Ruslan D. Novosiadly

Clinical and Translational Results of a Phase II, Randomized Trial of an AntiIGF-1R (Cixutumumab) in Women with Breast Cancer That Progressed on Endocrine Therapy

Purpose: This phase II trial evaluated the efficacy and safety of cixutumumab, a human anti–insulin-like growth factor receptor 1 (IGF-1R) monoclonal IgG1 antibody, and explored potential biomarkers in postmenopausal women with hormone receptor–positive breast cancer. Experimental Design: Patients with hormone receptor–positive breast cancer that progressed on antiestrogen therapy received (2:1 randomization) cixutumumab 10 mg/kg and the same antiestrogen (arm A) or cixutumumab alone (arm B) every 2 weeks (q2w). Primary endpoint was progression-free survival (PFS); secondary endpoints included overall survival (OS) and safety. Correlative analyses of IGF-1R, total insulin receptor (IR), and IR isoforms A (IR-A) and B (IR-B) expression in tumor tissue were explored. Results: Ninety-three patients were randomized (arm A, n = 62; arm B, n = 31). Median PFS was 2.0 and 3.1 months for arm A and arm B, respectively. Secondary efficacy measures were similar between the arms. Overall, cixutumumab was well tolerated. IGF-1R expression was not associated with clinical outcomes. Regardless of the treatment, lower IR-A, IR-B, and total IR mRNA expression in tumor tissue was significantly associated with longer PFS [IR-A: HR, 2.62 (P = 0.0062); IR-B: HR, 2.21 (P = 0.0202); and total IR: HR, 2.18 (P = 0.0230)] and OS [IR-A: HR, 2.94 (P = 0.0156); IR-B: HR, 2.69 (P = 0.0245); and total IR: HR, 2.72 (P = 0.0231)]. Conclusions: Cixutumumab (10 mg/kg) with or without antiestrogen q2w had an acceptable safety profile, but no significant clinical efficacy. Patients with low total IR, IR-A, and IR-B mRNA expression levels had significantly longer PFS and OS, independent of the treatment. The prognostic or predictive value of IR as a biomarker for IGF-1R–targeted therapies requires further validation. Clin Cancer Res; 22(2); 301–9. ©2015 AACR.

Intrinsic Resistance to Cixutumumab Is Conferred by Distinct Isoforms of the Insulin Receptor

Despite a recent shift away from anti–insulin-like growth factor I receptor (IGF-IR) therapy, this target has been identified as a key player in the resistance mechanisms to various conventional and targeted agents, emphasizing its value as a therapy, provided that it is used in the right patient population. Molecular markers predictive of antitumor activity of IGF-IR inhibitors remain largely unidentified. The aim of this study is to evaluate the impact of insulin receptor (IR) isoforms on the antitumor efficacy of cixutumumab, a humanized mAb against IGF-IR, and to correlate their expression with therapeutic outcome. The data demonstrate that expression of total IR rather than individual IR isoforms inversely correlates with single-agent cixutumumab efficacy in pediatric solid tumor models in vivo. Total IR, IR-A, and IR-B expression adversely affects the outcome of cixutumumab in combination with chemotherapy in patient-derived xenograft models of lung adenocarcinoma. IR-A overexpression in tumor cells confers complete resistance to cixutumumab in vitro and in vivo, whereas IR-B results in a partial resistance. Resistance in IR-B–overexpressing cells is fully reversed by anti–IGF-II antibodies, suggesting that IGF-II is a driver of cixutumumab resistance in this setting. The present study links IR isoforms, IGF-II, and cixutumumab efficacy mechanistically and identifies total IR as a biomarker predictive of intrinsic resistance to anti–IGF-IR antibody. Implications: This study identifies total IR as a biomarker predictive of primary resistance to IGF-IR antibodies and provides a rationale for new clinical trials enriched for patients whose tumors display low IR expression. Mol Cancer Res; 13(12); 1615–26. ©2015 AACR.

An Open-Label, Multicenter, Randomized, Phase II Study of Cisplatin and Pemetrexed With or Without Cixutumumab (IMC-A12) as a First-Line Therapy in Patients With Advanced Nonsquamous Non-Small Cell Lung Cancer

Introduction: Type 1 insulin‐like growth factor receptor is deregulated in solid tumors. Cixutumumab, a monoclonal antibody that inhibits the activity of type 1 insulin‐like growth factor receptor, was investigated in combination with pemetrexed/cisplatin in the frontline setting. Methods: In this open‐label, phase II study, patients with stage IV nonsquamous NSCLC and a performance status of 0 to 1 were randomized (1:1) to receive 20 mg/kg cixutumumab, 500 mg/m2 pemetrexed, and 75 mg/m2 cisplatin (cixutumumab [n = 87]) or pemetrexed and cisplatin (control [n = 85]). Eligible patients received pemetrexed‐based maintenance therapy with cixutumumab (cixutumumab arm) or without it (control arm). The primary end point was progression‐free survival. Secondary end points assessed overall survival, objective response rate, and safety. Survival was analyzed by the Kaplan‐Meier method and Cox proportional hazard model. Exploratory correlative analyses were also performed. Results: The mean age of the intent‐to‐treat population (n = 172) was 59 years (range 32–83). Median progression‐free survival was 5.45 months with cixutumumab versus 5.22 months in the control (hazard ratio = 1.15, 95% confidence interval: 0.81–1.61; p = 0.44). Median overall survival was 11.33 months with cixutumumab versus 10.38 months in the control (hazard ratio = 0.93, 95% confidence interval: 0.64–1.36). Objective response rate did not differ between treatments (p = 0.338). Grade 3 or 4 hyperglycemia occurred at a higher rate with cixutumumab than in the control (9.4% versus 1.2%). One death possibly related to cixutumumab occurred. Conclusions: Efficacy was not improved in patients with nonsquamous NSCLC when cixutumumab was added to pemetrexed/cisplatin. Combination therapy was well tolerated and no new safety concerns were reported.

Mouse PDX Trial Suggests Synergy of concurrent Inhibition of RAF and EGFR in Colorectal Cancer with BRAF or KRAS mutations

Purpose: To evaluate the antitumor efficacy of cetuximab in combination with LSN3074753, an analog of LY3009120 and pan-RAF inhibitor in 79 colorectal cancer patient-derived xenograft (PDX) models. Experimental Design: Seventy-nine well-characterized colorectal cancer PDX models were employed to conduct a single mouse per treatment group (n = 1) trial. Results: Consistent with clinical results, cetuximab was efficacious in wild-type KRAS and BRAF PDX models, with an overall response rate of 6.3% and disease control rate (DCR) of 20.3%. LSN3074753 was active in a small subset of PDX models that harbored KRAS or BRAF mutations. However, the combination treatment displayed the enhanced antitumor activity with DCR of 35.4%. Statistical analysis revealed that BRAF and KRAS mutations were the best predictors of the combinatorial activity and were significantly associated with synergistic effect with a P value of 0.01 compared with cetuximab alone. In 12 models with BRAF mutations, the combination therapy resulted in a DCR of 41.7%, whereas either monotherapy had a DCR of 8.3%. Among 44 KRAS mutation models, cetuximab or LSN3074753 monotherapy resulted in a DCR of 13.6% or 11.4%, respectively, and the combination therapy increased DCR to 34.1%. Molecular analysis suggests that EGFR activation is a potential feedback and resistant mechanism of pan-RAF inhibition. Conclusions: MAPK and EGFR pathway activations are two major molecular hallmarks of colorectal cancer. This mouse PDX trial recapitulated clinical results of cetuximab. Concurrent EGFR and RAF inhibition demonstrated synergistic antitumor activity for colorectal cancer PDX models with a KRAS or BRAF mutation. Clin Cancer Res; 23(18); 5547–60. ©2017 AACR.

Targeting the TGFβ pathway with galunisertib, a TGFβRI small molecule inhibitor, promotes anti-tumor immunity leading to durable, complete responses, as monotherapy and in combination with checkpoint blockade

BackgroundTGFβ signaling plays a pleotropic role in tumor biology, promoting tumor proliferation, invasion and metastasis, and escape from immune surveillance. Inhibiting TGFβ’s immune suppressive effects has become of particular interest as a way to increase the benefit of cancer immunotherapy. Here we utilized preclinical models to explore the impact of the clinical stage TGFβ pathway inhibitor, galunisertib, on anti-tumor immunity at clinically relevant doses.ResultsIn vitro treatment with galunisertib reversed TGFβ and regulatory T cell mediated suppression of human T cell proliferation. In vivo treatment of mice with established 4T1-LP tumors resulted in strong dose-dependent anti-tumor activity with close to 100% inhibition of tumor growth and complete regressions upon cessation of treatment in 50% of animals. This effect was CD8+ T cell dependent, and led to increased T cell numbers in treated tumors. Mice with durable regressions rejected tumor rechallenge, demonstrating the establishment of immunological memory. Consequently, mice that rejected immunogenic 4T1-LP tumors were able to resist rechallenge with poorly immunogenic 4 T1 parental cells, suggesting the development of a secondary immune response via antigen spreading as a consequence of effective tumor targeting. Combination of galunisertib with PD-L1 blockade resulted in improved tumor growth inhibition and complete regressions in colon carcinoma models, demonstrating the potential synergy when cotargeting TGFβ and PD-1/PD-L1 pathways. Combination therapy was associated with enhanced anti-tumor immune related gene expression profile that was accelerated compared to anti-PD-L1 monotherapy.ConclusionsTogether these data highlight the ability of galunisertib to modulate T cell immunity and the therapeutic potential of combining galunisertib with current PD-1/L1 immunotherapy.

Molecular Basis for Necitumumab Inhibition of EGFR Variants Associated with Acquired Cetuximab Resistance

Acquired resistance to cetuximab, an antibody that targets the EGFR, impacts clinical benefit in head and neck, and colorectal cancers. One of the mechanisms of resistance to cetuximab is the acquisition of mutations that map to the cetuximab epitope on EGFR and prevent drug binding. We find that necitumumab, another FDA-approved EGFR antibody, can bind to EGFR that harbors the most common cetuximab-resistant substitution, S468R (or S492R, depending on the amino acid numbering system). We determined an X-ray crystal structure to 2.8 Å resolution of the necitumumab Fab bound to an S468R variant of EGFR domain III. The arginine is accommodated in a large, preexisting cavity in the necitumumab paratope. We predict that this paratope shape will be permissive to other epitope substitutions, and show that necitumumab binds to most cetuximab- and panitumumab-resistant EGFR variants. We find that a simple computational approach can predict with high success which EGFR epitope substitutions abrogate antibody binding. This computational method will be valuable to determine whether necitumumab will bind to EGFR as new epitope resistance variants are identified. This method could also be useful for rapid evaluation of the effect on binding of alterations in other antibody/antigen interfaces. Together, these data suggest that necitumumab may be active in patients who are resistant to cetuximab or panitumumab through EGFR epitope mutation. Furthermore, our analysis leads us to speculate that antibodies with large paratope cavities may be less susceptible to resistance due to mutations mapping to the antigen epitope. Mol Cancer Ther; 17(2); 521–31. ©2017 AACR.

High-content molecular profiling of T-cell therapy in oncology

Recent clinical data have revealed the remarkable potential for T-cell-modulating agents to induce potent and durable responses in a subset of cancer patients. In this review, we discuss molecular approaches, platforms, and strategies that enable a broader interrogation of the activity of agents that modulate the activity of tumor-specific T cells, to more comprehensively understand how and why the agents succeed and fail, as well as examples of data sets generated in clinical trials that have provided important insights into the biological activity of T-cell therapies and that support further rational development of this exciting treatment modality.

Discovery and preclinical characterization of the antagonist anti-PD-L1 monoclonal antibody LY3300054

BackgroundModulation of the PD-1/PD-L1 axis through antagonist antibodies that block either receptor or ligand has been shown to reinvigorate the function of tumor-specific T cells and unleash potent anti-tumor immunity, leading to durable objective responses in a subset of patients across multiple tumor types.ResultsHere we describe the discovery and preclinical characterization of LY3300054, a fully human IgG1λ monoclonal antibody that binds to human PD-L1 with high affinity and inhibits interactions of PD-L1 with its two cognate receptors PD-1 and CD80. The functional activity of LY3300054 on primary human T cells is evaluated using a series of in vitro T cell functional assays and in vivo models using human-immune reconstituted mice. LY3300054 is shown to induce primary T cell activation in vitro, increase T cell activation in combination with anti-CTLA4 antibody, and to potently enhance anti-tumor alloreactivity in several xenograft mouse tumor models with reconstituted human immune cells. High-content molecular analysis of tumor and peripheral tissues from animals treated with LY3300054 reveals distinct adaptive immune activation signatures, and also previously not described modulation of innate immune pathways.ConclusionsLY3300054 is currently being evaluated in phase I clinical trials for oncology indications.

Abstract 583: The CDK4/6 inhibitor abemaciclib induces synergistic immune activation and antitumor efficacy in combination with PD-L1 blockade

Targeting cyclin dependent kinases 4 and 6 (CDK4/6) with inhibitors such as abemaciclib has shown promise in early and late phase clinical trials in both breast cancer and NSCLC. While there is evidence that patients benefit from single-agent abemaciclib, combination strategies leveraging this compound together with immunotherapy are of interest for the treatment of these and other cancers. Consequently, it is important to understand if and how a cell cycle inhibitor can be combined with immunotherapy. However, because most preclinical studies have been performed using xenograft tumors in immune-compromised mice, the potential immunomodulatory effects of abemaciclib have not been adequately ascertained. To investigate the immune combinatorial potential of abemaciclib, we studied the effects of treatment alone and in combination with checkpoint immunotherapy in a murine syngeneic tumor model sensitive to abemaciclib using immuno-competent mice. Abemaciclib monotherapy of established murine CT26 tumors, which harbor KRAS G12C mutation and CDKN2A deletion, caused a dose-dependent delay in tumor growth. Surprisingly, gene expression analysis showed that treatment was associated with an increase in intra-tumor immune inflammation without major alteration in immune subset frequencies. Testing of various dosing regimens in this preclinical model found that monotherapy abemaciclib pretreatment followed by combination with anti-PD-L1 antibody therapy, induced an enhanced anti-tumor response compared to abemaciclib and anti-PD-L1 monotherapies. Optimal combination therapy exhibited superior anti-tumor efficacy, resulting in complete tumor regression (CR) in 50-60% of mice in a setting where anti-PD-L1 monotherapy showed little or no efficacy (0% CRs). Mice which maintained CRs after cessation of combination therapy were able to resist later CT26 rechallenge, demonstrating that abemaciclib in combination with anti-PD-L1 enabled the generation of an immunologic memory. Examination of intra-tumor gene expression during treatment found that combination therapy further amplified the immune/T cell activation signature compared to both monotherapies. Intra-tumoral suppression of cell cycle genes, which are indicative of inhibition of CDK4/6, was also greater during the combination therapy, suggesting that the effects anti-PD-L1 therapy may augment the cell cycle arrest induced by abemaciclib. Although it was uncertain if agents that inhibit cell proliferation could be combined with immunotherapy, these preclinical results demonstrate that it is possible to combine CDK4/6 inhibition by abemaciclib with checkpoint immunotherapy to improve tumor efficacy. The synergistic responses observed in terms of tumor efficacy, immune activation, and cell cycle control provides support for the clinical investigation of this combination. Citation Format: Jack Dempsey, Lysiane Huber, Amelie Forest, Jennifer R. Stephens, Thompson N. Doman, Jason Manro, Andrew Capen, Robert S. Flack, Gregory P. Donoho, Sean Buchanan, Alfonso De Dios, Kyla Driscoll, Michael Kalos, Ruslan Novosiadly, Richard P. Beckmann, David A. Schaer. The CDK4/6 inhibitor abemaciclib induces synergistic immune activation and antitumor efficacy in combination with PD-L1 blockade [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 583. doi:10.1158/1538-7445.AM2017-583

Abstract 5590: Combination of an oncokinase inhibitor merestinib with anti-PD-L1 results in enhanced immune mediated antitumor activity in CT26 murine tumor model

The combination of tumor targeted therapeutics with PD-L1 checkpoint blockade is being explored as a method to increase the clinical benefits of immunotherapy, and expand response to additional cancer types. Merestinib (Mer) is a kinase inhibitor targeting several oncokinases1 (including MET, MST1R, AXL, MERTK, and MKNK1/2) that can potentially modulate immune function, angiogenesis, as well as target the tumor 1-5. To determine the combinatorial potential with immunotherapy, the effects of Mer were evaluated in vitro on human T cells, PBMCs and murine tumor lines CT26 colon carcinoma (harbors KRASmt G12D expresses low Met/no p-Met/high Axl/p-Axl) and B16F10 melanoma (expressing high Met/pMet/peIF4E). Additionally, the anti-tumor effect of Mer was tested in vivo on established CT26 and B16F10 tumors compared to MET specific TKIs (savolitinib, PF4217903) alone or in combination with PD-L1 antibody (Ab) blockade. In vitro, Mer showed no significant effects on either T cells or PBMCs, but was able to inhibit downstream signaling in both CT26 and B16F10 showing activity on murine tumor cell lines. In vivo, daily Mer monotherapy (6, 12 or 24 mg/kg) showed significant anti-tumor effect at all doses in both CT26 and B16F10, that was not seen with either savolitinib or PF4217903. Concurrent combination of Mer (12 mg/kg) and anti-PD-L1 Ab (0.5 mg qw) in CT26 was found to have anti-tumor activity that was synergistic as compared to each single agent alone. While the effect of Mer monotherapy was lost when treatment ended, tumors continued to regress in the combination group even upon cessation of therapy. The combination was well tolerated and resulted in 90% complete responders compared to 30% with anti-PD-L1 Ab alone, 35 days after completing dosing. To test the ability to generate immunologic memory, complete responders were re-challenged with CT26 cells on the contralateral side. All mice in the combination group resisted re-challenge, showing that Mer/PD-L1 Ab combination was triggering immunologic memory. Although there was no significant change in intra-tumor immune cell populations between groups, combination therapy showed an enhanced and unique intra-tumor immune activation/inflammation gene expression signature compared to PD-L1 Ab monotherapy. The enhanced immune activation of the combination therapy, leading to synergistic anti-tumor efficacy, demonstrates that merestinib has the potential to augment immunotherapy while targeting the tumor directly. This preclinical data provides the rationale for the clinical investigation of merestinib in combination with checkpoint therapies targeting the PD-L1/PD1 axis (NCT02791334). 1 - Yan et al. Invest New Drugs 2013;31:833-44 2 - Balan et al. J Biol Chem 2015;290:8110-20 3 - Eyob et al. Cancer Discov 2013;3:751-60 4 - Lemke G. CSH Persp Biol 2013;5:a009076 5 - Piccirillo et al. Nat Immunol 2014;15:503-11 Citation Format: Sau-Chi Betty Yan, Victoria L. Peek, Jennifer R. Stephens, Um L. Um, Amaladas Nelusha, Colleen A. Burns, Kelly M. Credille, Thompson N. Doman, Scott W. Eastman, Beverly L. Falcon, Gerald E. Hall, Philip W. Iversen, Bruce W. Konicek, Jason R. Manro, Any T. Pappas, Julie A. Stewart, Michael B. Topper, Swee-Seong Wong, Michael Kalos, Ruslan D. Novosiadly, Richard A. Walgren, David Schaer. Combination of an oncokinase inhibitor merestinib with anti-PD-L1 results in enhanced immune mediated antitumor activity in CT26 murine tumor model [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 5590. doi:10.1158/1538-7445.AM2017-5590