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Inhibition of TGFβ signaling in concert with a checkpoint blockade has been shown to provide improved and durable antitumor immune response in mouse models. However, on-target adverse cardiovascular effects have limited the clinical use of TGFβ receptor (TGFβR) inhibitors in cancer therapy. To restrict the activity of TGFβR inhibitors to tumor tissues and thereby widen the therapeutic index, a series of tumor-activated prodrugs of a selective small molecule TGFβR1 inhibitor 1 were prepared by appending 1 to a serine protease substrate and a half-life extension fatty acid carbon chain. The prodrugs were shown to be selectively metabolized in tumor tissues relative to the heart and blood and demonstrated a prolonged favorable increase in the tumor-to-heart ratio of the active drug in tissue distribution studies. Once-weekly administration of the most tissue-selective compound 10 provided anti-tumor efficacy comparable to the parent compound and reduced systemic exposure of the active drug.

Transforming growth factor beta (TGF-β) is a pleiotropic cytokine that has a wide array of biological effects. For decades, tumor biology implicated TGF-β as an attractive therapeutic target due to its immunosuppressive effects. Toward this end, multiple pharmaceutical companies developed a number of drug modalities that specifically target the TGF-β pathway. BMS-986260 is a small molecule, selective TGF-βR1 kinase inhibitor that was under preclinical development for oncology. In vivo studies across mouse, rat, dog, and monkey and cryopreserved hepatocytes predicted human pharmacokinetics (PK) and distribution of BMS-986260. Efficacy studies of BMS-986260 were undertaken in the MC38 murine colon cancer model, and target engagement, as measured by phosphorylation of SMAD2/3, was assessed in whole blood to predict the clinical efficacious dose. The human clearance is predicted to be low, 4.25 ml/min/kg. BMS-986260 provided a durable and robust antitumor response at 3.75 mg/kg daily and 1.88 mg/kg twice-daily dosing regimens. Phosphorylation of SMAD2/3 was 3.5-fold less potent in human monocytes than other preclinical species. Taken together, the projected clinical efficacious dose was 600 mg QD or 210 mg BID for 3 days followed by a 4-day drug holiday. Mechanism-based cardiovascular findings in the rat ultimately led to the termination of BMS-986260. This study describes the preclinical PK characterization and pharmacodynamics-based efficacious dose projection of a novel small molecule TGF-βR1 inhibitor.

[Image: see text] Novel imidazole-based TGFβR1 inhibitors were identified and optimized for potency, selectivity, and pharmacokinetic and physicochemical characteristics. Herein, we report the discovery, optimization, and evaluation of a potent, selective, and orally bioavailable TGFβR1 inhibitor, 10 (BMS-986260). This compound demonstrated functional activity in multiple TGFβ-dependent cellular assays, excellent kinome selectivity, favorable pharmacokinetic properties, and curative in vivo efficacy in combination with anti-PD-1 antibody in murine colorectal cancer (CRC) models. Since daily dosing of TGFβR1 inhibitors is known to cause class-based cardiovascular (CV) toxicities in preclinical species, a dosing holiday schedule in the anti-PD-1 combination efficacy studies was explored. An intermittent dosing regimen of 3 days on and 4 days off allowed mitigation of CV toxicities in one month dog and rat toxicology studies and also provided similar efficacy as once daily dosing.

C-terminal Src kinase (CSK) functions as a negative regulator of T cell activation through inhibitory phosphorylation of LCK, so inhibitors of CSK are of interest as potential immuno-oncology agent...

[Image: see text] Indoleamine 2,3-dioxygenase 1 (IDO1) is a heme-containing dioxygenase enzyme implicated in cancer immune response. This account details the discovery of BMS-986242, a novel IDO1 inhibitor designed for the treatment of a variety of cancers including metastatic melanoma and renal cell carcinoma. Given the substantial interest around this target for cancer immunotherapy, we sought to identify a structurally differentiated clinical candidate that performs comparably to linrodostat (BMS-986205) in terms of both in vitro potency and in vivo pharmacodynamic effect in a mouse xenograft model. On the basis of its preclinical profile, BMS-986242 was selected as a candidate for clinical development.

[Image: see text] C-terminal Src kinase (CSK) functions as a negative regulator of T cell activation through inhibitory phosphorylation of LCK, so inhibitors of CSK are of interest as potential immuno-oncology agents. Screening of an internal kinase inhibitor collection identified pyridazinone lead 1, and a series of modifications led to optimized compound 13. Compound 13 showed potent activity in biochemical and cellular assays in vitro and demonstrated the ability to increase T cell proliferation induced by T cell receptor signaling. Compound 13 gave extended exposure in mice upon oral dosing and produced a functional response (decrease in LCK phosphorylation) in mouse spleens at 6 h post dose.

Immunotherapy has fundamentally changed the landscape of cancer treatment. Despite the encouraging results with the checkpoint modulators, response rates vary widely across tumor types, with a majority of patients exhibiting either primary resistance without a significant initial response to treatment or acquired resistance with subsequent disease progression. Hematopoietic progenitor kinase 1 (HPK1) is predominantly expressed in hematopoietic cell linages and serves as a negative regulator in T cells and dendritic cells (DC). While HPK1 gene knockout (KO) studies suggest its role in anti-tumor immune responses, the involvement of kinase activity and thereof its therapeutic potential remain unknown. To investigate the potential of pharmacological intervention using inhibitors of HPK1, we generated HPK1 kinase dead (KD) mice which carry a single loss-of-function point mutation in the kinase domain and interrogated the role of kinase activity in immune cells in the context of suppressive factors or the tumor microenvironment (TME). Our data provide novel findings that HKP1 kinase activity is critical in conferring suppressive functions of HPK1 in a wide range of immune cells including CD4+, CD8+, DC, NK to Tregs, and inactivation of kinase domain was sufficient to elicit robust anti-tumor immune responses. These data support the concept that an HPK1 small molecule kinase inhibitor could serve as a novel agent to provide additional benefit in combination with existing immunotherapies, particularly to overcome resistance to current treatment regimens.

[Image: see text] The multifunctional cytokine TGFβ plays a central role in regulating antitumor immunity. It has been postulated that inhibition of TGFβ signaling in concert with checkpoint blockade will provide improved and durable immune response against tumors. Herein, we describe a novel series of 4-azaindole TGFβ receptor kinase inhibitors with excellent selectivity for TGFβ receptor 1 kinase. The combination of compound 3f and an antimouse-PD-1 antibody demonstrated significantly improved antitumor efficacy compared to either treatment alone in a murine tumor model.

To mount an effective immune response, a T-cell receptor (TCR) must first recognize a cognate antigen presented by malignant cells. Successful effector T-cell activation initiates an intracellular signaling cascade that results in the release of cytotoxins and proinflammatory cytokines that ultimately result in the elimination of antigen-presenting diseased cells. A key regulator of this signaling cascade is the C-terminal Src (CSK) kinase. In lymphocytes CSK inhibits T-cell activation by phosphorylating the SRC kinase family member LCK at tyrosine 505 and thereby terminates the signaling cascade. To assess whether relieving an intracellular blockade of TCR signaling will lead to a more robust T-cell response, and hence improved tumor immunity, we developed both genetic and pharmacologic approaches to inhibit CSK activity. A tamoxifen-inducible transgenic CSK knockout mouse was engineered to bypass the embryonic lethality observed with constitutive systemic CSK depletion. As expected, loss of CSK enhanced cytokine production and increased antitumor immune response in the MC38 syngeneic tumor mouse model. Potent and selective CSK small-molecule inhibitors, in the presence of TCR stimulation, similarly activated the proximal TCR signaling pathway and increased cytokine production and proliferation. In an in vivo efficacy study, however, the CSK inhibitor as monotherapy or in combination with aPD1 did not improve antitumor immune response at the dose levels tested. This seemingly discrepant finding in tumor response observed in the tamoxifen-inducible KO mice and mice treated with the CSK inhibitor may reflect the need for a more optimized dosing regimen and target coverage. In this poster, we will present our findings on the role of CSK inhibition in tumor immunity. Citation Format: Cindy Wang, Carolyn Cao, Lisa Berman-Booty, Jesse Swanson, Rukiye Eraslan, Miguel Sanjuan, Gregory Vite, Lan-Ying Qin, John Hunt, Brian Fink, Susan Wee. Targeting CSK kinase activity to enhance antitumor immunity [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr B198.

The enzyme indoleamine 2,3-dioxygenase 1 (IDO1) catalyzes the degradation of tryptophan along the kynurenine pathway, and is frequently expressed in human malignancies. The activity of IDO1 induces an immunosuppressive microenvironment in tissues by inhibiting T-cell function through local depletion of tryptophan and through generation of kynurenine pathway metabolites. Inhibition of IDO1 is expected to diminish the immunosuppressive tumor microenvironment and improve cancer patient outcomes, particularly when used in combination with cancer immunotherapy agents such as nivolumab and ipilimumab. In this presentation, we will disclose the chemical structure, enzyme inhibitory mechanism, in vitro potency and in vivo pharmacodynamic (PD) activity of BMS’ IDO1 inhibitor currently in Phase I clinical trials. The compound is a potent and selective IDO1 inhibitor with no activity against another tryptophan degrading enzyme, tryptophan 2,3-dioxygenase (TDO). It exhibited potent cellular activity, suppressing kynurenine production in HEK293 cells overexpressing human IDO1 (IC50 = 1.1 nM) and in HeLa cells stimulated with IFNγ (IC50 = 1.7 nM). The compound also potently restored T-cell proliferation in a co-culture of T cells and human cancer cells and in a mixed lymphocyte reaction where T cells were co-cultured with allogeneic IDO1-expressing dendritic cells (EC50 = 1.2 nM). In vivo, when given once a day orally, the compound exhibited significant PD activity in mouse tumors grown subcutaneously in syngeneic hosts and in human tumors grown as xenografts in nude mice. Citation Format: John T. Hunt, Aaron Balog, Christine Huang, Tai-An Lin, Tai-An Lin, Derrick Maley, Johnni Gullo-Brown, Jesse Swanson, Jennifer Brown. Structure, in vitro biology and in vivo pharmacodynamic characterization of a novel clinical IDO1 inhibitor [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4964. doi:10.1158/1538-7445.AM2017-4964

Topological insulators represent a novel state of matter with surface charge carriers having a massless Dirac dispersion and locked helical spin polarization. Many exciting experiments have been proposed by theory, yet, their execution have been hampered by the extrinsic conductivity associated with the unavoidable presence of defects in Bi$_2$Te$_3$ and Bi$_2$Se$_3$ bulk single crystals as well as impurities on their surfaces. Here we present the preparation of Bi$_2$Te$_3$ thin films that are insulating in the bulk and the four-point probe measurement of the conductivity of the Dirac states on surfaces that are intrinsically clean. The total amount of charge carriers in the experiment is of order 10$^{12}$ cm$^{-2}$ only and mobilities up to 4,600 cm$^2$/Vs have been observed. These values are achieved by carrying out the preparation, structural characterization, angle-resolved and x-ray photoemission analysis, and the temperature dependent four-point probe conductivity measurement all in-situ under ultra-high-vacuum conditions. This experimental approach opens the way to prepare devices that can exploit the intrinsic topological properties of the Dirac surface states., accepted for publication in Proceedings of the National Academy of Sciences of the United States of America (PNAS)