Abstract Background: Pre-clinical profile of GRC 54276, a clinical candidate with Phase 1/2 clinical trial ongoing, is presented here. GRC 54276 is a novel small molecule inhibitor of Hematopoietic progenitor kinase 1 (HPK1), a serine/threonine kinase1,2 that negatively regulates T and B cell receptor signaling3. Inhibition of HPK1 is an attractive therapeutic strategy for immuno-oncology based treatment of solid tumors3. Methods: GRC 54276 was designed and developed using SAR based medicinal chemistry design supported by computational approaches. In vitro profiling was done using a battery of biochemical assays, functional read-outs and primary human in vitro T-cell activation assays. In vivo efficacy was demonstrated in mouse colon tumor models of CT26 and MC38-hPD-L1. In vivo inhibition of biomarker pSLP76 Ser(376) by GRC 54276 was determined using CT26 tumor model. Detailed ADME-PK studies have been performed along with safety tolerability studies conducted in mice and monkeys. Results: GRC 54276, demonstrated excellent in vitro immune profile of target engagement and anti-tumor immune response activity in both human and mouse systems. As a single agent, GRC 54276 demonstrated strong inhibition of tumor growth and biomarker pSLP76 Ser(376) in the CT26 tumor model. Enhanced efficacy was demonstrated by combining GRC 54276 with check-point blocking antibodies anti-CTLA4 and Atezolizumab in the CT26 and MC38-hPD-L1 models, respectively. GRC 54276 robustly enhanced complete tumor rejections when combined with Atezolizumab in the MC38-hPD-L1 model, correlating with increased immune effector memory T cells. Pharmacokinetic profile of GRC 54276 is characterized by high permeability, rapid absorption and moderate oral bioavailable across species. GRC 54276 is non-gentoxic with no observed adverse effects in mice and no treatment related cardiovascular or respiratory effects in repeat dose toxicity study in monkeys. Conclusions: GRC 54276 is a novel HPK1 inhibtitor with acceptable pre-clincial profile and is currently undergoing a Phase 1/2 clinical trial. Acknowledgements: We thank Pooja S, Shital M, Rahul B, Ajit J, Sanjay G, Somesh K, Pramod S for their contributions to the project References: 1. F.Kiefer et al., The EMBO Journal 1996 2. Hu et al., Genes and Development 1996 3. Sawasdikosol and Burakoff. eLife 2020;9:e55122 Citation Format: Sravan Mandadi, Sanjib Das, Malini Bajpai, Jagmohan Saini, Murugan Chinnapattu, Sanjay Patale, Sandip Patil, Nanasaheb Kadlag, Nayan Waghmare, Balasaheb Gavhane, Ameya Deshpande, Dnyaneshwar Dahale, Vidya Kattige, Priyanka Pangre, Namrata Singh, Ekta Kashyap, Megha Marathe, Jiju Mani, Atul Akarte, Chandrasekhar Misra, Subhadip Das, Anuj Singh, Pandurang Lambade, Avratanu Das, Chaitanya Tirumalasetty, Raju Patole, Nilanjana Biswas, Vikas Karande, Heta Shah, Dayanidhi Behera, Pankaj Jain, Pavankumar Sancheti, Pramod Pawar, Vinod KR, Venkatesha Udupa, Sachin S. Chaudhari, Nagaraj Gowda, Pravin S. Iyer. GRC 54276, a novel small molecule inhibitor of HPK1 has entered phase 1/2 clinical trial insolid malignancies and Hodgkin’s/non Hodgkin’s lymphoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 463.

Background Hematopoietic progenitor kinase 1 (HPK1) is a member of the mitogen-activated protein kinase (MAP4K) family of protein serine/threonine kinases1,2 and is a negative regulator of T and B cell receptor signaling3. Inhibition of HPK1 is an attractive therapeutic strategy for immuno-oncology based treatment of solid tumors3. We present in vitro, in vivo, pharmacokinetic (PK) and early safety profiles of a novel and differentiated HPK1 inhibitor GRC 54276. Methods GRC 54276 is our clinical candidate, designed and developed using intuitive medicinal chemistry design and supported by computational approaches. SAR studies included a battery of biochemical assays, functional read-outs and primary human in vitro T-cell activation assays. In vivo efficacy was demonstrated in syngeneic mouse tumor models, both as a single agent and combination with immune check-point blockers (ICB), mouse anti-CTLA4 antibody or Atezolizumab (human anti-PD-L1 antibody). ADME-PK properties was evaluated cross-species. GLP and non-GLP safety tolerability studies were conducted in mice and monkeys. Results GRC 54276 demonstrated sub-nanomolar HPK1 potency, strong target engagement of pSLP76 inhibition, anti-tumor cytokines (IL-2 and IFN-γ) induction, reversal of immunosuppression by prostaglandin E2 (PGE2) or adenosine in both human and mouse systems. GRC 54276 demonstrated very strong tumor growth inhibition (TGI) efficacy as single agent and significantly enhanced TGI in combination with ICB antibodies anti-CTLA4 (CT26 model) or Atezolizumab (MC38-hPD-L1 model). The in vivo TGI efficacy mechanistically correlated with increased immune responses of cytokine induction, infiltration of cytotoxic T cells, tumor rejections accompanied by immune memory T cells induction. Pharmacokinetic profile of GRC 54276 included cross-species oral bioavailability (30 to 100%), predominant clearance by CYP3A4 with no significant inhibition of major CYP isoforms, negative activation potency in human PXR assay at several-fold over EDmax exposures. Safety profile demonstrated that GRC 54276 is non-genotoxic in the bone marrow micronucleus assay in mice and has no hERG liability. The no observed adverse effect levels in the 14-day and 17-day exploratory studies in mice and monkeys were 50 and 15 mg/kg/day, respectively. Conclusions GRC 54276, our clinical candidate is potent, selective, orally bioavailable HPK1 inhibitor demonstrating strong single-agent and combination efficacy, low DDI liability accompanied by acceptable early safety profile in mice and monkeys. GRC 54276 is undergoing IND enabling studies to advance to Phase 1 clinical trial. Acknowledgements We thank Vidya Kattige, Pooja Sawant, Shital More, Rahul B. Bhadane, Ajit Jagadale, Sanjay Gaikwad, Pramod Sagar for their contributions to the project References 1. The EMBO Journal 1996 2. Genes and Development 1996 3. eLife 2020;9:e55122 Citation Format: Sravan Mandadi, Sanjib Das, Jagmohan Saini, Sachin S. Chaudhari, Murugan Chinnapattu, Ameya Deshpande, Dnyaneshwar Dahale, Malini Bajpai, Priyanka Pangre, Namrata Singh, Ekta Kashyap, Megha Marathe, Jiju Mani, Atul Akarte, Chandrasekhar Misra, Subhadip Das, Anuj Singh, Avratanu Das, Pandurang Lambade, Chaitanya Tirumalasetty, Raju Patole, Nilanjana Biswas, Vikas Karande, Heta Shah, Dayanidhi Behera, Pankaj Jain, Pavankumar Sancheti, Somesh Kakade, Pramod K. Pawar, Vinod KR, Venkatesha Udupa, Nagaraj Gowda, Pravin S. Iyer. IND-ready clinical candidate for HPK1 developed with excellent efficacy and safety profile [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1804.

The role of RORγ as a transcription factor for Th17 cell differentiation and thereby regulation of IL-17 levels is well known. Increased RORγ expression along with IL-17A levels was observed in animal models, immune cells and BAL fluid of COPD patients. Increased IL-17A levels in severe COPD patients are positively correlated with decreased lung functions and increased severity symptoms and emphysema, supporting an urgency to develop novel therapies modulating IL-17 or RORγ for COPD treatment. We identified a potent RORγ inhibitor, PCCR-1 using hit to lead identification followed by extensive lead optimization by structure–activity relationship. PCCR-1 resulted in RORγ inhibition with a high degree of specificity in a biochemical assay, with > 300-fold selectivity over other isoforms of ROR. Our data suggest promising potency for IL-17A inhibition in human and canine PBMCs and mouse splenocytes with no significant impact on Th1 and Th2 cytokines. In vivo, PCCR-1 exhibited significant efficacy in the acute CS model with dose-dependent inhibition of the PD biomarkers that correlated well with the drug concentration in lung and BAL fluid, demonstrating an acceptable safety profile. This inhibitor effectively inhibited IL-17A release in whole blood and BALf samples from COPD patients. Overall, we identified a selective inhibitor of RORγ to pursue further development of novel scaffolds for COPD treatment.

Background Hematopoietic progenitor kinase 1 (HPK1, MAP4K1), is a negative regulator of T and B cell receptor signaling.1 2 3 A strong anti-tumor immunogenic response and tumor rejection was observed in mice with HPK1 gene knocked out.3 Treatment of HPK1 kinase dead mice with immune check-point blockers (ICBs) demonstrated enhanced tumor growth inhibition.3 Hence, HPK1 is an attractive therapeutic strategy for immuno-oncology based treatment in cancers. In comparison to our previous HPK1 small molecule inhibitor, PCC,4 we present here a differentiated novel HPK1 inhibitor, PCC-1 with good anti-T cell kinases selectivity and stronger anti-tumor efficacy in CT26 tumor model. In addition, using the syngeneic model of MC38 expressing human PD-L1, we present for the first time, the combination efficacy of a HPK1 inhibitor with the clinical ICB, Atezolizumab. Methods Intuitive medicinal chemistry complemented by structure-based drug design was used to identify & develop potent inhibitors of HPK1 with optimal kinase selectivity, PK and in vivo efficacy profile. The SAR efforts were guided by biochemical assays, functional read-outs and primary human in vitro T-cell activation assays. In vivo target engagement and pharmacodynamic data was generated using CT26 and MC38-hPD-L1 tumor models. Results PCC-1 has sub-nanomolar HPK1 inhibition potency and strong target engagement resulting in pSLP76 inhibition, enhanced anti-tumor cytokine production of IL-2 and/or IFNgamma in Jurkat cells, human PBMCs and human whole blood. PCC-1 also demonstrated nanomolar potency in inducing a complete reversal of PGE2 or adenosine mediated immunosuppression. Oral dosing of PCC-1 as a single agent, induced strong tumor growth inhibition (TGI) in the syngeneic model of CT26 and MC38-hPD-L1 tumor models. Combination of PCC-1 with anti-CTLA4 in CT26 tumor model induced significantly greater TGI than anti-CTLA4 alone. Moreover, as a first, the combination of PCC-1 with clinical ICB, Atezolizumab in MC38-hPD-L1 induced enhanced rejection of tumors. These results strongly suggest PCC-1 as a promising candidate for HPK1 inhibition and as a combination partner with ICBs in clinic. Conclusions PCC-1 is a novel, orally active HPK1 inhibitor that demonstrates excellent stand-alone efficacy and enhances current immunotherapy efficacy. Further evaluation of PCC-1 is ongoing to advance towards clinic. Acknowledgements We thank Dnyaneshwar Dahale, Sanjay Patale, Sandip Patil, Vidya Kattige, Jiju Mani, Namrata Singh, Ekta Kashyap, Sandeep Thorat, Pankaj Jain and Pramod Sagar for their contributions to the project Trial Registration N/A References Kiefer F, et al. The EMBO Journal 1996. Hu, et al. Genes and Development 1996. Sawasdikosol, Burakoff. eLife 2020;9:e55122. Sachin S Chaudhari, et al. Poster#1709, AACR Annual Meeting April-May 2021. Ethics Approval The studies involving animals have obtained ethics approval from Institutional Animal Ethics Committee (IAEC), The Committee for the Purpose of Control and Supervision of Experiments on Animals (CPCSEA), New Delhi, India, GRC/IAEC/472/2020-1. Participants of the studies have given informed consent before taking part.

Abstract Background Hematopoietic progenitor kinase 1 (HPK1, MAP4K1), a member of the MAP4K family of protein serine/threonine kinases1,2 is a negative regulator of TCR and BCR3. Immunosuppressive mediators PGE2 and adenosine regulate activation of HPK13. In vivo anti-tumor activity by HPK1 gene deletion, kinase dead HPK1 and small molecule HPK1 inhibitors have been demonstrated in multiple immunogenic syngeneic tumor models3,4. Enhanced anti-tumor efficacy in these models was achieved by combining HPK1 inhibitors with immune check-point blockers (ICBs) like anti-PD-1, anti-PD-L1or anti-CTLA4 antibodies3,4. HPK1 inhibition is a target for immuno-oncology treatment in cancers responsive or non-responsive to current ICBs. Methods Creative medicinal chemistry design complemented by structured-based support was used to identify & develop inhibitors of HPK1 with pico/nano-molar potency, optimal kinase selectivity, PK and efficacy profile. Our SAR efforts were guided by biochemical assays, functional read-outs and primary human in vitro T-cell activation assays. In vivo target engagement and pharmacodynamic data was generated using mouse models of the lung (LLC) and colon (MC38 and CT26) cancer. Results We describe novel and potent HPK1 inhibitors that can inhibit pSLP76 and enhance IL-2 production in Jurkat cells, IL-2 and IFN-γ production in human PBMCs, whole blood and primary T cells and TNF-α production by dendritic cells following stimulation. A reversal of PGE2 or adenosine mediated immunosuppression was also achieved by the inhibitors. These compounds on oral dosing showed strong tumor growth inhibition (TGI) in syngeneic models refractory to ICBs (subcutaneous LLC) as well as responsive to ICBs (subcutaneous MC38 and CT26; orthotopic LLC). TGI was accompanied by the immune response of increased cytokine (IL-2/IFNγ) levels and tumor infiltrating lymphocytes (TILs). Combination with ICBs induced enhanced TGI correlating with enhanced cytokine induction and TILs. We confirmed the immune-mediated mechanism of inhibitors by their lack of efficacy in immune-compromised mice. The lead compound demonstrated cardiac safety (hERG assay), lack of genotoxicity and very good safety margins in the mouse exploratory toxicology studies up to 28-days. Conclusion We have discovered a novel, orally active HPK1 inhibitor that demonstrates excellent stand-alone efficacy in multiple tumor models and also offers the potential to enhance current immunotherapy regimens in both responsive and refractory cancers. Further evaluation of our lead molecule towards the clinic is underway. Acknowledgements Sanjib D, Megha M, Jiju M, Sheetal K, Arti J, Swayam M, Srinivas K, Pradeep V, Vikram B, Abhay K, Jagmohan S, Ravi T, Mohammad Y, Rahul B, Ajit J, Sanjay G, Pramod S. References 1. Kiefer et al., EMBO Journal 1996; 2. Hu et al., Genes and Development 1996; 3. Sawasdikosol and Burakoff. eLife 2020;9:e55122; 4. AACR Annual Meeting June 22-24 2020 Citation Format: Sachin S. Chaudhari, Malini Bajpai, Sravan Mandadi, Vidya G. Kattige, Sandeep Thorat, Varada Potdar, Priyanka Pangre, Pooja Sawant, Chandrasekhar Misra, Subhadip Das, Atul Akarte, Anuj Singh, Sandeep Mahankali, Pandurang Lambade, Avratanu Das, Raju Patole, Venkatesha Udupa, Pavankumar Sancheti, Pramod K. Pawar, Somesh Kakade, Vinod K. R, Nilanjana Biswas, Heta Shah, Dayanidhi Behera, Pankaj Jain, Nagaraj Gowda, Pravin S. Iyer. Novel small molecule HPK1 inhibitor induces immunogenic anti-tumor effects [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1709.

and in vivo efficacy profile. The SAR efforts were guided by bio-chemical assays, functional read-outs and primary human in vitro T-cell activation assays. In vivo target engagement and pharmacodynamic data was generated using CT26 and MC38-hPD-L1 tumor models. Results PCC-1 has sub-nanomolar HPK1 inhibition potency and strong target engagement resulting in pSLP76 inhibition, enhanced anti-tumor cytokine production of IL-2 and/or IFN-gamma in Jurkat cells, human PBMCs and human whole blood. PCC-1 also demonstrated nanomolar potency in inducing a complete reversal of PGE2 or adenosine mediated immunosuppression. Oral dosing of PCC-1 as a single agent, induced strong tumor growth inhibition (TGI) in the syngeneic model of CT26 and MC38-hPD-L1 tumor models. Combination of PCC-1 with anti-CTLA4 in CT26 tumor model induced significantly greater TGI than anti-CTLA4 alone.

The retinoic acid receptor-related orphan nuclear receptor gamma t (RORγt) is a master regulator of Th17 cell differentiation and production of IL17A, IL17F and IL22. Recent evidences in animal models and COPD patients suggest a strong role of RORγt-IL17 axis in COPD inflammation and exacerbation. An inhaled RORγt inverse agonist is expected to provide efficacy without potential systemic side effects. We synthesized various novel, potent, selective RORγt inverse agonists for inhaled delivery. The lead compound, GRC39815, was evaluated in various in vitro and in vivo COPD models followed by toxicological assessment in preclinical species. GRC39815 is a highly selective and potent inverse agonist of RORγt with physicochemical properties suitable for inhalation. In RORγt binding and functional assays, GRC39815 displayed a potent IC50 of 3-7 nM and is >390 fold selective over Th1 and Th2 cytokines. It demonstrated excellent selectivity over a panel of receptors, ion channels, transporters and enzymes. In animal models of sub-acute and chronic cigarette smoke induced COPD, intranasal delivery of GRC39815 demonstrated a dose dependent inhibition of lung inflammation, emphysema and IL17 immunoreactivity with very low systemic concentrations. In COPD patient BAL cells stimulated with anti CD3/28, GRC 39815 showed potent IL17 inhibition. GRC 39815 had no meaningful effect on hERG, action potential duration in canine Purkinje fibers and was non-genotoxic. Conclusion: GRC39815 is a novel, selective and safe inhaled RORγt inverse agonist for potential antiinflammatory treatment of COPD and currently in Phase-1 clinical trial to evaluate safety, tolerability and pharmacokinetics in healthy adults.

Background: Antibody mediated IL-17 blockade is remarkably successful in autoimmune disease treatment. Recent studies implicate IL-17 in airway diseases. Increased IL-17 immuno-reactivity and Th17 cells in lung tissue and IL-17 level in bronchoalveolar lavage fluid (BALF) is reported in human COPD patients and tobacco-smoke-exposed rats. Anti-IL-17 Ab attenuates airway inflammation in these animals. In human COPD patients, lung IL-17 immuno-reactivity correlates with disease severity. Recent studies also implicate IL-17 in COPD exacerbations. Hence IL-17 blockade seems a very attractive approach in potentially addressing the anti-inflammatory treatment gap in COPD management. IL-17 production is modulated by RORγt / RORc receptor in mice / man and offers a novel mechanism for IL-17 blockade. Objective: We synthesized several novel, potent, selective RORc blockers and evaluated efficacy of the series lead called “Compound A” in animal model of COPD. Results: Compound A is highly selective and potent preclinical lead with IC50 ofless than 40 nM in RORc binding and PBMC- IL-17 release assay. In vivo, it produced ∼ 75 % inhibition of IL-17 release induced by LPS and anti-CD3. Its efficacy in a COPD was evaluated in cigarette smoke mouse model under acute (7 days) and chronic (3 months) conditions. Compound A showed promising efficacy in suppressing lung cellular infiltration by 50-70% upon intranasal administration. In chronic model, the compound showed significant decrease of lung emphysema, epithelial IL-17 immuno-reactivity and RORgt/IL-17 positive lung cells. Conclusion: Our RORc antagonist is a novel, inhalable IL-17 blocker for potential treatment of COPD.

We report the design and synthesis of novel 5,6-diarylated pyridin-2(1H)-one derivatives as pharmacophoric PDE10A inhibitors. This highly potent molecular scaffold was developed from an inactive diarylpyridine-2-amine derivative 3b by extensive and systematic analogue synthesis and SAR analysis. Further optimization of the scaffold resulted in identification of pyridin-2(1H)-one 18b as a lead compound with good potency (IC50 = 1.6 nM) and selectivity (>6000-fold) over other related PDEs but with a poor pharmacokinetic profile. Careful metabolite profiling of 18b revealed that poor systemic exposure in rats (Cmax = 44 ng/mL; AUC0-t = 359 ng · h/mL) at 10 mg/kg was due to the formation of O-glucuronide conjugate by phase 2 metabolism. The structure of the glucuronide metabolite was confirmed by retention time and LC-MS/MS fragmentation matching with the synthetic glucuronide 26. The problem of low exposure of 18b was effectively addressed by its conversion to an acetate prodrug 25b, which upon oral dosing resulted in an improved pharmacokinetic profile (Cmax = 359 ng.h/mL; AUC0-t = 2436 ng.h/mL) and a desirable brain to plasma ratio of 1.2. The prodrug 25b showed good efficacy in selected rodent models of psychosis.

We report analogue-based rational design and synthesis of two novel series of polycyclic heteroarenes, pyrrolo[3,2-b]quinolines and pyrido[2,3-b]indoles, tethered to a biaryl system by a methyl-, ethyl- or propyl ether as PDE10A inhibitors. A number of analogues were prepared with variable chain length and evaluated for their ability to block PDE10A enzyme using a radiometric assay. Detailed SAR analyses revealed that compounds with an ethyl ether linker are superior in potency compared to compounds with methyl or propyl ether linkers. These compounds, in general, showed poor metabolic stability in rat and human liver microsomes. The metabolic profile of one of the potent compounds was studied in detail to identify metabolic liabilities of these compounds. Structural modifications were carried out that resulted in improved metabolic stability without significant loss of potency.

The design, synthesis and structure activity relationship studies of a series of compounds from benzo[d]imidazo[5,1-b]thiazole scaffold as phosphodiesterase 10A (PDE10A) inhibitors are discussed. Several potent analogs with heteroaromatic substitutions (9a–d) were identified. The anticipated binding mode of these analogs was confirmed by performing the in silico docking experiments. Later, the heteroaromatics were substituted with saturated heteroalkyl groups which provided a tool compound 9e with excellent PDE10A activity, PDE selectivity, CNS penetrability and with favorable pharmacokinetic profile in rats. Furthermore, the compound 9e was shown to be efficacious in the MK-801 induced psychosis model and in the CAR model of psychosis.

The synthesis and structure–activity relationship studies of isothiazole and isoxazole fused pyrimidones as PDE7 inhibitors are discussed. The pharmacokinetic profile of 10 and 21 with adequate CNS penetration, required for in vivo Parkinson’s disease models, are disclosed.

The synthesis and structure-activity relationship studies of a series of compounds from imidazopyridazinone scaffold as PDE7 inhibitors are disclosed. Potent analogs such as compounds 7 (31nM), 8 (27nM), and 9 (12nM) were identified. The PDE selectivity and pharmacokinetic profile of compounds 7, 8 and 9 are also disclosed. The adequate CNS penetration of compound 7 in mice allowed it to be tested in the MPTP induced PD model and haloperidol induced catalepsy model to probe the differential pharmacology of PDE7 in the striatal pathway.

La presente invention a pour objet des inhibiteurs de la phosphodiesterase-10. En particulier, la presente invention concerne des derives tricycliques qui sont utiles en tant qu'inhibiteurs de la phosphodiesterase-10. La presente invention concerne aussi des procedes de preparation des composes selon la presente invention, des intermediaires utilises dans leur synthese, des compositions pharmaceutiques, et des methodes de traitement ou de prevention de maladies, d'etats pathologiques et/ou de troubles modules par la phosphodiesterase-10. (Formule I) (I)

La presente invention concerne des composes olefiniques a substitution aryle en tant qu'inhibiteurs de la phosphodiesterase 10A (PDE10A). En particulier, lesdits composes sont utiles pour le traitement ou la prevention de maladies, d'affections et/ou de troubles par inhibition de l'enzyme phosphodiesterase 10A. L'invention a egalement pour objet des procedes de preparation desdits composes, des intermediaires utilises dans leur synthese, leurs compositions pharmaceutiques.