeutic agent). Side effects are always undesirable, but undesirable effects are not necessarily adverse. Adverse effects of a therapy (e.g., a preventative or therapeutic agent) can be harmful, uncomfortable, or risky. Side effects include (but are not limited to) fever, chills, drowsiness, gastrointestinal toxicity (including gastrointestinal ulcers and erosions), nausea, vomiting, neurotoxicity, nephrotoxicity, kidney toxicity (including conditions such as papillary necrosis and chronic interstitial nephritis), hepatotoxicity (including elevated serum liver enzyme levels), bone marrow toxicity (including leukopenia, bone marrow suppression, thrombocytopenia, and anemia), dry mouth, metallic taste, prolonged pregnancy, weakness, somnolence, pain (including myasthenia gravis, bone pain, and headache), hair loss, fatigue, dizziness, extrapyramidal symptoms, akathisia, cardiovascular disturbances, and sexual dysfunction.

[0111] Some examples of substances that can serve as pharmaceutically acceptable carriers include (but are not limited to): ion exchangers; alumina; aluminum stearate; lecithin; serum proteins (e.g., human serum albumin); buffering substances (e.g., twin 80, phosphate esters, glycine, sorbic acid, or potassium sorbate); partial glyceride mixtures of saturated vegetable fatty acids, water, salts, or electrolytes (e.g., protamine sulfate, disodium hydrogen phosphate, potassium hydrogen phosphate, sodium chloride, or zinc salts); colloidal silica; magnesium trisilicate; polyvinylpyrrolidone; polyacrylates; waxes; polyethylene-polyoxypropylene block copolymers; methylcellulose; hydroxypropyl methylcellulose; lanolin; sugars such as lactose, glucose, and sucrose; starches such as corn starch and potato starch; cellulose and its derivatives such as sodium carboxymethyl cellulose, ethyl cellulose, and cellulose acetate; astragalus powder; malt; gelatin; slippery gelatin. Stone; excipients, such as cocoa butter and suppository wax; oils, such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil, and soybean oil; diols, such as propylene glycol or polyethylene glycol; esters, such as ethyl oleate and ethyl laurate; agar; buffers, such as magnesium hydroxide and aluminum hydroxide; alginic acid; pyrogen-free water; isotropic physiological saline; Ringer's solution; ethanol; and phosphate buffer solutions; and, at the discretion of the formulator, other non-toxic and compatible lubricants, such as sodium lauryl sulfate and magnesium stearate, as well as colorants, release agents, coating agents, sweeteners, flavoring agents, aromatizers, preservatives, and antioxidants may also be present in the composition.

[0112] formulations for transpulmonary delivery

[0113] In some embodiments, the pharmaceutical compositions described herein are suitable for direct administration to the lower respiratory tract (e.g., the lungs) via inhalation. Compositions administered by inhalation may be in the form of an inhalable powder composition or a liquid or powder spray, and may be administered using a standard powder inhaler or aerosol dispensing device. Such devices are well known. For administration by inhalation, the powder formulation typically contains the active compound along with an inert solid powdered diluent (e.g., lactose or starch). Inhalable dry powder compositions may be present in capsules and cartridges of gelatin or similar substances, or in blister packs of laminated aluminum foil for inhalers or blow-offs. Each capsule or cartridge may typically contain, for example, from about 10 mg to about 100 g of each active compound. Alternatively, the compositions described herein may be excipient-free.

[0114] Inhalable compositions can be packaged for single-dose or multi-dose delivery. For example, compositions can be packaged for multi-dose delivery in a manner similar to that described below: GB2242134, U.S. Patent Nos. 6,632,666, 5,860,419, 5,873,360, and 5,590,645 (all describing “Diskus” devices); or GB2178965, GB2129691, GB2169265, U.S. Patent Nos. 4,778,054, 4,811,731, and 5,035,237 (which describe “Diskhaler” devices); or EP 69715 (“Turbuhaler” device) or GB 2064336 and U.S. Patent No. 4,353,656 (“Rotahaler” device).

[0115] Spray compositions intended for local delivery to the lungs via inhalation can be formulated as aqueous solutions or suspensions or as aerosols delivered by pressurized packaging, such as in a controlled-dose inhaler (MDI), using suitable liquefied propellants, including hydrofluoroalkanes such as dichlorodifluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, and especially 1,1,1,2-tetrafluoroethane, 1,1,1,2,3,3,3-heptafluoro-n-propane and mixtures thereof. Aerosol compositions suitable for inhalation can be presented in suspension or solution form.

[0116] Medications administered via inhalation typically have a controlled particle size. The most preferred particle size for inhalation into the bronchial system is typically from about 1 μm to about 10 μm, and in some embodiments, from about 2 μm to about 5 μm. Particles with a size greater than about 20 μm are generally too large to reach the smaller airways during inhalation. To achieve these particle sizes, the particles of the active ingredient may undergo a particle size reduction process, such as micronization. The desired particle size fraction can be separated by air separation or sieving. Preferably, the particles are crystallized.

[0117] Nasal sprays can be formulated using aqueous or non-aqueous media, with the addition of agents such as thickeners, pH-adjusting buffer salts or acids or bases, isotonic regulators, or antioxidants.

[0118] Solutions intended for inhalation via nebulization can be formulated using aqueous media with added reagents such as acids or alkalis, buffer salts, isotropic modifiers, or antimicrobial agents. They can be sterilized by filtration or heating in an autoclave, or presented as a non-sterile product. The nebulizer supplies the aerosol in the form of a mist generated from the aqueous solution of the formulation.

[0119] In some embodiments, the pharmaceutical compositions described herein may be formulated together with supplemental active ingredients.

[0120] In some embodiments, the pharmaceutical compositions described herein are administered by a dry powder inhaler.

[0121] In other embodiments, the pharmaceutical compositions described herein are optionally combined via an aerosol dispensing device, for example... The inhalation chamber is used to administer the medication.

[0122] The carrier may be a solvent or dispersion medium containing, for example, water, ethanol, polyols (e.g., glycerol, propylene glycol, liquid polyethylene glycol, and the like), suitable mixtures thereof, and / or vegetable oils. Appropriate flowability may be maintained, for example, by using a coating such as lecithin, by maintaining the desired particle size in the case of a dispersion, and by using a surfactant. Prevention of microbial activity in the compositions described herein may be achieved by adding antibacterial and antifungal agents, such as parabens, chlorobutanol, phenol, sorbic acid, thimerosal, and the like. In many cases, isotonic agents, such as sugars or sodium chloride, are preferred. The absorption of the injectable composition may be prolonged by using an absorption-retarding agent (e.g., aluminum monostearate and gelatin) in the composition.

[0123] In some embodiments, the pharmaceutical compositions described herein may be contained within a matrix that controls the release of the composition. In some embodiments, the matrix may comprise: lipids, polyvinyl alcohol, polyvinyl acetate, polycaprolactone, poly(glycolic acid), poly(lactic acid), polycaprolactone, polylactic acid, polyanhydride, polylactide-co-glycolic acid, polyamino acids, polyethylene oxide, acrylic-terminated polyethylene oxide, polyamide, polyethylene, polyacrylonitrile, polyphosphazene, poly(orthoester), sucrose acetate isobutyrate (SAIB), and combinations thereof, such as U.S. Patent Nos. 6,667,371 and 6,667,371. Other polymers disclosed in Nos. 13,355, 6,596,296, 6,413,536, 5,968,543, 4,079,038, 4,093,709, 4,131,648, 4,138,344, 4,180,646, 4,304,767, and 4,946,931, each of which is expressly incorporated herein by reference in its entirety. In these embodiments, the matrix sustains drug release.

[0124] Pharmaceutically acceptable carriers and / or diluents may also include any solvents, dispersion media, coatings, antibacterial and / or antifungal agents, isotonics, and absorption delay agents, as well as the like. The use of such media and agents for pharmaceutically active substances is well known in the art. Unless any conventional media or reagent is incompatible with the active ingredient, its use in pharmaceutical compositions should be considered.

[0125] The pharmaceutical compositions described herein can be formulated for administration using conventional techniques. See, for example, Remington, *The Science and Practice of Pharmacy* (20th edition, 2000). For instance, the intranasal pharmaceutical compositions of this disclosure can be formulated as aerosols (this term includes both liquid and dry powder aerosols). As is known to those skilled in the art, aerosols of liquid particles can be generated by any suitable means, such as using a pressure-driven aerosol nebulizer or an ultrasonic nebulizer. See, for example, U.S. Patent No. 4,501,729. Aerosols of solid particles (e.g., lyophilized, freeze-dried, etc.) can also be generated using any solid particulate pharmaceutical aerosol generator by techniques known in the pharmaceutical field. As another example, the pharmaceutical compositions of this disclosure can be formulated as an on-demand soluble form, providing a lyophilized portion of the pharmaceutical composition and a dissolved solution portion of the pharmaceutical composition.

[0126] In some embodiments of this disclosure, the pharmaceutical composition is in the form of an aqueous suspension, which may be prepared from a solution or suspension. For solutions or suspensions, the dosage form may consist of microcells of lipophilic substances, liposomes (phospholipid vesicles / membranes), and / or fatty acids (e.g., palmitic acid). In specific embodiments, the pharmaceutical composition is a solution or suspension capable of dissolving in fluids secreted by the epithelial mucosa of the tissue to which the pharmaceutical composition is administered, applied, and / or delivered, which may advantageously enhance absorption.

[0127] Pharmaceutical compositions may be aqueous solutions, non-aqueous solutions, or a combination of aqueous and non-aqueous solutions.

[0128] Suitable aqueous solutions include (but are not limited to): hydrogels, aqueous suspensions, aqueous microsphere suspensions, aqueous microsphere dispersions, aqueous liposome dispersions, aqueous microcells of liposomes, aqueous microemulsions, and any combination thereof, or any other aqueous solution soluble in fluids secreted by the nasal mucosa. Exemplary non-aqueous solutions include (but are not limited to): non-aqueous gels, non-aqueous suspensions, non-aqueous microsphere suspensions, non-aqueous microsphere dispersions, non-aqueous liposome dispersions, non-aqueous emulsions, non-aqueous microemulsions, and any combination thereof, or any other non-aqueous solution soluble in or mixable with fluids secreted by the mucosa.

[0129] Examples of powder formulations include (but are not limited to): pure powder mixtures, micronized powders, freeze-dried powders, lyophilized powders, powder microspheres, coated powder microspheres, liposome dispersions, and any combination thereof. Powder microspheres may be formed from various polysaccharides and celluloses, including (but not limited to) starch, methylcellulose, gum arabic, carboxymethylcellulose, hydroxypropylcellulose, carbomer, polyvinyl alcohol alginate, gum arabic, polyglucosamine, and any combination thereof.

[0130] In certain embodiments, the inhalable composition is one that is at least partially or even substantially (e.g., at least 80%, 90%, 95% or more) soluble in fluids secreted by mucous membranes to facilitate absorption. The composition may be formulated with a carrier and / or other substances that promote the dissolution of the agent in the secretions, including (but not limited to) fatty acids (e.g., palmitic acid), gangliosides (e.g., GM-1), phospholipids (e.g., phosphatidylserine), and emulsifiers (e.g., polysorbate 80).

[0131] Those skilled in the art will understand that, for intranasal administration or delivery, nasal secretions can alter the pH of the administered dose because the amount of the administered pharmaceutical composition is typically small, as the pH range in the nasal cavity can be as wide as 5 to 8. Such alterations can affect the concentration of unionized drug available for absorption. Therefore, in representative embodiments, the pharmaceutical composition further includes a buffer to maintain or adjust the pH in situ. Typical buffers include (but are not limited to) ascorbate, acetate, citrate, gluten, carbonate, and phosphate buffers.

[0132] In some embodiments, the pH of the pharmaceutical composition is selected such that the internal environment of the mucosal tissue after administration is neutral or acidic, which (1) provides an active compound in an unionized form for absorption, (2) inhibits the growth of pathogens that are more likely to exist in an alkaline environment, and (3) reduces the likelihood of irritation to the mucosa.

[0133] For liquid and powder sprays or aerosols, pharmaceutical compositions can be formulated to have any suitable and desired particle size or droplet size. In illustrative examples, the majority and / or average particle or droplet size ranges from about 1, 2.5, 5, 10, 15, or 20 micrometers and / or from about 25, 30, 40, 45, 50, 60, 75, 100, 125, 150, 175, 200, 225, 250, 275, 300, 325, 350, 375, 400, or 425 micrometers (inclusive of all combinations thereof). Representative examples of suitable ranges of most and / or average particle size or droplet size include (but are not limited to) about 5 to 100 micrometers, about 10 to 60 micrometers, about 175 to 325 micrometers, and about 220 to 300 micrometers, which facilitate the safe and effective deposition of active compounds, for example, in the nasal cavity (e.g., in the upper third of the nasal cavity, the superior nasal meatus, the olfactory region, and / or the sinus regions leading to the olfactory nerve pathway). Generally, particles or droplets smaller than about 5 micrometers will deposit in the trachea or even the lungs, while particles or droplets of about 50 micrometers or larger generally do not reach the nasal cavity and deposit in the anterior part of the nose.

[0134] International Patent Publication WO 2005 / 023335 describes particles and droplets having diameter sizes suitable for practicing representative embodiments of this disclosure. In particular embodiments, the particles or droplets have an average diameter of about 5 to 30 micrometers, about 10 to 20 micrometers, about 10 to 17 micrometers, about 10 to 15 micrometers, about 12 to 17 micrometers, about 10 to 15 micrometers, or about 10 to 12 micrometers. The particles may have an average diameter or size “substantially” as described herein, i.e., at least about 50%, 60%, 70%, 80%, 90%, or 95% or more of the particles have a specified diameter or size range.

[0135] The pharmaceutical compositions described herein can be delivered in a sprayed or atomized liquid form with droplet sizes as described above.

[0136] According to specific embodiments including intranasal delivery methods, it may be desirable to prolong the residence time of the pharmaceutical composition in the nasal cavity (e.g., in the upper third of the nasal cavity, the superior nasal meatus, the olfactory region, and / or the sinus region), for example, to enhance absorption. Therefore, the pharmaceutical composition may optionally be formulated with bioadhesive polymers, gums (e.g., saccharin), polyglucosamine (e.g., highly purified cationic polysaccharides), pectin (or any carbohydrate that thickens or emulsifies like a gel when applied to the nasal mucosa), microspheres (e.g., starch, albumin, polydextrose, cyclodextrin), gelatin, liposomes, carbomer, polyvinyl alcohol, alginate, gum arabic, polyglucosamine, and / or cellulose (e.g., methyl or propyl cellulose; hydroxy or carboxy cellulose; carboxymethyl or hydroxypropyl cellulose) as agents to increase the residence time in the nasal cavity. As another method, increasing the viscosity of the formulation can also provide a means of prolonging the contact between the agent and the nasal epithelium. The pharmaceutical composition may be formulated as a nasal lotion, ointment, or gel, which offers the advantage of topical application due to its viscosity.

[0137] A moist and highly vascularized membrane promotes rapid absorption. Therefore, pharmaceutical compositions may optionally contain humectants (especially in the case of gel compositions) to ensure adequate intranasal moisture content. Examples of suitable humectants include (but are not limited to) glycerin / glycerol, mineral oil, vegetable oil, membrane conditioners, soothing agents, and / or sugar alcohols (e.g., xylitol, sorbitol; and / or mannitol). The concentration of the humectant in the pharmaceutical composition will vary depending on the selected pharmaceutical agent and formulation.

[0138] Pharmaceutical compositions may optionally include absorption enhancers, such as agents that inhibit enzyme activity, reduce the viscosity or elasticity of mucus, reduce the mucociliary clearance effect, open tightly bound and / or dissolve active compounds. Chemical enhancers are known in the art and include chelating agents (e.g., EDTA), fatty acids, bile salts, surfactants and / or preservatives. Permeation enhancers are particularly suitable when formulating compounds that exhibit poor membrane permeability, lack lipophilicity, and / or are degraded by aminopeptidase. The concentration of the absorption enhancer in the pharmaceutical composition will vary depending on the selected pharmaceutical agent and formulation.

[0139] To extend shelf life, preservatives may optionally be added to the pharmaceutical composition. Suitable preservatives include (but are not limited to) benzyl alcohol, parabens, thimerosal, chlorobutanol, and benzylamino chloride, and combinations thereof. The concentration of the preservative will vary depending on the preservative used, the compound being formulated, the formulation, and the like. In representative embodiments, the preservative is present in an amount of about 2% by weight or less.

[0140] The pharmaceutical compositions described herein may optionally contain odorants, such as those described in EP 0 504263B1, to provide a sense of odor in order to facilitate inhalation of the composition, thereby promoting delivery to the olfactory region and / or triggering transmission via olfactory neurons.

[0141] As an alternative, the composition may contain flavoring agents, for example, to enhance the taste and / or individual acceptability of the composition.

[0142] Porous particles delivered via the lungs

[0143] In some embodiments, the particles are porous, resulting in an appropriate density to prevent deposition in the posterior pharynx when administered via an inhaler. The combination of relatively large particle size and relatively low density avoids phagocytosis in the lungs, provides appropriately targeted delivery, avoids systemic delivery of the component, and delivers high concentrations of the component within the lungs.

[0144] Representative methods for preparing these particles and for delivering these particles are described, for example, in U.S. Patent Nos. 7,384,649, 7,182,961, 7,146,978, 7,048,908, 6,956,021, 6,766,799, and 6,732,732.

[0145] Other patents disclosing these particles include U.S. Patent Nos. 7,279,182, 7,252,840, 7,032,593, 7,008,644, 6,848,197, and 6,749,835.

[0146] U.S. Patent No. 7,678,364 discloses a method for delivering particles to the pulmonary system, comprising: administering a safe and effective amount of dry powder to the respiratory tract of a patient requiring treatment, prevention, or diagnosis, said dry powder comprising: a) a polyvalent metal cation in combination with a therapeutic, preventive, or diagnostic agent; b) a pharmaceutically acceptable carrier; and c) a component containing polyvalent metal cations, wherein said dry powder is spray-dried and has a total polyvalent metal cation content of about 10% w / w or more of the total weight of the pharmaceutical agent, about 0.4 g / cm³. 3 Or lower knock tightness, median geometric diameter of about 5 micrometers to about 30 micrometers and aerodynamic diameter of about 1 to about 5 micrometers.

[0147] The amount of the compound or its salt described herein present in the particles can range from about 0.1% by weight to about 95% by weight, although in some cases it can even be up to 100%. For example, about 1% to about 50% by weight, such as about 5% to about 30% by weight. Particles in which the drug is distributed throughout the entire particle are preferred.

[0148] In some embodiments, the particles comprise surfactants other than the phospholipids described above. As used herein, the term "surfactant" refers to any agent that is preferably absorbed at the interface between two immiscible phases (e.g., the interface between water and an organic polymer solution, a water / air interface, or an organic solvent / air interface). Surfactants generally have both hydrophilic and lipophilic moieties, which facilitates the formation of an external environment that does not attract coated particles upon absorption, thus reducing particle aggregation. Surfactants can also promote the absorption of therapeutic or diagnostic agents and improve the bioavailability of pharmaceuticals.

[0149] Suitable surfactants that can be used to manufacture the particles described herein include (but are not limited to): hexadecyl alcohol; fatty alcohols, such as polyethylene glycol (PEG); polyoxyethylene-9-lauryl ether; surfactant fatty acids, such as palmitic acid or oleic acid; cholates; surfactantin; poloxamer; and sorbitol fatty acid esters, such as sorbitol trioleate. 80; and tyloxapol.

[0150] The surfactant may be present in the particles in an amount ranging from about 0 to about 5% by weight. Preferably, it may be present in the particles in an amount ranging from about 0.1 to about 1.0% by weight, for example, 1.0% by weight.

[0151] It has a content of less than approximately 0.4 g / cm³ 3 Particles with a high knock-tightness, a median diameter of at least about 5 μm, and an aerodynamic diameter of about 1 μm to about 5 μm or about 1 μm to about 3 μm are better able to avoid inertial and gravitational deposition in the oropharyngeal region and target the respiratory tract or deep lungs. Using larger, more porous particles is advantageous because they can be atomized more effectively than smaller, denser particles (such as those currently used in inhalation therapy).

[0152] Liposome delivery

[0153] The compositions described herein are advantageously delivered to the lungs to provide the compound at the site of actual or potential influenza infection. This can be achieved via transpulmonary delivery through a pre-dose inhaler or other transpulmonary delivery device, and also by bringing the particles into the microvascular bed surrounding the alveoli in the lungs.

[0154] Nanocarriers, including smaller monolayer vesicles (such as liposomes), exhibit several advantages over other conventional methods for drug delivery to the lungs, including prolonged drug release and cell-specific targeted drug delivery. Nanoscale drug carriers can also facilitate the delivery of poorly water-soluble drugs, and some of the compounds described herein are poorly water-soluble. Additional advantages include their ability to provide controlled release, protection against metabolism and degradation, reduced drug toxicity, and improved targeting ability.

[0155] Liposomes (preferably monolayer vesicles) are characterized by a size of less than 200 nm as measured by dynamic light scattering, and are preferably characterized by being composed of chemically pure synthetic phospholipids, most preferably having side chains of at least 16 carbons in length, and containing one or more of the compounds described herein or a pharmaceutically acceptable salt sufficient to preferentially deliver (i.e., target) a certain amount of such compounds to the microvascular bed surrounding the alveoli. Vesicle diameter can be measured, for example, by dynamic light scattering using a helium-neon 100mW NEC gas laser and a Malvern K7027 correlator, ideally generating at least two or three measurements each time for size determination.

[0156] The term "chemically pure phospholipid" is intended to define phospholipids that are substantially free of harmful purification components and impurities that cause aggregation of smaller monolayer vesicles (SUVs) formed therefrom, and have a purity exceeding 97%. Preferably, the liposomes have a diameter predominantly between about 50 and about 160 nm, are substantially neutral in charge, and incorporate side chains with a length of 16 to 18 carbon atoms. More preferably, the liposomes are prepared from distearate phosphatidylcholine (DSPC) and include cholesterol as a vesicle stabilizer (most preferably in an amount of 10% to 50% of the total lipids).

[0157] Liposomes having a melting point above body temperature (i.e., above 37°C) is also advantageous. For this reason, the use of pure phospholipids, preferably saturated phospholipids having a carbon chain length of at least 16 carbons, preferably between 16 and 18 carbons, is advantageous. Distearylphosphatidylcholine (DSPC) is a preferred phospholipid. Cholesterol helps stabilize the liposomes and is preferably added in an amount sufficient to provide liposome stability. Most preferably, the liposomes further comprise polyethylene glycol-modified phospholipids, such as DSPEPEG. The method involves introducing a quantity of liposomes into the bloodstream of a patient, the liposomes being less than 200 nm in size (preferably monolayer vesicles) and preferably characterized by comprising chemically pure synthetic phospholipids, most preferably having side chains of at least 16 carbons in length, and containing a compound described herein or a pharmaceutically acceptable salt or prodrug sufficient to preferentially deliver (i.e., target) a quantity of the compound to the microvascular bed surrounding the alveoli in the lungs.

[0158] The compounds described herein can be combined with other anti-influenza agents as also described herein. Such additional agents may also be present in the liposomes, may be present in different liposomes, or may be administered via different routes.

[0159] Liposomes comprise one or more of the compounds described herein or pharmaceutically acceptable salts thereof, and may optionally include other anti-influenza agents. Liposomes can be prepared by dissolving phospholipids and cholesterol in a suitable organic solvent, such as chloroform, and evaporating the solvent to form a lipid membrane. If an ion carrier is used to load the compounds described herein into the liposomes, the ion carrier may be added to the lipid solution prior to evaporation. The dried lipid membrane is then rehydrated in a suitable aqueous phase, such as phosphate-buffered saline or other physiologically suitable solution. Water-soluble drugs or therapeutic agents may be contained in the hydrated solution, but if long-distance loading is required, a loading agent, such as a chelating agent described above, may be added to the hydrated solution to encapsulate it within the internal water space of the liposome.

[0160] Upon addition of a hydration solution, liposomes of varying sizes spontaneously form and encapsulate a portion of the aqueous phase. Subsequently, the liposomes and the aqueous suspension are subjected to, for example, shear forces from compression, sonication, or homogenization as described in U.S. Patent No. 4,753,788, to produce vesicles of specific sizes.

[0161] The liposomes can then be processed to remove unwanted compounds, such as unencapsulated drugs, from the suspension, which can be achieved through processes such as gel chromatography or ultrafiltration.

[0162] The use of liposomes in dry powder aerosols for targeted lung delivery is described, for example, in Willis et al., Lung, June 2012, 190(3):251-262.

[0163] Administration method

[0164] Depending on the severity of the infection being treated, the compounds and pharmaceutically acceptable compositions described above may be administered to humans and other animals orally, rectally, non-intestinally, intracerebrospinally, intravaginally, intraperitoneally, topically (as a powder, ointment or drops), buccally, as an oral or nasal spray, to the pulmonary system (e.g., by using an inhaler, such as a dose-dependent inhaler (MDI)) or similar routes.

[0165] Liquid dosage forms for oral administration include (but are not limited to) pharmaceutically acceptable emulsions, microemulsions, solutions, suspensions, syrups, and elixirs. In addition to the active compound, liquid dosage forms may contain inert diluents (e.g., water or other solvents) commonly used in the field, solubilizers and emulsifiers (e.g., ethanol, isopropanol, ethyl carbonate, EtOAc, benzyl alcohol, methyl benzoate, propylene glycol, 1,3-butanediol, dimethylformamide), oils (especially cottonseed oil, peanut oil, corn oil, germ oil, olive oil, castor oil, and sesame oil), glycerin, tetrahydrofurfuryl alcohol, polyethylene glycol, and fatty acid esters and mixtures thereof of sorbitol. In addition to inert diluents, oral compositions may also include adjuvants such as wetting agents, emulsifiers and suspending agents, sweeteners, flavoring agents, and aromatizers.

[0166] Injectable formulations (e.g., sterile injectable aqueous or oily suspensions) can be formulated using suitable dispersants, wetting agents, and / or suspending agents according to known techniques. Sterile injectable formulations can also be sterile injectable solutions, suspensions, or emulsions in non-toxic, non-enteric-acceptable diluents or solvents, such as solutions in 1,3-butanediol. Among acceptable mediators and solvents, water, Ringer's solution, USP, and isotonic sodium chloride solution can be used. Furthermore, sterile non-volatile oils are routinely used as solvents or suspension media. For this purpose, any mild non-volatile oil can be used, including synthetic monoglycerides or diglycerides. Additionally, fatty acids (e.g., oleic acid) are used in the preparation of injectable formulations.

[0167] Injectable formulations can be sterilized, for example, by filtration through a bacterial trapping filter or by incorporating a sterilizing agent in the form of a sterile solid composition that can be dissolved or dispersed in sterile water or other sterile injectable media before use.

[0168] To prolong the effects of the compounds described herein, it is generally desirable to slow the absorption of compounds administered subcutaneously or intramuscularly. This can be achieved by using liquid suspensions of poorly water-soluble crystalline or amorphous substances. The absorption rate of the compound depends on its solubility, which in turn may depend on crystal size and crystal form. Alternatively, delayed absorption of compounds administered non-enterogestally can be achieved by dissolving or suspending the compound in an oil-based medium. Injectable accumulation formulations are manufactured by forming microcapsule matrices of the compound within a biodegradable polymer, such as polylactic-co-glycolic acid. The release rate of the compound can be controlled depending on the compound-to-polymer ratio and the properties of the specific polymer used. Examples of other biodegradable polymers include poly(orthoester) and poly(anhydride). Injectable accumulation formulations can also be prepared by encapsulating the compound in tissue-compatible liposomes or microemulsions.

[0169] Compositions intended 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 suppository waxes, which are solid at ambient temperature but liquid at body temperature and thus melt in the rectal or vaginal cavity to release the active compound.

[0170] 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 (e.g., sodium citrate or dicalcium phosphate) and / or the following: a) fillers or extenders, such as starch, lactose, sucrose, glucose, mannitol, and silica; b) binders, such as carboxymethyl cellulose, alginate, gelatin, polyvinylpyrrolidone, sucrose, and gum arabic; c) humectants, such as glycerin; d) disintegrants, such as agar-agar, calcium carbonate, potato or cassava starch, alginate, certain silicates, and sodium carbonate; e) solution blockers, such as paraffin; f) absorption accelerators, such as quaternary ammonium compounds; g) wetting agents, such as cetyl alcohol and glycerol monostearate; h) adsorbents, such as kaolin and bentonite; and i) lubricants, such as talc, calcium stearate, magnesium stearate, solid polyethylene glycol, sodium lauryl sulfate, and mixtures thereof. In the case of capsules, tablets, and pills, the dosage form may also include a buffer.

[0171] Similar types of solid compositions can also be used as fillers in soft-filled and hard-filled gelatin capsules, which use excipients such as lactose / milk sugar and high molecular weight polyethylene glycol and the like. Solid dosage forms such as tablets, sugar-coated pills, capsules, pellets, and granules can be prepared with coatings and shells, such as enteric coatings and other coatings well known in pharmaceutical formulation techniques. They may optionally contain emulsifiers and may also have compositions that release or optionally delay the release of the active ingredient only or preferentially in a portion of the intestine. Examples of encapsulation compositions that can be used include polymeric substances and waxes. Similar types of solid compositions can also be used as fillers in soft-filled and hard-filled gelatin capsules, which use excipients such as lactose / milk sugar and high molecular weight polyethylene glycol and the like.

[0172] The active compound may also be in a microencapsulated form having one or more excipients as mentioned above. Solid dosage forms such as tablets, sugar-coated pills, capsules, pellets, and granules may be prepared with coatings and shells, such as enteric coatings, release-controlled coatings, and other coatings well known in pharmaceutical formulation techniques. In these solid dosage forms, the active compound may be mixed with at least one inert diluent (e.g., sucrose, lactose, or starch). As is common practice, such dosage forms may also include substances other than inert diluents, such as tablet-making lubricants and other tablet-making aids, such as magnesium stearate and microcrystalline cellulose. In the case of capsules, tablets, and pellets, the dosage form may also contain a buffer. It may optionally contain an emulsifier and may also have a composition that releases the active ingredient only or preferentially in a portion of the intestine or optionally in a delayed manner. Examples of encapsulation compositions that can be used include polymers and waxes.

[0173] Dosage forms for topical or transdermal administration of the compounds described herein include ointments, pastes, creams, lotions, gels, powders, solutions, sprays, inhalers, or patches. The active ingredient is mixed under sterile conditions with a pharmaceutically acceptable carrier and any desired preservatives or buffers, if applicable. Ocular formulations, ear drops, and eye drops that fall within the scope of this disclosure are also covered. Additionally, this disclosure covers the use of transdermal patches, which offer the added advantage of controlled delivery of the compound to the body. Such dosage forms can be prepared by dissolving or dispensing the compound in a suitable medium. Absorption enhancers may also be used to increase the flux of the compound across the skin. The rate can be controlled by providing a rate-controlled membrane or by dispersing the compound in a polymer matrix or gel.

[0174] The compositions described herein may be administered orally, non-enterically, by inhalation spray, topically, rectally, nasally, buccally, vaginally, or via an implantable reservoir. As used herein, the term "non-enterically" includes (but is not limited to) subcutaneous, intravenous, intramuscular, intra-articular, intrasynovial, intrasternal, intrathecal, intrahepatic, intralesional, and intracranial injection or infusion techniques. Specifically, the compositions are administered orally, intraperitoneally, or intravenously.

[0175] The sterile injectable forms of the compositions described herein may be aqueous or oily suspensions. These suspensions may be formulated using suitable dispersants or wetting agents and suspending agents according to techniques known in the art. Sterile injectable formulations may also be sterile injectable solutions or suspensions in nontoxic, non-enteric-acceptable diluents or solvents, such as solutions in 1,3-butanediol. Among acceptable mediators and solvents, water, Ringer's solution, and isotonic sodium chloride solution may be used. Furthermore, sterile, non-volatile oils are routinely used as solvents or suspension media. For this purpose, any mild, non-volatile oil may be used, including synthetic monoglycerides or diglycerides. Fatty acids (e.g., oleic acid and its glycerol derivatives) are suitable for the preparation of injectable formulations, as are pharmaceutically acceptable natural oils (e.g., olive oil or castor oil, especially their polyoxyethylene forms). These oil solutions or suspensions may also contain long-chain alcohol diluents or dispersants, such as carboxymethyl cellulose or similar dispersants commonly used to formulate pharmaceutically acceptable dosage forms (including emulsions and suspensions). Other commonly used surfactants (such as Tween, Span, and other emulsifiers) or bioavailability enhancers commonly used in the manufacture of pharmaceutically acceptable solid, liquid, or other dosage forms may also be used for formulation purposes.

[0176] The pharmaceutical compositions described herein can be administered orally 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, common carriers include (but are not limited to) lactose and corn starch. Lubricants, such as magnesium stearate, are often added. For oral administration in capsule form, suitable diluents include lactose and dried corn starch. When an aqueous suspension is required for oral use, the active ingredient is combined with an emulsifier and a suspending agent. Certain sweeteners, flavoring agents, or coloring agents may also be added if desired.

[0177] Alternatively, the pharmaceutical compositions described herein can be administered in the form of suppositories for rectal administration. These suppositories can be prepared by mixing the reagent with a suitable non-irritating excipient that is solid at room temperature but liquid at rectal temperature and thus melts in the rectum to release the drug. Such materials include (but are not limited to) cocoa butter, beeswax, and polyethylene glycol.

[0178] The pharmaceutical compositions described herein can also be administered topically, especially when the therapeutic target includes areas or organs easily accessible for topical application, such as diseases of the eye, skin, or lower intestine. Suitable topical formulations for each of these areas or organs are readily prepared.

[0179] Topical application for the lower intestine can be achieved via rectal suppository formulation (see above) or in a suitable enema formulation. Topical percutaneous patches may also be used.

[0180] For topical application, pharmaceutical compositions may be formulated in a suitable ointment form containing an active ingredient suspended or dissolved in one or more carriers. Carriers for topical administration of the compounds described herein include, but are not limited to, mineral oil, liquid petrolatum, white petrolatum, propylene glycol, polyoxyethylene, polyoxypropylene compounds, emulsified waxes, and water. Alternatively, pharmaceutical compositions may be formulated in a suitable lotion or cream form containing an active ingredient 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 wax, cetearyl alcohol, 2-octyldodecyl alcohol, benzyl alcohol, and water.

[0181] For ocular use, the pharmaceutical composition may be formulated as a micronized suspension in isotonic, pH-adjusted sterile saline, with or without a preservative (e.g., benzalkonium chloride), or specifically as a solution in isotonic, pH-adjusted sterile saline. Alternatively, for ocular use, the pharmaceutical composition may be formulated in an ointment such as petrolatum.

[0182] The compounds used in the methods described herein can be formulated in unit dosage forms. The term "unit dosage form" refers to a physically discrete unit suitable for administration to a treated individual in unit dose form, wherein each unit contains a predetermined amount of active substance calculated to produce the desired therapeutic effect, optionally combined with a suitable pharmaceutical carrier. Unit dosage forms can be used for a single daily dose or multiple daily doses (e.g., about 1 to 4 or more times per day). When multiple daily doses are used, the unit dosage form used for each dose may be the same or different.

[0183] This disclosure is intended to provide a more comprehensive understanding of the examples described herein, which detail exemplary embodiments. However, these examples should not be construed as limiting the scope of this disclosure. All references throughout this disclosure are expressly incorporated herein by reference.

[0184] Example

[0185] Example 1 - Analysis of Cytopathic Effects

[0186] The study was conducted to evaluate the in vitro combination effect of the test compound 3-(2-(5-chloro-1H-pyrrolo[2,3-b]pyridin-3-yl)-5-fluoro-7H-pyrrolo[2,3-d]pyrimidin-7-yl)bicyclo[2.2.2]octane-2-carboxylic acid with other influenza antiviral agents in an in vitro cytopathic effect (CPE) assay based on influenza virus cells.

[0187] Influenza virus type A / PR / 8 / 34 (H1N1) (ATCC VR-1469) was obtained from ATCC. MDCK cells (ATCC CCL-34) were obtained from ATCC and maintained in minimally essential Eagle medium (Sigma M2279) supplemented with 10% fetal bovine serum (Corning R35-076-CV), 1% L-glutamic acid (Gibco 25030081), 1% non-essential amino acids (NEAA, Gibco 11140050), and 1% penicillin-streptomycin (PS, HycloneSV30010).

[0188] The test medium used for MDCK cells was OptiPROSFM medium (Gibco 12309019) supplemented with 1% L-glutamic acid, 1% NEAA and 1% PS, and 2.5 μg / mL trypsin was used as the test medium for MDCK cells.

[0189] In 384-well plates, MDCK cells were seeded at 2,000 cells / well and cultured overnight at 37°C and 5% CO2. The next day, each of the two drug combination pairs was tested using a checkerboard crossover pattern of seven drug concentrations of each compound, including three replicates of each compound individually. Pimodivir (VX-787) was evaluated in parallel as a control compound. The compounds (in DMSO) were seeded using a Tecan HP D300 digital dispenser. Virus (2 TCID50) was then added. 90 / well). The compound was tested at concentrations of 0.125, 0.25, 0.5, 1, 2, 4 and 8 × EC. 50 Values. Compound combinations are listed in Table 4. The final concentration of DMSO (Sigma 34869) in the cell culture medium was 0.5%. The resulting cultures were incubated at 37°C and 5% CO2 for another 5 days until the virus infection in the virus control showed obvious CPE (as assessed by a reduction in viable cell count). To assess cell viability, CCK-8 kit solution (one vial of colorimetric system, Biolite 35004) was subsequently added to each well and the cells were incubated at 37°C for 3 hours. Absorbance (460 nm) was measured using a SpectraMax 340PC384 disc reader (molecular device).

[0190] The antiviral activity and cytotoxicity of the compound combination are expressed as inhibition % and survival % respectively, and are calculated using the following equation:

[0191] Inhibition (%) = (Original Data) cpd -average VC ) / (average CC -average VC )×100

[0192] Survival (%) = (Original Data) cpd -average MC ) / (average CC -average MC )×100

[0193] Raw data cpd Indicates the absorbance value of the pores treated with the compound; average VC ,average CC and average MC The values ​​represent the mean absorbance values ​​for the virus control (VC; virus-infected cells, no compound), cell control (CC; cells without virus or compound), and culture medium control (MC; culture medium only). Data were obtained using MacSynergy. TM II software (Prichard and Shipman, 1990) was used to calculate the combination index. A synergistic effect curve (95%) was then calculated. Positive combination index values ​​indicate synergistic effects, while negative combination index values ​​indicate antagonistic effects.

[0194] As shown in the table below, 3-(2-(5-chloro-1H-pyrrolo[2,3-b]pyridin-3-yl)-5-fluoro-7H-pyrrolo[2,3-d]pyrimidin-7-yl)bicyclo[2.2.2]octane-2-carboxylic acid exhibits a strong synergistic effect with influenza antiviral drugs (including baloxavir (without the hydroxyl form of baloxavir masocylate prodrug), oseltamivir, and favipiravir) in an antiviral CPE analysis targeting IFV A / PR / 8 / 34 (H1N1).

[0195]

[0196] An absolute combination index value <25 (i.e., 95% synergistic effect) indicates a cumulative effect. An absolute combination index value of 25 to 50 indicates a slight synergistic or antagonistic effect. An absolute combination index value of 50 to 100 indicates a moderate synergistic or antagonistic effect. An absolute combination index value >100 indicates a strong synergistic or antagonistic effect.

[0197] Figures 1 to 3 Provides visual representations of the calculation curves for each of the three test combinations.

[0198] Compound 1, along with the same three combinations of baloxavir (without the hydroxyl form of baloxavir masocylate prodrug), oseltamivir, and favipiravir, also exhibited antiviral activity against influenza B.

Claims

1. (1) 3-(2-(5-chloro-1H-pyrrolo[2,3-b]pyridin-3-yl)-5-fluoro-7H-pyrrolo[2,3-d]pyrimidin-7-yl)bicyclo[2.2.2]octane-2-carboxylic acid or a pharmaceutically acceptable salt thereof, and (2) the use of a second antiviral agent in the preparation of a medicament for the treatment or prevention of influenza, the second antiviral agent being selected from baloxavir, baloxavir macozide, oseltamivir, oseltamiviric acid, favipiravir, or a pharmaceutically acceptable salt thereof.

2. The use according to claim 1, wherein the second antiviral agent is oseltamivir, oseltamivir acid, or a pharmaceutically acceptable salt thereof.

3. The use according to claim 1, wherein the second antiviral agent is baloxavir macozide, baloxavir, or a pharmaceutically acceptable salt thereof.

4. The use according to claim 1, wherein the second antiviral agent is favipiravir or a pharmaceutically acceptable salt thereof.

5. The use according to any one of claims 1 to 4, wherein the 3-(2-(5-chloro-1H-pyrrolo[2,3-b]pyridin-3-yl)-5-fluoro-7H-pyrrolo[2,3-d]pyrimidin-7-yl)bicyclo[2.2.2]octane-2-carboxylic acid or a pharmaceutically acceptable salt thereof is administered prior to the second antiviral agent.

6. The use according to any one of claims 1 to 4, wherein the 3-(2-(5-chloro-1H-pyrrolo[2,3-b]pyridin-3-yl)-5-fluoro-7H-pyrrolo[2,3-d]pyrimidin-7-yl)bicyclo[2.2.2]octane-2-carboxylic acid or a pharmaceutically acceptable salt thereof is administered after the second antiviral agent.

7. The use according to any one of claims 1 to 4, wherein the 3-(2-(5-chloro-1H-pyrrolo[2,3-b]pyridin-3-yl)-5-fluoro-7H-pyrrolo[2,3-d]pyrimidin-7-yl)bicyclo[2.2.2]octane-2-carboxylic acid or a pharmaceutically acceptable salt thereof is administered simultaneously with the second antiviral agent.

8. The use according to claim 7, wherein the 3-(2-(5-chloro-1H-pyrrolo[2,3-b]pyridin-3-yl)-5-fluoro-7H-pyrrolo[2,3-d]pyrimidin-7-yl)bicyclo[2.2.2]octane-2-carboxylic acid or a pharmaceutically acceptable salt thereof is co-formulated with the second antiviral agent.

9. The use according to claim 7, wherein the 3-(2-(5-chloro-1H-pyrrolo[2,3-b]pyridin-3-yl)-5-fluoro-7H-pyrrolo[2,3-d]pyrimidin-7-yl)bicyclo[2.2.2]octane-2-carboxylic acid or a pharmaceutically acceptable salt thereof is formulated separately with the second antiviral agent.

10. A combination comprising 3-(2-(5-chloro-1H-pyrrolo[2,3-b]pyridin-3-yl)-5-fluoro-7H-pyrrolo[2,3-d]pyrimidin-7-yl)bicyclo[2.2.2]octane-2-carboxylic acid or a pharmaceutically acceptable salt thereof, and a second antiviral agent selected from baloxavir, baloxavir macozide, oseltamivir, oseltamiviric acid, favipiravir, or a pharmaceutically acceptable salt thereof.

11. The combination according to claim 10, wherein the second antiviral agent is oseltamivir, oseltamivir acid, or a pharmaceutically acceptable salt thereof.

12. The combination according to claim 10, wherein the second antiviral agent is baloxavir macozide, baloxavir, or a pharmaceutically acceptable salt thereof.

13. The combination according to claim 10, wherein the second antiviral agent is favipiravir or a pharmaceutically acceptable salt thereof.

14. A combination comprising a) a therapeutically effective amount of 3-(2-(5-chloro-1H-pyrrolo[2,3-b]pyridin-3-yl)-5-fluoro-7H-pyrrolo[2,3-d]pyrimidin-7-yl)bicyclo[2.2.2]octane-2-carboxylic acid or a pharmaceutically acceptable salt thereof and b) a therapeutically effective amount of a second antiviral agent selected from baloxavir, baloxavir macozide, oseltamivir, oseltamiviric acid, favipiravir, or a pharmaceutically acceptable salt thereof.

15. The combination according to claim 14, wherein the second antiviral agent is oseltamivir, oseltamivir acid, or a pharmaceutically acceptable salt thereof.

16. The combination according to claim 14, wherein the second antiviral agent is baloxavir macozide, baloxavir, or a pharmaceutically acceptable salt thereof.

17. The combination according to claim 14, wherein the second antiviral agent is favipiravir or a pharmaceutically acceptable salt thereof.

18. Use of the combination according to any one of claims 10 to 17 in the preparation of a medicament for treating or preventing influenza virus infection.

19. The use according to claim 18, wherein the influenza is a pandemic or drug-resistant pandemic / seasonal influenza.

20. Use of the combination according to any one of claims 10 to 17 in the preparation of a medicament for inhibiting the endonuclease activity of influenza polymerase in influenza A or B viruses.

21. The use according to claim 20, wherein the drug further comprises a therapeutically effective amount of a third antiviral agent.

22. The use according to claim 20, wherein the drug is suitable for administration with an influenza vaccine before, after, or simultaneously with the combination.

23. The use according to claim 20, wherein the influenza virus is a pandemic or drug-resistant pandemic / seasonal influenza virus.

24. Use of a therapeutic amount of the combination of any one of claims 10 to 17 in the preparation of a medicament for treating or preventing infection with influenza A or influenza B in a host.

25. The use according to claim 24, wherein the drug further comprises a therapeutically effective amount of a third antiviral agent.

26. The use according to claim 24, wherein the drug is suitable for administration with an influenza vaccine before, after, or simultaneously with the combination.

27. The use according to claim 24, wherein the influenza virus is a pandemic or drug-resistant pandemic / seasonal influenza virus.

28. Use of a therapeutic amount of the combination according to any one of claims 10 to 17 in the preparation of a medicament for reducing the endonuclease activity of influenza polymerase in influenza A or influenza B in a host.

29. The use according to claim 28, wherein the drug further comprises a therapeutically effective amount of a third antiviral agent.

30. The use according to claim 28, wherein the drug is suitable for administration with an influenza vaccine before, after, or simultaneously with the combination.

31. The use according to claim 28, wherein the influenza virus is a pandemic or drug-resistant pandemic / seasonal influenza virus.

32. Use of a therapeutic amount of the combination of any one of claims 10 to 17 in the preparation of a medicament for reducing the replication of influenza virus in a host.

33. The use according to claim 32, wherein the medicament further comprises a therapeutically effective amount of a third antiviral agent.

34. The use according to claim 32, wherein the drug is suitable for administration before, after, or simultaneously with the combination.

35. The use according to claim 32, wherein the influenza virus is a pandemic or drug-resistant pandemic / seasonal influenza virus.

36. Use of a combination according to any one of claims 10 to 17 in the preparation of a medicament for treating influenza A or influenza B virus infection.

37. A pharmaceutical composition for treating or preventing influenza virus infection or replication in a patient, comprising 3-(2-(5-chloro-1H-pyrrolo[2,3-b]pyridin-3-yl)-5-fluoro-7H-pyrrolo[2,3-d]pyrimidin-7-yl)bicyclo[2.2.2]octane-2-carboxylic acid or a pharmaceutically acceptable salt thereof, wherein the composition is administered in combination with a second antiviral agent selected from baloxavir, baloxavir macozide, oseltamivir, oseltamiviric acid, favipiravir, or a pharmaceutically acceptable salt thereof.

38. A combination comprising a) 3-(2-(5-chloro-1H-pyrrolo[2,3-b]pyridin-3-yl)-5-fluoro-7H-pyrrolo[2,3-d]pyrimidin-7-yl)bicyclo[2.2.2]octane-2-carboxylic acid or a pharmaceutically acceptable salt thereof, b) baloxavir, baloxavir macozylate, or a pharmaceutically acceptable salt thereof, and c) a neuraminidase inhibitor.

39. The combination according to claim 38, wherein the neuraminidase inhibitor is oseltamivir, oseltamivir acid, zanamivir, lanamivir, peramivir, or a pharmaceutically acceptable salt thereof.

40. The combination according to claim 39, wherein the neuraminidase inhibitor is oseltamivir, oseltamivir acid, or a pharmaceutically acceptable salt thereof.

Images