Jyoti Asundi

An Antibody–Drug Conjugate Targeting the Endothelin B Receptor for the Treatment of Melanoma

Purpose: To identify and evaluate targets amenable to antibody therapy in melanoma. Experimental Design: We searched for mRNA transcripts coding for cell-surface proteins with expression patterns similar to that of the melanoma oncogene MITF. One such candidate, the endothelin B receptor (EDNBR), was first analyzed for a functional contribution to tumor growth by conditional induction of shRNA. Second, antibodies were raised to the receptor, conjugated with monomethyl auristatin E, and tested for efficacy against melanoma tumor models generated from cell lines. Results: Conditional knockdown of the receptor in tumor xenograft models resulted in only a modest impact on tumor growth. A monoclonal antibody reactive with the N-terminal tail of EDNBR was found to internalize rapidly into melanoma cells. When conjugated with monomethyl auristatin E, the antibody–drug conjugate (ADC) showed remarkable efficacy against human melanoma cell lines and xenograft tumor models that was commensurate with levels of receptor expression. Comparative immunohistochemistry revealed a range of EDNBR expression across a panel of human melanomas, with the majority expressing levels equivalent to or greater than that in the models responsive to the ADC. Conclusion: An ADC targeting the EDNBR is highly efficacious in preclinical models of melanoma. Clin Cancer Res; 17(5); 965–75. ©2011 AACR.

An Antibody–Drug Conjugate Directed against Lymphocyte Antigen 6 Complex, Locus E (LY6E) Provides Robust Tumor Killing in a Wide Range of Solid Tumor Malignancies

Purpose: Chemotherapies are limited by a narrow therapeutic index resulting in suboptimal exposure of the tumor to the drug and acquired tumor resistance. One approach to overcome this is through antibody–drug conjugates (ADC) that facilitate greater potency via target-specific delivery of highly potent cytotoxic agents. Experimental Design: In this study, we used a bioinformatics approach to identify the lymphocyte antigen 6 complex locus E (LY6E), an IFN-inducible glycosylphosphatidylinositol (GPI)-linked cell membrane protein as a promising ADC target. We developed a monoclonal anti-LY6E antibody and characterized in situ LY6E expression in over 750 cancer specimens and normal tissues. Target-dependent anti-LY6E ADC killing was investigated both in vitro and in vivo using patient-derived xenograft models. Results: Using in silico approaches, we found that LY6E was significantly overexpressed and amplified in a wide array of different human solid tumors. IHC analysis revealed high LY6E protein expression in a number of tumor types, such as breast, lung, gastric, ovarian, pancreatic, kidney and head/neck carcinomas. Characterization of the endocytic pathways for LY6E revealed that the LY6E-specific antibody is internalized into cells leading to lysosomal accumulation. Consistent with this, a LY6E-specific ADC inhibited in vitro cell proliferation and produced durable tumor regression in vivo in clinically relevant LY6E-expressing xenograft models. Conclusions: Our results identify LY6E as a highly promising molecular ADC target for a variety of solid tumor types with current unmet medical need. Clin Cancer Res; 21(14); 3252–62. ©2015 AACR.

MAPK Pathway Inhibition Enhances the Efficacy of an Anti-Endothelin B Receptor Drug Conjugate by Inducing Target Expression in Melanoma

Therapies targeting the mitogen-activated protein (MAP) kinase pathway in melanoma have produced significant clinical responses; however, duration of response is limited by acquisition of drug resistance. Rational drug combinations may improve outcomes in this setting. We assessed the therapeutic combination of an antibody–drug conjugate (ADC) targeting the endothelin B receptor (EDNRB) with small-molecule inhibitors of the MAP kinase signaling pathway in melanoma. Cell lines and tumor models containing either mutant BRAF or NRAS, or wild-type for both, were exposed to small-molecule inhibitors of BRAF and MEK. Expression of EDNRB was analyzed and the therapeutic impact of combining the anti-EDNRB ADC with the BRAF and MEK inhibitors was assessed. Increased expression of EDNRB in response to inhibition of BRAF and/or MEK was observed and augmented the antitumor activity of the ADC. Enhanced target expression and ADC antitumor activity were realized irrespective of the response of the tumor model to the BRAF or MEK inhibitors alone and could be achieved in melanoma with mutant NRAS, BRAF, or neither mutation. Cells that acquired resistance to BRAF inhibition through long-term culture retained drug-induced elevated levels of EDNRB expression. Expression of EDNRB was not enhanced in normal human melanocytes by inhibition of BRAF and the combination of the ADC with MAPK inhibitors was well-tolerated in mice. The anti-EDNRB ADC combines well with BRAF and MEK inhibitors and could have therapeutic use in the majority of human melanoma cases. Mol Cancer Ther; 13(6); 1599–610. ©2014 AACR.

Abstract CT233: A first-in-human phase I study of the safety and pharmacokinetic (PK) activity of DEDN6526A, an anti-endothelin B receptor (ETBR) antibody-drug conjugate (ADC), in patients with metastatic or unresectable melanoma

Background: ETBR, a G-protein coupled receptor that can activate RAF/MEK signaling, is overexpressed in metastatic melanoma compared with normal skin. DEDN6526A is an ADC with the anti-mitotic agent monomethyl auristatin (MMAE) linked to the humanized IgG1 anti-ETBR monoclonal antibody and represents a novel targeted treatment against melanoma. In preclinical models, DEDN6526A shows dose-dependent anti-tumor activity in ETBR-expressing tumor xenografts. Methods: DEDN6526A (0.3-2.8 mg/kg) was given intravenously every 3 weeks (q3w) to pts with metastatic or unresectable cutaneous, mucosal, or ocular (uveal) melanoma in a 3+3 design to determine the maximum-tolerated dose, followed by cohort expansion at the recommended phase II dose (RP2D). PK samples were collected, and archival tumor tissue was assessed for ETBR expression by immunohistochemistry. Clinical activity was evaluated per RECIST v1.1. Results: As of 25 November 2013, 28 pts with median age 65 (range 24-82), ECOG PS 0-1, and median 2.5 prior therapies (range 0-5) enrolled in dose escalation and received median 6 doses of DEDN6526A (range 1-28). Based on cumulative safety data, the RP2D of DEDN6526A is 2.4 mg/kg q3w. Three DLTs occurred: G3 infusion-related reaction (IRR) (1.8 mg/kg), G3 transaminitis (2.4 mg/kg), and G3 elevated ALT and AST and transient G2 elevated bilirubin meeting criteria for drug induced liver injury (2.8 mg/kg). The most common treatment-emergent adverse events (AEs) of any grade (G) in ≥ 20% of pts were fatigue (57%), chills (39%), alopecia (32%), diarrhea (32%), nausea (29%), decreased appetite (25%), headache (25%), IRRs (25%), asthenia (21%), peripheral sensory neuropathy (21%), and vomiting (21%). The protocol was amended to allow steroid pre-medications prior to DEDN6526A administration, which mitigated the IRRs. Treatment-related G≥3 AEs in ≥ 5% of pts, assessed by investigators, were anemia (11%), neutropenia (11%), leukopenia (7%), and white blood count decreased (7%). Exposures of the conjugate (antibody-conjugated MMAE), total antibody, and unconjugated MMAE increased with dose. Among 19 pts treated at doses ≥ 1.8 mg/kg, there were four confirmed partial responses (21%); 6 pts (32%) had stable disease ≥ 6 months. Clinical benefit in pts did not seem to be associated with melanoma origin (cutaneous/mucosal vs. ocular), tumor BRAF mutation status, and/or progression on prior immune therapies. Correlation of ETBR expression with clinical activity will be presented. Conclusions: DEDN6526A administered at the RP2D of 2.4 mg/kg q3w is safe and tolerable. There is early evidence of anti-tumor activity in melanoma pts. Enrollment in the expansion cohort is ongoing. Citation Format: Jeffrey R. Infante, Shahneen K. Sandhu, Catriona M. McNeil, Omar Kabbarah, Chunze Li, Wei Zhong, Jyoti Asundi, Katie Wood, Yu-Waye Chu, Omid Hamid. A first-in-human phase I study of the safety and pharmacokinetic (PK) activity of DEDN6526A, an anti-endothelin B receptor (ETBR) antibody-drug conjugate (ADC), in patients with metastatic or unresectable melanoma. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr CT233. doi:10.1158/1538-7445.AM2014-CT233

Abstract 947: An antibody drug conjugate (ADC) directed to lymphocyte antigen 6 complex, locus E (LY6E) delivers targeted chemotherapy to a wide range of solid tumor malignancies

Chemotherapies are limited by a narrow therapeutic index resulting in suboptimal exposure of the tumor to the drug and acquired tumor resistance. One approach to overcome this is through Antibody Drug Conjugates (ADCs); that facilitate greater potency via target specific chemotherapy delivery. In this study, we used a bioinformatics approach to identify Lymphocyte antigen 6, locus E, (LY6E), an interferon-inducible glycosylphosphatidylinositol (GPI)-linked cell membrane protein, as a promising ADC target. Immunohistochemical analysis revealed LY6E was overexpressed in a number of tumor types, such as, breast, including triple negative breast cancer, lung, gastric, ovarian, pancreatic and head/neck carcinomas. Characterization of the endocytic pathways for LY6E revealed rapid uptake of the LY6E specific antibody into cells leading to lysosomal accumulation. Consistent with this, a LY6E specific ADC inhibited in vitro cell proliferation and produced durable tumor regression in vivo in high LY6E expressing cancers. Our results identify LY6E as a highly promising molecular ADC target for a variety of solid tumor types with current unmet medical need. Citation Format: Jyoti Asundi, Lisa Crocker, Jarrod Tremayne, Paul Polakis, Ron Firestein. An antibody drug conjugate (ADC) directed to lymphocyte antigen 6 complex, locus E (LY6E) delivers targeted chemotherapy to a wide range of solid tumor malignancies. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 947. doi:10.1158/1538-7445.AM2015-947

Abstract 2924: MAPK pathway inhibition enhances the efficacy of an anti-endothelin B receptor drug conjugate by inducing target expression in melanoma

Proceedings: AACR Annual Meeting 2014; April 5-9, 2014; San Diego, CA Therapies targeting the MAP kinase pathway in melanoma have produced significant clinical responses; however, duration of response is limited by acquisition of drug resistance. Rationale drug combinations may improve outcomes in this setting. We assessed the therapeutic combination of an antibody drug conjugate (ADC) targeting the Endothelin B receptor (EDNRB) with small molecule inhibitors of the MAP kinase signaling pathway in melanoma. Cell lines and tumor models containing either mutant BRAF or NRAS, or wildtype for both, were exposed to small molecule inhibitors or BRAF and MEK. Expression of EDNRB was analyzed and the therapeutic impact of combining the anti-EDNRB ADC with the BRAF and MEK inhibitors was assessed. Increased expression of EDNRB in response to inhibition of BRAF and/or MEK was observed and augmented the antitumor activity of the ADC. Enhanced target expression and ADC anti-tumor activity were realized irrespective of the response of the tumor model to the BRAF or MEK inhibitors alone and could be achieved in melanoma with mutant NRAS, BRAF or neither mutation. Cells that acquired resistance to BRAF inhibition through long-term culture retained drug-induced elevated levels of EDNRB expression. Expression of EDNRB was not enhanced in normal human melanocytes by inhibition of BRAF and the combination of the ADC with MAPK inhibitors was well tolerated in mice. The anti-EDNRB ADC combines well with BRAF and MEK inhibitors and could have therapeutic utility in the majority of human melanoma cases. Citation Format: Jyoti Asundi, Paul Polakis, Jennifer A. Lacap, Michelle Nannini. MAPK pathway inhibition enhances the efficacy of an anti-endothelin B receptor drug conjugate by inducing target expression in melanoma. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2924. doi:10.1158/1538-7445.AM2014-2924

Abstract B192: Pharmacokinetic characterization of anti-ETBR antibody drug conjugate in preclinical species.

Purpose. Anti-ETBR-MC-vc-PAB-MMAE (anti-ETBR-vc-E) is an antibody-drug conjugate (ADC) targeting the Endothelin B receptor. It comprises the humanized IgG1 anti-ETBR monoclonal antibody and a potent anti-mitotic agent, monomethyl auristatin E (MMAE), linked through MC-VC-PAB linker. Anti-ETBR antibody and its conjugate cross-react with mouse, rat, cynomolgus monkey and human, which provides an opportunity to compare and evaluate the impact of normal tissue expression on ADC disposition in preclinical species. Methods. Anti-ETBR-vc-E was administered intravenously in mice (0.5 and 5 mg/kg), rats (0.5 and 5 mg/kg), and cynomolgus monkeys (0.3 and 1 mg/kg). The samples were assayed for total antibody and conjugated antibody concentrations in mice and rats while total antibody and antibody-conjugated MMAE concentrations were determined in monkeys. Concentration-time data were used to estimate the pharmacokinetic parameters. Results. The pharmacokinetic behavior of the ADC was characterized by monitoring and comparing total antibody, conjugated antibody, and antibody-conjugated MMAE disposition. The clearances for all three analytes were dose-independent after intravenous administration in mice, rats, or cynomolgus monkeys over the dose range studied. Conjugated antibody and antibody-conjugated MMAE clearances were approximately 2-fold higher than total antibody clearance. Clearance of total antibody in rat was approximately 2-fold higher than in mouse and monkey, with values of 19.4, 7.91, and 8.99 mL/day/kg, respectively. Half-life ranged from 8 to 10 days and volume of distribution ranged from 37 to 40 mL/kg. Conclusions. The pharmacokinetics of anti-ETBR-vc-E total antibody was dose proportional in rodents and monkey suggesting that the presence of cross-reactive antigen in the preclinical species had no impact on clearance. All three species showed a relatively higher deconjugation clearance compared to total antibody clearance. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2011 Nov 12-16; San Francisco, CA. Philadelphia (PA): AACR; Mol Cancer Ther 2011;10(11 Suppl):Abstract nr B192.