Anti-EFNA4 Calicheamicin Conjugates Effectively Target Triple-Negative Breast and Ovarian Tumor-Initiating Cells to Result in Sustained Tumor Regressions

Abstract

Purpose: Triple-negative breast cancer (TNBC) and ovarian cancer each comprise heterogeneous tumors, for which current therapies have little clinical benefit. Novel therapies that target and eradicate tumor-initiating cells (TIC) are needed to significantly improve survival.Experimental Design: A panel of well-annotated patientderived xenografts (PDX) was established, and surface markers that enriched for TIC in specific tumor subtypes were empirically determined. The TICs were queried for overexpressed antigens, one of which was selected to be the target of an antibody-drug conjugate (ADC). The efficacy of the ADC was evaluated in 15 PDX models to generate hypotheses for patient stratification.Results: We herein identified E-cadherin (CD324) as a surface antigen able to reproducibly enrich for TIC in well-annotated, low-passage TNBC and ovarian cancer PDXs. Gene expression analysis of TIC led to the identification of Ephrin-A4 (EFNA4) as a prospective therapeutic target. An ADC comprising a humanized anti-EFNA4 monoclonal antibody conjugated to the DNA-damaging agent calicheamicin achieved sustained tumor regressions in both TNBC and ovarian cancer PDX in vivo. Non-claudin low TNBC tumors exhibited higher expression and more robust responses than other breast cancer subtypes, suggesting a specific translational application for tumor subclassification.Conclusions: These findings demonstrate the potential of PF-06647263 (anti-EFNA4-ADC) as a first-in-class compound designed to eradicate TIC. The use of well-annotated PDX for drug discovery enabled the identification of a novel TIC target, pharmacologic evaluation of the compound, and translational studies to inform clinical development.

Highly Potent, Anthracycline-based Antibody–Drug Conjugates Generated by Enzymatic, Site-specific Conjugation

“…As a result, such biparatopic ADCs show enhanced internalization and lysosomal trafficking and, consequently, superior potency when compared with a conventional, monospecific ADC (28). Finally, alternative payloads with substantially higher potency than microtubuledisrupting agents are being investigated, including pyrrolobenzodiazepine (PBD) dimers (29), enedyenes such as calicheamicin (30), indilino-benzodiazepine (IBD) pseudodimers (termed IGNs; ref. 31), as well as novel anthracylines (32,33).…”

Therapeutic potential of targeting the Eph/ephrin signaling complex

“…This highlights the potential of developing anti-angiogenic antibodies, although the wide expression of both EphB4 and ephrin-B2 in the normal human vasculature could nullify their use as therapeutic agents. An anti-ephrin-A4 mAb conjugated to the DNA-damaging agent calicheamicin has been shown to cause sustained tumor regressions in triple-negative breast and ovarian cancer in vivo (Damelin et al, 2015) and is now in a clinical trial.…”

Therapeutic targets and new directions for antibodies developed for ovarian cancer

“…Recently EFNA4 has also been identified as a novel tumor-initiating cell (TIC)-associated target 43 . TICs are a subpopulation of tumor cells that drive tumor growth, resistance to treatment, and disease recurrence.…”

“…PF-06647263 is a first-in-class, humanized IgG2 mAb (huE22) specific for EFNA4, and linked to calicheamicin, a DNA-damaging cytotoxic agent 43 . Initial results from a Phase 1 study in patients with advanced solid tumors unselected for EFNA-4 expression show a DLT in only 1 patient, and 2 patients (1 with ovarian cancer and one with triple negative breast cancer) have experienced a PR 151 …”