Bicacinabine Dosage Regimen

Abstract

The present invention relates to (3S)-1-[5-tert-butyl-3-[(1-methyltetrazol-5-yl)methyl]triazolo[4,5-d]pyrimidin-7-yl]pyrrolidin-3-ol for use in the treatment of diabetic retinopathy and methods of treatment thereof.

Description

[Background technology]

[0001] background Diabetes mellitus affects 463 million people worldwide, and its prevalence is expected to increase to 700 million by 2045 (IDF 2019; Nair et al. 2016). Diabetic retinopathy (DR) affects approximately one-third of patients with type 1 and type 2 diabetes (Yau et al. 2012). Chronic hyperglycemia causes retinal microvascular changes, inflammation, and neurodegeneration (Fong et al. 2004). At any stage of DR, patients can develop diabetic macular edema (DME), the most common cause of central vision loss in patients with diabetes (Resnikoff et al. 2004). DME, along with the progression of DR (i.e., proliferative diabetic retinopathy [PDR]), can impair vision and lead to blindness (Leasher et al. 2016). Overall, DR is estimated to be the most frequent cause of new cases of blindness among working-age adults worldwide (American Diabetes Association 2002; Yau et al. 2012).

[0002] Unlike other parts of the world, patients in the United States with diabetic retinopathy (DR) can be treated with intravitreal (IVT) injections of anti-vascular endothelial growth factor (VEGF) (Lucentis USPI; Eylea USPI), regardless of disease severity or the presence of visual impairment, PDR, or DME. However, clinical practice trends indicate that a high proportion of patients with nonproliferative diabetic retinopathy (NPDR) and DME resulting in minimal visual impairment are not readily treated (Cantrell et al. 2020). Instead, treatment initiation is postponed until the disease progresses, causing further damage to the retinal anatomy and worsening vision. Patients with severe NPDR do not yet have clear visual impairment, but the underlying retinal pathology is progressive. If left untreated, moderate to severe NPDR can progress to sight-threatening disease in approximately 41% and 58% of patients within one and two years, respectively (Regeneron Pharmaceuticals, Inc. 2020). Therefore, there remains an unmet need for the development of novel therapies that provide non-invasive treatment options for severe NPDR and prevent progression to visual impairment.

[0003] Chronic hyperglycemia can cause DR or DME through pathological mechanisms that are poorly understood but appear to involve four major pathways: the polyol pathway, the advanced glycation end products pathway, the protein kinase C pathway, and the hexosamine pathway (Brownlee 2005). All of these pathways result in increased oxidative stress, inflammation, vascular dysfunction, and neurodegeneration (Fong et al. 2004). Müller cells, microglia, and endothelial cells produce chemokines that induce leukostasis, extravasation, the influx of systemic immune cells such as monocytes into the retina, and increased production of cytokines, including VEGF, tumor necrosis factor-α, interleukin (IL)-1β, IL-6, metalloproteinases, and angiopoietin-2. These inflammatory mediators then lead to the breakdown of endothelial cell-cell junctions that form the blood-retinal barrier. Retinal microglia are activated early in the disease process, resulting in increased apoptosis of ganglion cells and amacrine cells. In retinal capillaries, pericyte shedding and basement membrane thickening also occur as a result of hyperglycemia, all of which contribute to increased vascular leakage. The above factors lead to vascular pathology that ultimately causes vision loss (Das et al. 2015).

[0004] DR can be broadly classified into two stages based on the level of microvascular degeneration and associated ischemic damage: (a) non-specific retinal dysplasia (NPDR), characterized by microaneurysms and possible intraretinal hemorrhage, hard exudates, and cotton-wool patches; and (b) progressive retinal dysplasia (PDR), characterized by retinal neovascularization, iris neovascularization (NVI), and vitreous hemorrhage (Wilkinson et al. 2003). Another complication of DR is DME, which can occur across all DR severity levels and is the most common cause of vision loss in patients with DR. DME results from diabetes-induced breakdown of the blood-retinal barrier, resulting in vascular leakage of fluid and circulating proteins into the neural retina. This leads to abnormal retinal thickening and often cystoid edema of the macula (Frey and Antonetti 2011; Zhang et al. 2014; Stitt et al. 2016). Hyperglycemic conditions also cause retinal ischemia, inducing vascular leakage, neovascularization, and increased VEGF production, which contributes to PDR.

[0005] Current Treatments and Unmet Medical Needs Intraocular treatment modalities for diabetic eye disease include laser photocoagulation and intravenous steroid (IVT) anti-VEGF therapy. Current treatment paradigms, including laser photocoagulation and / or IVT anti-VEGF therapy, focus on treating progressive disease once PDR or DME develops. Anti-VEGF agents are approved in the United States for the treatment of all forms of DR (Lucentis USPI; Eylea® USPI) and elsewhere for the treatment of DME and PDR (Lucentis SmPC), and constitute the first-line therapy for most eyes with DME involving the fovea. IVT injections of steroids can also be effective in treating DME (Campochiaro et al. 2012; Boyer et al. 2014), but the use of IVT steroids is usually limited by more frequent ocular side effects, such as cataracts and glaucoma. According to the 2017 Guidelines for Diabetic Eye Care from the International Council of Ophthalmology, NPDR warrants close monitoring and laser photocoagulation can be considered, but the use of anti-VEGF therapy is not mentioned (Wong et al. 2018). Despite widespread labeling of anti-VEGF therapy for DR in the United States (Lucentis USPI; Eylea® USPI), clinical practice trends indicate that a high percentage of patients with NPDR, with or without DME, do not readily undergo treatment despite minimal visual impairment (PAT Survey; Cantrell et al. 2020). Instead, treatment initiation is postponed until the disease progresses, causing further damage to the retinal anatomy and worsening vision. Several factors may drive the deferral of treatment for moderately severe to severe NPDR in current practice and could potentially be addressed with oral RO6868847. These include: Invasiveness of available treatments: All currently available ocular treatments for DR / DME (e.g., laser, IVT injections) require invasive procedures and are associated with infrequent but serious risks. Furthermore, patients report anxiety regarding the initiation of IVT injection procedures (Senra et al. 2017). Given that DM is a systemic condition and DR is usually bilateral, the sponsor is developing RO6868847 for oral administration, thus offering a non-invasive alternative to IVT injection procedures and associated risks to improve patient experience. Higher frequency of treatments / visits with IVT therapy: Initiating anti-VEGF therapy for DR / DME means more frequent treatments and / or follow-up visits to establish and maintain benefit, at least in the first year (Lally et al. 2016). As an oral medication, RO6868847 can be self-administered continuously, and its effects are not subject to the inclusion and exclusion of IVT medications. Therefore, the interval between follow-up visits may be longer with RO6868847 than with IVT anti-VEGF therapy, potentially reducing the burden on patients and the healthcare system. Low perceived urgency to treat: Compared to neovascular age-related macular degeneration (AMD), DR is perceived as having slower disease progression and relatively more tolerable disease progression, leading to the mistaken belief that the retinal damage associated with disease progression is largely reversible. Extrapolation from pivotal trials using anti-VEGF therapy for DME (Brown et al. 2013; Schmidt-Erfurth et al. 2014; Heier et al. 2016) has shown that deferred therapy can result in retinal thickness reduction (structural benefit) comparable to immediate therapy, but delayed anti-VEGF therapy appears to reduce the likelihood of visual recovery (functional benefit). There is consensus that earlier detection and treatment of DR / DME can minimize or prevent both disease progression and the risk of vision loss (Nair et al. 2016). Lack of consensus on how to treat: In contrast to DME, there is no clear consensus on the management of patients with DR and good visual acuity (BCVA >73 letters) with anti-VEGF therapy, as demonstrated by the 2018 PAT survey (Singh 2018). New evidence from the PANORAMA trial (Regeneron Pharmaceuticals, Inc. 2020) was recently published, suggesting a treatment protocol of bimonthly injections of aflibercept (followed by 5 months of monthly treatments) to improve DR. Furthermore, the clinical trial Protocol W from the Diabetic Retinopathy Clinical Research Network (Protocol W) demonstrated that anti-VEGF therapy can prevent the development of sight-threatening complications in eyes at high risk for PDR.

[0006] Although patients with moderate to severe NPDR do not have obvious visual impairment, the underlying retinal pathology is progressive (Morello 2007). If left untreated, moderate to severe NPDR can progress to sight-threatening disease in approximately 41% of patients within one year and in 58% of patients within two years (Regeneron Pharmaceuticals, Inc. 2020). Therefore, there remains an unmet need for the development of novel therapies that offer earlier intervention and non-invasive treatment options before irreversible retinal damage occurs.

[0007] In summary, in clinical practice, patients presenting with NPDR are often not immediately treated with IVT anti-VEGF therapy for several possible reasons. Delaying DR therapy in these patients is likely to be disadvantageous due to their progressive disease. Many of the given reasons for postponing treatment with IVT anti-VEGF therapy could potentially be addressed by RO6868847, an oral small molecule agonist highly specific for cannabinoid receptor 2 (CB2). Oral small molecule agonists such as RO6868847 (also known as RG7774 or bicasinabin) may be appropriate for treating patients with all stages of NPDR and early stages of advanced complications of DR (PDR, DME), even though IVT anti-VEGF injections are not approved / covered or administered in clinical practice. DR is often bilateral, and based on its intended oral route, RO6868847 may benefit both eyes simultaneously. Numerous studies have shown that cannabinoids can suppress cytokine production in innate and adaptive immune responses in both animal models and human cell cultures (Klein 2005). The suppression of pro-inflammatory cytokine and chemokine production indicates that synthetic CB2 agonists may have anti-inflammatory effects and therefore can be used to treat chronic inflammatory diseases such as DR.

[0008] RO6868847 is a white to off-white powder that exhibits moderate to good solubility in organic solvents and slight solubility in aqueous solutions. RO6868847 is fairly stable to temperature stress and to light in the solid state and in solution. For Phase I, oral film-coated tablets containing 0.05, 0.75, 10, or 100 mg of RO6868847 drug substance were developed for clinical use. Three placebo film-coated tablets were formulated to match the size and appearance of the active film-coated tablets: one size matching the 0.05 mg and 0.75 mg active film-coated tablets, and one size each matching the 10 mg and 100 mg active film-coated tablets.

[0009] Rationale for RO6868847 Treatment RO6868847 is a highly selective CB2 agonist intended for oral treatment of all stages of NPDR and the early stages of advanced complications of DR (PDR, DME). CB2 is primarily expressed in immune cells, including retinal microglia. In the eye, CB2 activation by RO6868847 has been shown to inhibit leukocyte adhesion and microglial activation, reduce vascular permeability, and consequently maintain endothelial barrier function, resulting in an anti-inflammatory effect. Furthermore, RO6868847 is highly selective for CB2, thereby eliminating the psychotropic effects of central CB1 activation (Topol et al. 2010). Current data suggest that RO6868847 exhibits the highest selectivity compared to known CB2 agonists (Soethoudt et al. 2017). Oral administration may benefit patients with DR by achieving systemic and bilateral retinal exposure. Based on preclinical evidence, systemic exposure may target leukocytes to prevent their adhesion to retinal endothelial cells, while retinal exposure may activate CB2 receptors in microglia, preserving them in a non-activated, branched state. Furthermore, preclinical evidence suggests that systemic exposure may have benefits for other diabetic complications, such as diabetic nephropathy, which was shown to improve in the same preclinical model used for retinal microglial analysis (Zoja et al. 2016). In summary, due to the advantages of this non-invasive oral administration and its expected bilateral efficacy, physicians may more easily initiate treatment for all patients with NR, who currently receive only delayed IVT therapy. Furthermore, the use of a highly selective CB2 agonist is expected to lack CB1 modulation and its respective side effects. The development program for RO6868847 was initiated to provide a treatment option for patients with moderate to severe NPDR, for whom photocoagulation and intravitreal anti-VEGF therapy are typically postponed in current clinical practice.

[0010] A key challenge in selecting a dose range for clinical development of RO6868847 was the wide predicted pharmacologically active dose range in humans, ranging from 0.22 mg to 42 mg per day based on different nonclinical animal models. In addition to nonclinical in vivo data, we used CB2 receptor target engagement (TE) information from human whole blood to select the clinical dose range. Because in vivo monitoring of CB2R TE on ocular microglial cells is not feasible, we instead predicted CB2R TE by RO6868847 using observed plasma PK in humans in combination with derived affinity data based on peripheral ex vivo CB2R TE on B cells, a circulating immune cell type that expresses CB2 receptors. CB2R TE in the retina was estimated by assuming that the plasma concentration of RO6868847 reflects the concentration in the retina and that the binding affinity of RO6868847 to CB2 receptors on B cells assessed in human blood using an ex vivo target engagement assay represents binding to CB2 receptors within the retina. Surprisingly, in vitro experiments evaluating CB2R-TE in whole blood samples from elderly individuals and age- and demographically-matched diabetic and non-diabetic subjects revealed that approximately three-fold higher concentrations of RO6868847 were required to achieve similar CB2R-target engagement effects as observed in the blood of young, healthy participants.

[0011] In a Phase I single ascending dose (SAD) and multiple ascending dose (MAD) study (BP40387), CB2R-TE was measured in blood from healthy participants who received single and multiple 0.75-300 mg doses of RO6868847. Based on clinical RO6868847 pharmacokinetic (PK) and pharmacodynamic (PD; measured by CB2R-TE) data, accounting for differences between healthy and diabetic subjects, the lower end of the 0.75 mg-300 mg dose range is expected to achieve near half-maximal CB2R-TE, while the upper end of that dose range is expected to achieve maximum CB2R-TE. These doses are surprisingly higher than predicted based on nonclinical models.

[0012] In addition to the CB2R-TE whole blood assay, an in vitro LPS whole blood challenge assay was applied using blood from healthy participants who received multiple 300 mg doses of RO6868847 6 in the MAD study. Surprisingly, a signal for attenuation of LPS-induced cytokine release was observed only at the higher end of the dose range, suggesting additional pharmacological effects that support the use of doses providing exposure in the high exposure range.

[0013] In summary, the clinical dose range was established based on a combination of three data sets: preclinical biomarker readouts of CB2 for target engagement and effect on LPS-induced cytokine production in blood from healthy participants, elderly diabetic patients and age- and demographically matched controls, and finally clinical and biomarker data obtained in healthy participants in a Phase I clinical study. Summary of the Invention

[0014] Quick Overview A first aspect of the present invention relates to the compound (3S)-1-[5-tert-butyl-3-[(1-methyltetrazol-5-yl)methyl]triazolo[4,5-d]pyrimidin-7-yl]pyrrolidin-3-ol for use as an agonist of cannabinoid receptor 2 in a patient at a dose of 0.75 mg to 300 mg.

[0015] A second aspect of the present invention relates to a method for the treatment of a cannabinoid receptor 2 agonist, comprising administering to a patient, particularly a patient in need thereof, (3S)-1-[5-tert-butyl-3-[(1-methyltetrazol-5-yl)methyl]triazolo[4,5-d]pyrimidin-7-yl]pyrrolidin-3-ol in a dose of 0.75 mg to 300 mg.

[0016] A third aspect of the present invention relates to the use of the compound (3S)-1-[5-tert-butyl-3-[(1-methyltetrazol-5-yl)methyl]triazolo[4,5-d]pyrimidin-7-yl]pyrrolidin-3-ol in the manufacture of a medicament as an agonist of cannabinoid receptor 2 in a dose of 0.75 mg to 300 mg. [Brief explanation of the drawings]

[0017] [Figure 1] Figure 1 shows the ratio of CB2 surface expression on B cells to T cells in whole blood from healthy participants after exposure to increasing concentrations of RO6868847. [Figure 2] 1 shows the determination of IC50 values ​​for RO6868847 in B cells from diabetic patients and age-matched healthy participants. [Figure 3] Figure 1 shows IC50 determination for the effect of RO6868847 on G-CSF, IFNγ, IL1β and TNFα levels in an in vitro LPS whole blood challenge assay in healthy participants. [Figure 4] Figure 1 shows IC50 determination for the effect of RO6868847 on G-CSF, IFNγ, IL1β and TNFα levels in an in vitro LPS whole blood challenge assay in diabetic patients and matched healthy participants. [Figure 5] Shown is the mean (+SD) CB2 receptor target engagement (% change from baseline) on B cells versus time profile after 14 days of multiple oral doses of 20 mg to 300 mg of RO6868847 or placebo. [Figure 6] Individual median baseline normalized TNF-α levels versus plasma Cavg,ss for RO6868847 on day 14 are shown. [Figure 7] Individual median baseline normalized G-CSF levels versus plasma Cavg,ss for RO6868847 on day 14 are shown. [Figure 8] Individual median baseline normalized IL-1β levels versus plasma Cavg,ss for RO6868847 on day 14 are shown. [Figure 9] Individual median baseline normalized IFNγ levels versus plasma Cavg,ss for RO6868847 on day 14 are shown. [Figure 10] Figure 1 shows disease markers versus free exposure of the active S-epimer RO6868847 in several animal models. AUC = area under the concentration-time curve from 0 to 24 hours; cyno = cynomolgus monkey; GLP = Good Laboratory Practice; Ki = equilibrium dissociation constant; LCNV = laser-induced angiogenesis; LPS = lipopolysaccharide; NOAEL = no observed adverse effect level; OD = optical density; RO6868847 = bicasinabin; STZ = streptozotocin. Disease markers (hyperfluorescent area (mm²), OD retina-to-plasma fluorescein ratio, leukocyte count per mm², or total process length (mm²)) as a percentage of control values ​​(100% represents vehicle-treated diabetic controls, and 0% represents vehicle-treated healthy controls) in different in vivo PD models as a function of free AUC of the active S-epimer RO6868847. The rat Ki (33.3 nM = 12 ng / mL) was extrapolated to free AUC as follows: free AUC = 12'24 = 286 ng·hr / mL. The green area represents the estimated pharmacologically active exposure range in humans. DETAILED DESCRIPTION OF THE INVENTION

[0018] Unless otherwise stated, the following terms used in the specification and claims have the meanings indicated below.

[0019] "Active pharmaceutical ingredient" (or "API") refers to a compound or molecule in a pharmaceutical composition that has a specific biological activity.

[0020] "Chronic degenerative disease" refers to a disease in which the function and / or structure of the affected tissue or organ deteriorates over time. Examples of chronic degenerative diseases include, but are not limited to, neurodegenerative inflammatory disorders, atherosclerosis, liver fibrosis, or diabetic microvascular complications.

[0021] "Diabetes mellitus" or "DM" refers to various forms of glucose metabolic disorders with different etiologies and symptoms. A common feature is relative or absolute insulin deficiency. Diabetic mellitus is characterized by persistently elevated blood glucose levels (hyperglycemia) or improper utilization of supplied glucose. Diabetes mellitus is subdivided into type I (insulin-dependent; IDDM) and type II (non-insulin-dependent; NIDDM). Diabetes-specific and diabetes-related chronic complications include microangiopathy, such as retinopathy, nephropathy, and neuropathy, polyneuropathy, diabetic foot, skeletal, supporting, and connective tissue disorders, and macroangiopathy, particularly coronary heart disease, cerebrovascular disease, and peripheral arterial occlusive disease.

[0022] "Diabetic retinopathy" or DR refers to microvascular damage in the retina that occurs in diabetes mellitus. The forms of diabetic retinopathy include non-proliferative retinopathy (NPDR) (background retinopathy), such as retinal hemorrhage, microaneurysms, hard exudates, and retinal edema accompanied by vision loss, and proliferative retinopathy (PDR), which involves the additional development of cotton wool spots on and before the retina, accompanied by vitreous hemorrhage due to retinal ischemia from vascular occlusion, and neovascularization. Proliferative retinopathy can lead to tractional retinal detachment, neovascular glaucoma, and blindness. The term DR encompasses all stages of PDR and NPDR.

[0023] "Diabetic macular edema" or DME refers to swelling of the macula caused by retinal vascular leakage that occurs in patients with diabetes. DME is the leading cause of vision loss in people with diabetic retinopathy. People with diabetes have a 10 percent lifetime risk of developing DME. DME affects up to 30% of people who have had diabetes for more than 20 years. If left untreated, DME can lead to moderate to severe vision loss.

[0024] "Individual" or "subject" refer to mammals and are used interchangeably. Mammals include, but are not limited to, livestock animals (e.g., cows, sheep, cats, dogs, and horses), primates (e.g., humans and non-human primates such as monkeys), rabbits, and rodents (e.g., mice and rats). In certain embodiments, the individual or subject is a human.

[0025] "Diabetic microvascular complications" refers to diabetic retinopathy, nephropathy, and neuropathy that can lead to kidney failure, peripheral arterial disease, or limb amputation.

[0026] "Nonproliferative diabetic retinopathy" or "NPDR" refers to the early stage of diabetic retinopathy, characterized by edema and hard white spots in the central retina, lipids leaking from abnormal blood vessels, resulting in blurred central vision. It may also involve vascular blockage, restricting the blood supply to the retina, and increased macular edema.

[0027] "Patient" refers to a human (such as a male or female) in need of treatment with RO6868847.

[0028] Detailed Description All publications, patent applications, patents, and other references mentioned herein are incorporated by reference in their entirety.

[0029] The nomenclature used in this application is based on IUPAC systematic nomenclature unless otherwise indicated.

[0030] While various features and embodiments of the present invention are disclosed herein, other features, modifications, and equivalents of the present invention will be apparent to those of ordinary skill in the art based on the teachings provided. The invention as described is not limited to the examples and embodiments provided, and those of ordinary skill in the art will recognize various alternative equivalents. As used herein, the singular forms "a," "an," and "the" include the plural unless the context clearly dictates otherwise. For example, "an" individual also includes "individuals."

[0031] (3S)-1-[5-tert-butyl-3-[(1-methyltetrazol-5-yl)methyl]triazolo[4,5-d]pyrimidin-7-yl]pyrrolidin-3-ol according to the present invention refers to the compound of formula (I). [ka] It is also known as RO6868847, bicasinabin, CAS number 1433361-02-4, and methods for making and using the compound are described in WO 2022 / 106669 and WO 2012EP71788.

[0032] In one embodiment, the present invention provides the compound (3S)-1-[5-tert-butyl-3-[(1-methyltetrazol-5-yl)methyl]triazolo[4,5-d]pyrimidin-7-yl]pyrrolidin-3-ol for use as an agonist of cannabinoid receptor 2 in a patient at a dose of 0.75 mg to 300 mg.

[0033] In certain embodiments, the present invention provides (3S)-1-[5-tert-butyl-3-[(1-methyltetrazol-5-yl)methyl]triazolo[4,5-d]pyrimidin-7-yl]pyrrolidin-3-ol for use in the treatment or prevention of a chronic degenerative disease in a patient at a dose of 0.75 mg to 300 mg.

[0034] In certain embodiments, the present invention provides (3S)-1-[5-tert-butyl-3-[(1-methyltetrazol-5-yl)methyl]triazolo[4,5-d]pyrimidin-7-yl]pyrrolidin-3-ol for use in the treatment or prevention of neurodegenerative inflammatory disorders, atherosclerosis, liver fibrosis, or diabetic microvascular complications in a patient at a dose of 0.75 mg to 300 mg.

[0035] In certain embodiments, the present invention provides (3S)-1-[5-tert-butyl-3-[(1-methyltetrazol-5-yl)methyl]triazolo[4,5-d]pyrimidin-7-yl]pyrrolidin-3-ol for use in the treatment or prevention of diabetic microvascular complications in a patient, wherein the diabetic microvascular complication is diabetic retinopathy, diabetic macular edema, or nonproliferative diabetic retinopathy, at a dose of 0.75 mg to 300 mg.

[0036] In certain embodiments, the present invention provides (3S)-1-[5-tert-butyl-3-[(1-methyltetrazol-5-yl)methyl]triazolo[4,5-d]pyrimidin-7-yl]pyrrolidin-3-ol for use in the treatment or prevention of nonproliferative diabetic retinopathy (NPDR) in a patient at a dose of 0.75 mg to 300 mg.

[0037] In certain embodiments, the present invention provides (3S)-1-[5-tert-butyl-3-[(1-methyltetrazol-5-yl)methyl]triazolo[4,5-d]pyrimidin-7-yl]pyrrolidin-3-ol for use in the treatment or prevention of proliferative diabetic retinopathy (PDR) in a patient at a dose of 0.75 mg to 300 mg.

[0038] In certain embodiments, the present invention provides (3S)-1-[5-tert-butyl-3-[(1-methyltetrazol-5-yl)methyl]triazolo[4,5-d]pyrimidin-7-yl]pyrrolidin-3-ol for use in the treatment or prevention of diabetic macular edema (DME) in a patient at a dose of 0.75 mg to 300 mg.

[0039] In certain embodiments, the present invention provides (3S)-1-[5-tert-butyl-3-[(1-methyltetrazol-5-yl)methyl]triazolo[4,5-d]pyrimidin-7-yl]pyrrolidin-3-ol for use in the treatment or prevention of diabetic nephropathy (DN) in a patient at a dose of 0.75 mg to 300 mg.

[0040] In a more specific embodiment, the present invention provides (3S)-1-[5-tert-butyl-3-[(1-methyltetrazol-5-yl)methyl]triazolo[4,5-d]pyrimidin-7-yl]pyrrolidin-3-ol for use in the treatment or prevention as described above, wherein the patient is a human (such as a male or female).

[0041] In a more specific embodiment, the present invention provides orally administered (3S)-1-[5-tert-butyl-3-[(1-methyltetrazol-5-yl)methyl]triazolo[4,5-d]pyrimidin-7-yl]pyrrolidin-3-ol for use in the treatment or prevention of the above.

[0042] In a more particular embodiment, the present invention provides (3S)-1-[5-tert-butyl-3-[(1-methyltetrazol-5-yl)methyl]triazolo[4,5-d]pyrimidin-7-yl]pyrrolidin-3-ol for use in the treatment or prevention as above, wherein the patient is a human (such as a male or female) and the compound is administered at a dose of between 0.75 mg and 300 mg, particularly orally.

[0043] In a more specific embodiment, the present invention provides (3S)-1-[5-tert-butyl-3-[(1-methyltetrazol-5-yl)methyl]triazolo[4,5-d]pyrimidin-7-yl]pyrrolidin-3-ol for use in the treatment or prevention of the above, administered once daily.

[0044] In a more particular embodiment, the present invention provides (3S)-1-[5-tert-butyl-3-[(1-methyltetrazol-5-yl)methyl]triazolo[4,5-d]pyrimidin-7-yl]pyrrolidin-3-ol for use in the treatment or prevention described herein above, administered in combination with other treatments, particularly in combination with laser, anti-VEGF treatment, or other treatments for diabetic retinopathy.

[0045] In one embodiment, the present invention provides a method for treatment of a cannabinoid receptor 2 agonist, comprising administering to a patient, particularly a patient in need thereof, (3S)-1-[5-tert-butyl-3-[(1-methyltetrazol-5-yl)methyl]triazolo[4,5-d]pyrimidin-7-yl]pyrrolidin-3-ol at a dose of 0.75 mg to 300 mg.

[0046] In certain embodiments, the present invention provides a method for the treatment or prevention of a chronic degenerative disease, comprising administering to a patient, particularly a patient in need thereof, (3S)-1-[5-tert-butyl-3-[(1-methyltetrazol-5-yl)methyl]triazolo[4,5-d]pyrimidin-7-yl]pyrrolidin-3-ol at a dose of 0.75 mg to 300 mg.

[0047] In certain embodiments, the present invention provides a method for the treatment or prevention of neurodegenerative inflammatory disorders, atherosclerosis, liver fibrosis, or diabetic microvascular complications, comprising administering to a patient, particularly a patient in need thereof, (3S)-1-[5-tert-butyl-3-[(1-methyltetrazol-5-yl)methyl]triazolo[4,5-d]pyrimidin-7-yl]pyrrolidin-3-ol at a dose of 0.75 mg to 300 mg.

[0048] In certain embodiments, the present invention provides a method for the treatment or prevention of a diabetic microvascular complication, wherein the diabetic microvascular complication is diabetic retinopathy, diabetic macular edema, or nonproliferative diabetic retinopathy, comprising administering to a patient, particularly a patient in need thereof, (3S)-1-[5-tert-butyl-3-[(1-methyltetrazol-5-yl)methyl]triazolo[4,5-d]pyrimidin-7-yl]pyrrolidin-3-ol at a dose of 0.75 mg to 300 mg.

[0049] In certain embodiments, the present invention provides a method for the treatment or prevention of nonproliferative diabetic retinopathy (NPDR), comprising administering to a patient, particularly a patient in need thereof, (3S)-1-[5-tert-butyl-3-[(1-methyltetrazol-5-yl)methyl]triazolo[4,5-d]pyrimidin-7-yl]pyrrolidin-3-ol at a dose of 0.75 mg to 300 mg.

[0050] In certain embodiments, the present invention provides a method for the treatment or prevention of proliferative diabetic retinopathy (PDR), comprising administering to a patient, particularly a patient in need thereof, (3S)-1-[5-tert-butyl-3-[(1-methyltetrazol-5-yl)methyl]triazolo[4,5-d]pyrimidin-7-yl]pyrrolidin-3-ol at a dose of 0.75 mg to 300 mg.

[0051] In certain embodiments, the present invention provides a method for the treatment or prevention of diabetic macular edema (DME), comprising administering to a patient, particularly a patient in need thereof, (3S)-1-[5-tert-butyl-3-[(1-methyltetrazol-5-yl)methyl]triazolo[4,5-d]pyrimidin-7-yl]pyrrolidin-3-ol at a dose of 0.75 mg to 300 mg.

[0052] In certain embodiments, the present invention provides a method for the treatment or prevention of diabetic nephropathy (DN), comprising administering to a patient (particularly a patient in need thereof) (3S)-1-[5-tert-butyl-3-[(1-methyltetrazol-5-yl)methyl]triazolo[4,5-d]pyrimidin-7-yl]pyrrolidin-3-ol at a dose of 0.75 mg to 300 mg.

[0053] In more particular embodiments, the present invention provides methods for the treatment or prevention of the above, particularly wherein the patient is a human (such as male or female).

[0054] In more particular embodiments, the present invention provides methods for the above treatment or prevention which are administered orally.

[0055] In a more particular embodiment, the invention provides a method for the treatment or prevention as described above, wherein the patient is a human (such as male or female) and the compound is administered in a dose of between 0.75 mg and 300 mg, particularly orally.

[0056] In a more particular embodiment, the present invention provides a method for the treatment or prevention as described above, which is administered once daily.

[0057] In more particular embodiments, the present invention provides methods for the above treatment or prevention administered in combination with other treatments, particularly in combination with laser, anti-VEGF treatment, or other treatments for diabetic retinopathy.

[0058] In one embodiment, the present invention provides the use of the compound (3S)-1-[5-tert-butyl-3-[(1-methyltetrazol-5-yl)methyl]triazolo[4,5-d]pyrimidin-7-yl]pyrrolidin-3-ol in the manufacture of a medicament as an agonist of cannabinoid receptor 2 at a dose of 0.75 mg to 300 mg.

[0059] In a particular embodiment, the present invention provides use of the compound (3S)-1-[5-tert-butyl-3-[(1-methyltetrazol-5-yl)methyl]triazolo[4,5-d]pyrimidin-7-yl]pyrrolidin-3-ol for the manufacture of a medicament for the treatment or prevention of a chronic degenerative disease at a dose of 0.75 mg to 300 mg.

[0060] In a particular embodiment, the present invention provides use of the compound (3S)-1-[5-tert-butyl-3-[(1-methyltetrazol-5-yl)methyl]triazolo[4,5-d]pyrimidin-7-yl]pyrrolidin-3-ol for the manufacture of a medicament for the treatment or prevention of neurodegenerative inflammatory disorders, atherosclerosis, liver fibrosis, or diabetic microvascular complications at a dose of 0.75 mg to 300 mg.

[0061] In a specific embodiment, the present invention provides use of the compound (3S)-1-[5-tert-butyl-3-[(1-methyltetrazol-5-yl)methyl]triazolo[4,5-d]pyrimidin-7-yl]pyrrolidin-3-ol at a dose of 0.75 mg to 300 mg for the manufacture of a medicament for the treatment or prevention of diabetic microvascular complications, wherein the diabetic microvascular complication is diabetic retinopathy, diabetic macular edema, or nonproliferative diabetic retinopathy.

[0062] In a particular embodiment, the present invention provides use of the compound (3S)-1-[5-tert-butyl-3-[(1-methyltetrazol-5-yl)methyl]triazolo[4,5-d]pyrimidin-7-yl]pyrrolidin-3-ol for the manufacture of a medicament for the treatment or prevention of nonproliferative diabetic retinopathy (NPDR) at a dose of 0.75 mg to 300 mg.

[0063] In a particular embodiment, the present invention provides use of the compound (3S)-1-[5-tert-butyl-3-[(1-methyltetrazol-5-yl)methyl]triazolo[4,5-d]pyrimidin-7-yl]pyrrolidin-3-ol for the manufacture of a medicament for the treatment or prevention of proliferative diabetic retinopathy (PDR) at a dose of 0.75 mg to 300 mg.

[0064] In a particular embodiment, the present invention provides use of the compound (3S)-1-[5-tert-butyl-3-[(1-methyltetrazol-5-yl)methyl]triazolo[4,5-d]pyrimidin-7-yl]pyrrolidin-3-ol for the manufacture of a medicament for the treatment or prevention of diabetic macular edema (DME) at a dose of 0.75 mg to 300 mg.

[0065] In a particular embodiment, the present invention provides use of the compound (3S)-1-[5-tert-butyl-3-[(1-methyltetrazol-5-yl)methyl]triazolo[4,5-d]pyrimidin-7-yl]pyrrolidin-3-ol for the manufacture of a medicament for the treatment or prevention of diabetic nephropathy (DN) at a dose of 0.75 mg to 300 mg.

[0066] In a more particular embodiment, the present invention provides the use of the compound (3S)-1-[5-tert-butyl-3-[(1-methyltetrazol-5-yl)methyl]triazolo[4,5-d]pyrimidin-7-yl]pyrrolidin-3-ol as defined above, particularly where the patient is a human being (such as a male or female).

[0067] In a more particular embodiment, the present invention provides the use of the above compound (3S)-1-[5-tert-butyl-3-[(1-methyltetrazol-5-yl)methyl]triazolo[4,5-d]pyrimidin-7-yl]pyrrolidin-3-ol, administered orally.

[0068] In a more particular embodiment, the present invention provides the use of the compound (3S)-1-[5-tert-butyl-3-[(1-methyltetrazol-5-yl)methyl]triazolo[4,5-d]pyrimidin-7-yl]pyrrolidin-3-ol as defined above, wherein the patient is a human (such as male or female) and the compound is administered at a dose of 0.75 mg to 300 mg, in particular orally.

[0069] In a more particular embodiment, the present invention provides the use of the compound (3S)-1-[5-tert-butyl-3-[(1-methyltetrazol-5-yl)methyl]triazolo[4,5-d]pyrimidin-7-yl]pyrrolidin-3-ol as defined above, administered once daily.

[0070] In a more particular embodiment, the present invention provides the use of the above compound (3S)-1-[5-tert-butyl-3-[(1-methyltetrazol-5-yl)methyl]triazolo[4,5-d]pyrimidin-7-yl]pyrrolidin-3-ol administered in combination with other treatments, particularly in combination with laser, anti-VEGF treatment, or other treatments for diabetic retinopathy. [Example]

[0071] Non-clinical research CB2 in vitro pharmacology Pharmacological studies were conducted in vitro and in vivo to examine the potency, selectivity, and activity of RO6868847. Pharmacological effects were determined in vivo in two different diabetic animal models, including a streptozotocin (STZ)-induced rat type 1 diabetic mouse model and a genetic type 2 diabetic mouse model, as well as in two acute models with acute inflammatory stimuli (LPS injection) or laser injury (CNV). Additional in vitro pharmacological studies were conducted to examine the pharmacological effects of RO6868847 in human whole blood obtained from either healthy volunteers or diabetic patients, as well as the assay's suitability as a biomarker of target engagement and pharmacodynamic effects (PD).

[0072] RO6868847 binds to human CB2 with a Ki of 51.3 nM and 62.8 nM when measured in Chinese hamster ovary (CHO) cells expressing recombinant CB2 or U968M cells expressing endogenous CB2. Among several species tested, RO6868847 exhibits the highest affinity for human and cynomolgus monkey CB2, with comparable Ki values ​​of 51.3 nM and 66.8 nM, respectively. Table 3 summarizes the affinity of RO6868847 for the CB2 receptor. Based on physiological expression levels and receptor activity states in endogenous systems, the human Ki of 62.8 nM in U698M cells is proposed for use in human dose prediction. [Table 1]

[0073] Selectivity of CB2 over CB1 and other targets Inhibition of cAMP accumulation in Chinese hamster ovary cells stably expressing recombinant CB2 was used to determine the functional potency of RO6868847 against different CB2 species and its selectivity over the closely related CB1. RO6868847 is a potent full agonist of human CB2 and exhibits similar potency for CB2 in other species, including cynomolgus monkeys, rats, and mice. Table 4 summarizes the potency across different CB2 species. [Table 2]

[0074] RO6868847 showed no affinity for human CB1, resulting in no receptor activation or inhibition at the highest concentration tested of 10 μM, and yielding >195-fold binding selectivity and >3600-fold functional selectivity over human CB1 (see Table 5). [Table 3]

[0075] The in vitro pharmacological profile of RO6868847 was evaluated at a concentration of 30 μM on a panel of 131 targets, including 57 CNS-specific and abuse-potential-specific targets. RO6868847 demonstrated high selectivity for CB2 across all panels. Further in vitro assays evaluated the selectivity of RO6868847 against 78 additional receptors and ion channels using radioligand binding assays, which demonstrated excellent selectivity for CB2. In functional cellular assays, RO6868847 activity was undetectable against both human and mouse 15-lipoxygenase, MAGL, diacylglycerol lipase, or FAAH enzymes in the endocannabinoid pathway (10 μM in mice and up to 500 μM for human targets). These results suggest that RO6868847 is unlikely to be responsible for off-target effects, and therefore, follow-up investigations in functional assays were not considered necessary.

[0076] Example 1: In vitro target engagement by monitoring CB2 surface expression on peripheral B cells in whole blood from healthy participants and diabetic patients To monitor target engagement by RO6868847 in humans, we developed a flow cytometry assay (1078220) utilizing an in-house monoclonal anti-human CB2 antibody in combination with established blood cell markers (CD45, CD14, CD3, and CD19). Using this assay, assessment of CB2 surface expression on peripheral leukocyte subsets in whole blood from healthy participants confirmed published findings by Castaneda and colleagues (Castaneda et al. 2013) that B cells exhibit low but robust surface expression of CB2, while CB2 surface expression is not detectable on other peripheral blood cell populations, such as T cells, granulocytes, or monocytes. Therefore, we selected B cells to monitor CB2 surface expression and evaluate possible drug effects on this measure in peripheral blood. Like most GPCR agonists, RO6868847 is expected to induce CB2 receptor internalization upon CB2 activation, reducing surface detection. To establish this measure as a surrogate target engagement biomarker, freshly isolated whole blood samples (n = 9) from healthy participants were incubated in vitro in duplicate with eight different increasing concentrations of RO6868847 for 1 hour at 37°C, followed by flow cytometry assays (1078220). Data from individual donors (n = 9) were expressed as the ratio of CB2 surface expression on B cells to T cells and normalized to the same ratio in each donor's whole blood without drug incubation (see Figure 1), revealing a concentration-dependent decrease in CB2 surface expression on B cells upon RO6868847 treatment.

[0077] Utilizing data from nine individual donors, a mean IC50 (TE) ± SD = 96.5 ± 27.1 nM was estimated, along with a respective mean Emax = 55.0 ± 4.6% (34.6 ng / mL). Furthermore, this in vitro data suggests that monitoring target engagement in human plasma using this flow cytometry assay may be feasible in human peripheral blood, and therefore was used in a Phase I study of BP40387 in healthy participants.

[0078] The availability of this in vitro target engagement assay also allowed us to examine target engagement-exposure responses in blood samples obtained from patients with diabetes mellitus (type 1 and type 2) and age-, sex-, and BMI-matched healthy controls (Table 6) (1097891). This population was selected because it resembles the target patient population envisioned for proof-of-concept Phase II trials, which encompasses patients with diabetes according to definitions by the World Health Organization and / or the American Diabetes Association. [Table 4]

[0079] These experiments demonstrated that in whole blood obtained from diabetic patients and matched healthy participants, RO6868847 induced dose-dependent CB2 receptor internalization on the surface of B cells compared to CB2 receptor levels without exposure to RO6868847 (Figure 2).

[0080] However, by comparing the respective mean IC50 ± SD values ​​of CB2 receptor levels for the three groups of participants (Table 7), the data show a trend that diabetic (DM) patients and matched healthy participants may require approximately 2-3 fold higher exposure to achieve half-maximal internalization of CB2 receptors on B cells (one-way anova with Tukey's multiple comparisons correction, p = 0.0575) compared to previously analyzed healthy participants. This was taken into consideration during dose selection for the Phase 2 study. [Table 5]

[0081] Example 2: In vitro pharmacodynamic effects of RO6868847 in an LPS challenge assay in whole blood from healthy volunteers and diabetic patients To investigate the putative anti-inflammatory pharmacodynamic effects mediated by the CB2 agonist RO6868847 in potentially clinically relevant human samples, we performed an ex vivo whole blood lipopolysaccharide (LPS) challenge assay. Sodium heparin whole blood samples from healthy participants (n = 8) were incubated in vitro with eight increasing concentrations of RO6868847 (0-30 μM) for 1 h at 37 °C and then exposed to LPS (100 ng / ml). After 18 h at 37 °C, supernatants were analyzed for changes in various chemokines and cytokines using a Luminex bead-based multiplex ELISA. These experiments revealed dose-dependent changes in specific chemokine and cytokine levels (decreases in IFNγ, IL1β, and TNFα and increases in G-CSF) following LPS stimulation in whole blood, thus suggesting an overall anti-inflammatory / immunomodulatory downstream pharmacodynamic effect of the CB2 agonist in vitro at high exposures.

[0082] Furthermore, this in vitro LPS whole blood challenge assay was also used in samples from diabetic participants (types I and II) and matched healthy participants (Table 6) to investigate whether the RO6868847 exposure (0–30 μM)-response relationship and respective changes in cytokine levels could be altered by any chronic inflammatory condition in diabetic (DM) and / or elderly healthy participants (HP), who are expected to be the study populations for the phase II proof-of-concept study.

[0083] In diabetic patients and matched healthy participants, we observed a dose-dependent attenuation of IFNγ and IL1β responses at high exposures in a small number of participants (Figure 4), confirming previous in vitro observations in healthy participants (Figure 3). However, for G-CSF and TNFα levels, we were unable to conclude an overall drug response over the range of exposures investigated due to the high variability observed and the fact that EC50 / IC50 values ​​could only be calculated for one or two individual samples (Figure 4, Table 8). [Table 6]

[0084] Overall, the data support a general immunomodulatory / anti-inflammatory effect of RO6868847 in elderly and diabetic patients. Whether these effects are similar but of different magnitude due to a less sensitive immune response or whether the differences are more significant remains to be investigated.

[0085] In vivo pharmacodynamic-based exposure-response analysis Several animal models have demonstrated the effects of bicasinabin relevant to DR pathophysiology, including inhibition of ocular leukostasis (rodent diabetic and non-diabetic models), reduction of retinal hyperfluorescence (rodent laser injury model), and preservation of retinal permeability, function, and microglial morphology (rodent diabetic model). The diversity of models tested (including acute antigen-stimulated vs. disease-modifying models) and the lack of recommended non-clinical DME / DR models with demonstrated clinical translation present challenges in estimating pharmacologically active exposure in humans.

[0086] Figure 10 shows the relationship between disease markers and the free AUC of the active S-epimer vicacinabin. 0-24 The data are presented as percent of control values ​​in various in vivo PD models as a function of exposure. Exposures were derived from satellite animals or extrapolated from previous PK or PD studies in the same species.

[0087] Based on this integrated assessment, the pharmacologically active exposure range observed in rats spanned 2 logs, with an upper limit (unbound AUC 0-24 [S-epimer] = 546 ng·hr / mL) corresponds to the full effect at 10 mg / kg in the rat STZ-induced type 1 diabetes model, and the lower limit (unbound AUC 0-24 [S-epimer] = 2.8 ng hr / mL) corresponds to a 50% reduction in disease markers (fluorescent area) in a laser-induced NV rat model. i unbound AUC resulting in plasma concentrations close to 0-24represents 52% of the upper limit of pharmacologically active exposure observed in rats.

[0088] Effects in humans Completed and ongoing Phase I / II clinical studies of RO6868847 are summarized below. All studies in the clinical development program are detailed in Appendix 4 and were conducted in accordance with the principles of Good Clinical Practice (GCP). The BP40387 study was a Phase I study conducted in healthy male and female participants and included five parts. Part 1 (SAD) consisted of an adaptive, single-ascending-dose, investigator- / participant-blinded, randomized, placebo-controlled, parallel study to investigate the safety, tolerability, and pharmacokinetics of RO6868847. Part 2 (FE) was designed as an open-label, randomized, two-period crossover, single-dose investigation of the effect of food on the pharmacokinetics of RO6868847. Part 3 (MAD) was an adaptive, multiple-ascending-dose, investigator- / participant-blinded, randomized, placebo-controlled, parallel study to investigate the safety, tolerability, and pharmacokinetics of RO6868847 administered once daily for 14 days. Part 4 (DDI-CYP induction) was a non-randomized, open-label, fixed-sequence, two-period study to investigate RO6868847-mediated CYP induction at 300 mg administered once daily for 14 days, using midazolam, repaglinide, and bupropion as probe substrates for CYP3A, CYP2C8, and CYP2B6, respectively. Part 5 (DDI-transporter-mediated inhibition) was a non-randomized, open-label, fixed-sequence, four-period study to investigate the inhibitory effects of RO6868847 on drug transporters, using metformin and atorvastatin as probe substrates for OCT2, MATE2-K, and OATP1B1, respectively. Overall, in Part 1 (SAD), a total of 33 healthy participants received a single oral dose of RO6868847 ranging from 0.75 mg to 300 mg, and 12 participants received placebo under fasting conditions. In Part 2 (FE), eight participants received a single oral dose of 100 mg of RO6868847 after at least 10 hours of overnight fasting and once within 30 minutes of starting a high-calorie, high-fat breakfast. In Part 3 (MAD), a total of 30 participants received multiple oral doses of 6 mg to 300 mg of RO6868847 QD for 14 days, and 10 participants received placebo.

[0089] Clinical Pharmacokinetics In Study BP40387, RO6868847 was rapidly absorbed under fasting conditions, with median maximum plasma concentrations (Tmax) between 1 and 4 hours. No significant deviations from dose proportionality were observed for Cmax and AUCInf / tau up to 100 mg. However, at doses above 100 mg, Cmax and AUCInf / tau appeared to increase in a slightly less than dose-proportional manner. AUCtau at steady-state drug correlated well with AUCInf after single-dose administration, indicating no relevant PK time-dependence. After multiple daily doses of RO6868847, steady state was achieved, on average, approximately 2 days after dosing, with only minor accumulation across the dose range investigated. Renal excretion of RO6868847 after single and multiple doses was minimal, accounting for less than 5% of the administered dose on average. Overall, food had no relevant effects on the pharmacokinetics of RO6868847.

[0090] In Part 3 (MAD), coadministration of a single oral dose of 0.100 mg midazolam with multiple oral doses of RO6868847 ranging from 6 mg to 300 mg once daily for 14 days demonstrated that RO6868847 was a weak inducer of CYP3A at the 300 mg dose administered, but not at the lower doses tested. In Study BP40387, administration of single and multiple oral doses of RO6868847 resulted in reversible, dose-dependent, and sustained CB2 receptor target engagement on blood B cells.

[0091] Pharmacokinetics after single and multiple dose oral administration to healthy subjects, single dose plasma pharmacokinetics In Part 1 (SAD) of Study BP40387, RO6868847 was rapidly absorbed overall under fasting conditions, but Tmax appeared to shift with increasing dose. Individual Tmax ranged from 1 to 6 hours for 100 mg and 300 mg of RO6868847. The main plasma PK parameters of RO6868847 are summarized in Table 20. AUCinf and Cmax increased approximately dose-proportionally up to 100 mg of RO6868847, but beyond 100 mg to 300 mg of RO6868847, Cmax and AUCinf increased in a slightly less than dose-proportional manner. After reaching peak plasma concentrations (Cmax), RO6868847 plasma concentrations declined monoexponentially, and the apparent terminal elimination half-life of RO6868847 ranged from 6.95 to 18.5 hours. Furthermore, dose variability in half-life increased with increasing dose due to individual participants exhibiting a biphasic decline after peak plasma concentrations. [Table 7]

[0092] In Part 2 (FE) of Study BP40387, the time to reach peak plasma concentration (Tmax) was slightly longer after dosing after a high-fat, high-calorie breakfast, consistent with delayed gastric emptying due to a prolonged meal. The median Tmax was achieved 4 hours after dosing in the fed state compared with 3 hours after dosing in the fasted state. On average, Cmax remained unaffected by food, and AUCInf was only slightly increased in the fed state compared with the fasted state.

[0093] Single-dose urinary pharmacokinetics Following administration of a single oral dose in Part 1 (SAD) of Study BP40387, RO6868847 was quantifiable in urine, but the proportion of the administered dose (Fe) recovered in urine as RO6868847 over 72 hours post-dose was low, with geometric mean values ​​ranging from 2.16% to 4.27% over the dose range studied.

[0094] Multiple-dose plasma pharmacokinetics In Part 3 (MAD) of Study BP40387, RO6868847 was rapidly absorbed, with overall individual times to peak plasma concentrations (Tmax) achieved between 1 and 4 hours post-dose for all dose groups on Days 1, 7, and 14 (except for the 6 mg dose group on Day 1, where Tmax ranged from 0.5 to 3 hours) (Table 21). Table 21 summarizes plasma PK parameters for RO6868847 at steady state on Day 14. At the end of the 14-day treatment period, RO6868847 plasma concentrations declined monoexponentially at the lowest dose investigated, 6 mg of RO6868847, whereas biexponential declines were observed at doses greater than 6 mg. The elimination phase was characterized by a geometric mean apparent terminal elimination half-life ranging from 8.27 hours to 39.0 hours across the dose range investigated. Statistical analysis showed that Cmax and AUCtau at steady state increased in a slightly less than dose-proportional manner for doses above 100 mg. However, this effect was more pronounced for Cmax, especially at the 300 mg dose of RO6868847. After multiple dose administration, both Cmax and AUCtau reached steady state approximately 2 days after treatment, with slight accumulation regardless of dose (Table 21). Statistical analysis of log-transformed accumulation rates showed no evidence of time-dependent pharmacokinetics. [Table 8]

[0095] Multiple-dose urinary pharmacokinetics At steady state, RO6868847 was quantifiable in urine, but the Fe recovered in urine as RO6868847 over the dosing interval (24 h) was low, with geometric mean values ​​ranging from 0.738% to 1.85% over the dose range studied.

[0096] Safety in Phase 1 BP40387 In Study BP40387, 11 participants (24.4%) reported at least one AE in Part 1, 1 participant (12.5%) in Part 2, 16 participants (40%) in Part 3, 12 participants (70.6%) in Part 4, and 2 participants (11.1%) in Part 5 (see Table 22). One participant (5.9%) was discontinued by the investigator in Part 4 of the study due to an AE. AEs reported in more than two participants in Part 1 were device site erythema (six participants in each PT) and vascular procedural complication (two participants in each PT); in Part 3, device site erythema (five participants in each PT), constipation (three participants in each PT), device site papule, device site pruritus, vascular procedural complication, and headache (two participants each); and in Part 4, constipation (two participants in each PT). In Study BP40387, one SAE was reported in Part 4 (DDI-CYP induction) of severe intensity (significantly altered T-wave morphology compared to baseline) and moderate intensity (synchronous tachycardia) in a subject treated with 300 mg bicasinabine QD. This event was considered related to bicasinabine by the investigator and resolved without sequelae within 1 week after treatment cessation.

[0097] Example 3 In Example 2, target engagement of RO6868847 was assessed on circulating B cells from whole blood samples using a newly developed exploratory flow cytometry assay to monitor RO6868847-induced CB2 receptor internalization on B cells. Figure 5 shows the average CB2 receptor target engagement versus time profile after administration of multiple doses of RO6868847 in Example 3, excluding data from the 6 mg dose (as it was not available for reporting). Reversible, dose-dependent, and sustained CB2 receptor target engagement was observed after administration of multiple doses of RO6868847 up to 300 mg as described in Example 2. Upon cessation of treatment, CB2 receptor surface expression returned to baseline within approximately one week. Target engagement appeared to plateau at a dose of 100 mg of RO6868847, with only a slight increase at higher doses of RO6868847. In Example 2, pro-inflammatory cytokine production was induced by ex vivo stimulation with LPS in freshly obtained whole blood samples to measure downstream markers of RO6868847-mediated CB2 receptor internalization and investigate its pharmacological profile. Participants in Study BP40387 were evaluated for serum levels of TNF-α, G-CSF, IL-1β, and interferon-γ to investigate changes in these cytokines in the presence of repeated daily doses of up to 300 mg of RO6868847 for 14 days. Under steady-state conditions, concentration-dependent effects on TNF-α, G-CSF, and IL1-β could be observed compared to baseline, as shown in Figures 6, 7, and 8, respectively. However, IFN-γ showed no detectable trends within the investigated concentration range of RO6868847 (see Figure 9). It should be noted that IL1-β analysis was confounded by a significant proportion of baseline data and could not be reported, especially at the highest tested dose of RO6868847, 300 mg (see Figure 8). Substantial variability in these measurements was observed. However, given the small cohort size in this study evaluation, this was expected, and dedicated routine statistical analysis was not performed.

[0098] conclusion Safety and Tolerability RO6868847 was safe and very well tolerated in healthy male and female participants after single oral doses of up to 300 mg. RO6868847 was generally well tolerated in healthy male and female participants following multiple oral doses of up to 300 mg RO6868847 administered once daily for 14 days, with the sole exception of one participant who experienced an SAE that was accompanied by a transient elevation of plasma trough concentrations of unknown cause(s) that was approximately 4-5 times higher than the average for the rest of the cohort at the 300 mg dose of RO6868847. Following single and multiple dose administration of RO6868847, there was no dose-related increase in the incidence or severity of reported AEs, and no clusters of adverse events were observed. No specific concerns were identified regarding clinical laboratory parameters, ECG, vital signs, C-SSRS, Bowdle VAS, Bond and Lader VAS, TBNK assay, or neurological examination.

[0099] Pharmacokinetics and Pharmacodynamics RO6868847 was rapidly absorbed, and no significant deviations from dose-proportionality were observed for AUC and Cmax after single and multiple oral doses of RO6868847 up to a dose of 100 mg. However, at doses greater than 100 mg, RO6868847 AUC and Cmax appeared to increase in a slightly less than dose-proportional manner. Following multiple once-daily doses of RO6868847, steady state was achieved, on average, approximately 2 days after dosing, with only minor accumulation. Overall, food had no relevant effects on the pharmacokinetics of RO6868847. Renal excretion of RO6868847 after single and multiple doses was minimal, accounting on average for less than 5% of the administered dose. Single and multiple doses of RO6868847 resulted in dose-dependent, sustained, but reversible CB2 receptor internalization on blood B cells. Multiple doses of RO6868847 appeared to attenuate ex vivo LPS-induced pro-inflammatory cytokine production (i.e., TNF-α, G-CSF, and IL1-β) in whole blood compared to baseline in an overall dose-dependent manner.

[0100] Aspects of the present invention Aspect 1. The compound (3S)-1-[5-tert-butyl-3-[(1-methyltetrazol-5-yl)methyl]triazolo[4,5-d]pyrimidin-7-yl]pyrrolidin-3-ol for use as an agonist of cannabinoid receptor 2 in a patient at a dose of between 0.75 mg and 300 mg.

[0101] Embodiment 2. A compound according to embodiment 1 for use in the treatment or prevention of a chronic degenerative disease.

[0102] Embodiment 3. A compound according to embodiment 1 or 2 for use in the treatment or prevention of neurodegenerative inflammatory disorders, atherosclerosis, liver fibrosis, or diabetic microvascular complications.

[0103] Aspect 4. The compound of any one of Aspects 1-3 for use in the treatment or prevention of a diabetic microvascular complication, wherein the diabetic microvascular complication is diabetic retinopathy, diabetic macular edema, or non-proliferative diabetic retinopathy.

[0104] Embodiment 5. A compound according to any one of embodiments 1 to 4 for use in the treatment or prevention of non-proliferative diabetic retinopathy (NPDR).

[0105] Aspect 6. The compound according to any one of Aspects 1 to 5, particularly wherein the patient is a human (e.g., male or female).

[0106] Aspect 7. The compound of any one of Aspects 1 to 6, which is administered orally.

[0107] Aspect 8. The compound of any one of Aspects 1 to 7, wherein the patient is a human (e.g., male or female) and the compound is administered at a dose of 0.75 mg to 300 mg, particularly orally.

[0108] Embodiment 9. The compound of any one of embodiments 1 to 8, which is administered once daily.

[0109] Embodiment 10. The compound according to any one of embodiments 1 to 9, which is administered in combination with other treatments, in particular in combination with laser, anti-VEGF treatment, or other treatments for diabetic retinopathy.

[0110] Embodiment 11. A method for the treatment of a cannabinoid receptor 2 agonist, comprising administering to a patient (particularly a patient in need thereof) (3S)-1-[5-tert-butyl-3-[(1-methyltetrazol-5-yl)methyl]triazolo[4,5-d]pyrimidin-7-yl]pyrrolidin-3-ol at a dose of 0.75 mg to 300 mg.

[0111] Embodiment 12. The method of embodiment 11 for the treatment or prevention of a chronic degenerative disease.

[0112] Embodiment 13. A method according to embodiment 11 or 12 for the treatment or prevention of neurodegenerative inflammatory disorders, atherosclerosis, liver fibrosis, or diabetic microvascular complications.

[0113] Embodiment 14. The method of any one of embodiments 11 to 13 for the treatment or prevention of diabetic microvascular complications, wherein the diabetic microvascular complication is diabetic retinopathy, diabetic macular edema, or non-proliferative diabetic retinopathy.

[0114] Embodiment 15. A method according to any one of embodiments 11 to 14, for the treatment or prevention of non-proliferative diabetic retinopathy (NPDR).

[0115] Aspect 16. The method of any one of Aspects 11 to 15, in particular, wherein the patient is a human (such as a male or female).

[0116] Aspect 17. The compound of any one of Aspects 11 to 16, which is administered orally.

[0117] Aspect 18. The method of any one of Aspects 11 to 17, wherein the patient is a human (e.g., male or female) and the compound is administered at a dose of 0.75 mg to 300 mg, particularly orally.

[0118] Embodiment 19. The compound according to any one of embodiments 11 to 18, which is administered once daily.

[0119] Embodiment 20. The method according to any one of embodiments 11 to 19, wherein the method is administered in combination with another treatment, in particular in combination with a laser, an anti-VEGF treatment, or another treatment for diabetic retinopathy.

[0120] Aspect 21. Use of the compound (3S)-1-[5-tert-butyl-3-[(1-methyltetrazol-5-yl)methyl]triazolo[4,5-d]pyrimidin-7-yl]pyrrolidin-3-ol in the manufacture of a medicament as an agonist of cannabinoid receptor 2 at a dosage of 0.75 mg to 300 mg.

[0121] Embodiment 22. The use according to embodiment 21 for the manufacture of a medicament for the treatment or prevention of a chronic degenerative disease.

[0122] Embodiment 23. Use according to embodiment 21 or 22 for the manufacture of a medicament for the treatment or prevention of neurodegenerative inflammatory disorders, atherosclerosis, liver fibrosis, or diabetic microvascular complications.

[0123] Aspect 24. The use according to any one of aspects 21 to 23 for the manufacture of a medicament for the treatment or prevention of diabetic microvascular complications, wherein the diabetic microvascular complication is diabetic retinopathy, diabetic macular edema, or non-proliferative diabetic retinopathy.

[0124] Embodiment 25. Use according to any one of embodiments 21 to 24 for the manufacture of a medicament for the treatment or prevention of non-proliferative diabetic retinopathy (NPDR).

[0125] Aspect 26. The use according to any one of Aspects 21 to 25, in particular, wherein the patient is a human (e.g., male or female).

[0126] Embodiment 27. The use according to any one of embodiments 21 to 26, wherein the use is administered orally.

[0127] Aspect 28. The use according to any one of aspects 21 to 27, wherein the patient is a human (e.g., male or female) and the compound is administered at a dose of 0.75 mg to 300 mg, in particular orally.

[0128] Embodiment 29. The use according to any one of embodiments 21 to 28, wherein the use is administered once daily.

[0129] Embodiment 30. The use according to any one of embodiments 21 to 28, wherein the use is administered in combination with another treatment, in particular in combination with a laser, an anti-VEGF treatment, or another treatment for diabetic retinopathy.

Claims

1. A pharmaceutical composition as an agonist for cannabinoid receptor 2, comprising the compound (3S)-1-[5-tert-butyl-3-[(1-methyltetrazole-5-yl)methyl]triazolo[4,5-d]pyrimidine-7-yl]pyrrolidine-3-ol, A pharmaceutical composition in which, when administered to a subject, the compound is administered to the subject in a dose of 0.75 mg to 300 mg.

2. A pharmaceutical composition according to claim 1 for the treatment or prevention of chronic degenerative diseases.

3. The pharmaceutical composition according to claim 1 for the treatment or prevention of neurodegenerative inflammatory disorders, atherosclerosis, hepatic fibrosis, or diabetic microvascular complications.

4. A pharmaceutical composition according to claim 1 for the treatment or prevention of diabetic microvascular complications, wherein the diabetic microvascular complications are diabetic retinopathy, diabetic macular edema, or nonproliferative diabetic retinopathy.

5. The pharmaceutical composition according to claim 1 for the treatment or prevention of nonproliferative diabetic retinopathy (NPDR).

6. The pharmaceutical composition according to any one of claims 1 to 5, wherein the subject is a human.

7. The pharmaceutical composition according to claim 6, wherein the human is a male or female human.

8. A pharmaceutical composition according to any one of claims 1 to 5, which is administered orally.

9. The pharmaceutical composition according to any one of claims 1 to 5, wherein the subject is a human and the pharmaceutical composition is administered orally.

10. The pharmaceutical composition according to claim 9, wherein the human is a male or female human.

11. A pharmaceutical composition according to any one of claims 1 to 5, which is administered once a day.

12. A pharmaceutical composition according to any one of claims 1 to 5, for administration in combination with other treatments.

13. The pharmaceutical composition according to claim 12, wherein the other treatment is laser, anti-VEGF treatment, or other treatment for diabetic retinopathy.

14. Use of the compound (3S)-1-[5-tert-butyl-3-[(1-methyltetrazole-5-yl)methyl]triazolo[4,5-d]pyrimidine-7-yl]pyrrolidine-3-ol for the manufacture of a pharmaceutical as an agonist of cannabinoid receptor 2, When the aforementioned pharmaceutical is administered to a subject, the aforementioned compound is administered to the subject in a dose of 0.75 mg to 300 mg.

15. The use according to claim 14, wherein the pharmaceutical product is for the treatment or prevention of a chronic degenerative disease.

16. The use according to claim 14, wherein the pharmaceutical is for the treatment or prevention of neurodegenerative inflammatory disorders, atherosclerosis, hepatic fibrosis, or diabetic microvascular complications.

17. The use according to claim 14, wherein the pharmaceutical product is for the treatment or prevention of diabetic microvascular complications, the use wherein the diabetic microvascular complications are diabetic retinopathy, diabetic macular edema, or nonproliferative diabetic retinopathy.

18. The use according to claim 14, wherein the pharmaceutical is for the treatment or prevention of nonproliferative diabetic retinopathy (NPDR).

19. The use according to any one of claims 14 to 18, wherein the subject is a human.

20. The use according to claim 19, wherein the human is a male or female human.

21. The use according to any one of claims 14 to 18, wherein the pharmaceutical product is administered orally.

22. The use according to any one of claims 14 to 18, wherein the subject is a human and the pharmaceutical product is administered orally.

23. The use according to claim 22, wherein the human is a male or female human.

24. The use according to any one of claims 14 to 18, wherein the pharmaceutical product is administered once a day.

25. The use according to any one of claims 14 to 18, wherein the pharmaceutical is to be administered in combination with other treatments.

26. The use according to claim 25, wherein the other treatment is a laser, an anti-VEGF treatment, or another treatment for diabetic retinopathy.

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