Orit Foord

A DLL3-targeted antibody-drug conjugate eradicates high-grade pulmonary neuroendocrine tumor-initiating cells in vivo.

Targeting DLL3 with an antibody-drug conjugate eliminates tumor-initiating cells in high-grade pulmonary neuroendocrine cancers. Not just another Notch Pulmonary neuroendocrine tumors, such as small cell lung cancer, are among the most difficult cancers to treat. Although standard chemotherapy regimens are available for this type of cancer, their effects are only transient, and the tumors typically acquire resistance to the drugs very quickly. Saunders et al. have discovered that DLL3, a ligand in the Notch signaling pathway, is associated with the neuroendocrine cancer phenotype. The authors targeted DLL3 with an antibody conjugated to a cytotoxic drug, which proved to be much more effective than standard chemotherapy for treating patient-derived tumor xenografts. Unlike chemotherapy, the anti-DLL3 treatment appeared to be particularly effective against tumor-initiating cells, which may account for its efficacy. The high-grade pulmonary neuroendocrine tumors, small cell lung cancer (SCLC) and large cell neuroendocrine carcinoma (LCNEC), remain among the most deadly malignancies. Therapies that effectively target and kill tumor-initiating cells (TICs) in these cancers should translate to improved patient survival. Patient-derived xenograft (PDX) tumors serve as excellent models to study tumor biology and characterize TICs. Increased expression of delta-like 3 (DLL3) was discovered in SCLC and LCNEC PDX tumors and confirmed in primary SCLC and LCNEC tumors. DLL3 protein is expressed on the surface of tumor cells but not in normal adult tissues. A DLL3-targeted antibody-drug conjugate (ADC), SC16LD6.5, comprised of a humanized anti-DLL3 monoclonal antibody conjugated to a DNA-damaging pyrrolobenzodiazepine (PBD) dimer toxin, induced durable tumor regression in vivo across multiple PDX models. Serial transplantation experiments executed with limiting dilutions of cells provided functional evidence confirming that the lack of tumor recurrence after SC16LD6.5 exposure resulted from effective targeting of DLL3-expressing TICs. In vivo efficacy correlated with DLL3 expression, and responses were observed in PDX models initiated from patients with both limited and extensive-stage disease and were independent of their sensitivity to standard-of-care chemotherapy regimens. SC16LD6.5 effectively targets and eradicates DLL3-expressing TICs in SCLC and LCNEC PDX tumors and is a promising first-in-class ADC for the treatment of high-grade pulmonary neuroendocrine tumors.

Potent High-Affinity Antibodies for Treatment and Prophylaxis of Respiratory Syncytial Virus Derived from B Cells of Infected Patients

Native human Abs represent attractive drug candidates; however, the low frequency of B cells expressing high-quality Abs has posed a barrier to discovery. Using a novel single-cell phenotyping technology, we have overcome this barrier to discover human Abs targeting the conserved but poorly immunogenic central motif of respiratory syncytial virus (RSV) G protein. For the entire cohort of 24 subjects with recent RSV infection, B cells producing Abs meeting these stringent specificity criteria were rare, <10 per million. Several of the newly cloned Abs bind to the RSV G protein central conserved motif with very high affinity (Kd 1–24 pM). Two of the Abs were characterized in detail and compared with palivizumab, a humanized mAb against the RSV F protein. Relative to palivizumab, the anti-G Abs showed improved viral neutralization potency in vitro and enhanced reduction of infectious virus in a prophylaxis mouse model. Furthermore, in a mouse model for postinfection treatment, both anti-G Abs were significantly more effective than palivizumab at reducing viral load. The combination of activity in mouse models for both prophylaxis and treatment makes these high-affinity human-derived Abs promising candidates for human clinical testing.

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

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 patient-derived 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. Clin Cancer Res; 21(18); 4165–73. ©2015 AACR.

A PTK7-targeted antibody-drug conjugate reduces tumor-initiating cells and induces sustained tumor regressions

PTK7 is a tumor-initiating cell antigen, which can be targeted with an antibody-drug conjugate to confer sustained tumor regressions. Initiating an antitumor attack Cancer is notorious for relapsing after treatment, making it difficult to eradicate from a patient’s body. Such relapses are driven by tumor-initiating cells, a type of stem cells that give rise to tumors. Damelin et al. determined that a protein called PTK7 is frequently present on tumor-initiating cells and developed an antibody-drug conjugate for targeting it. The authors demonstrated the effectiveness of this therapy in mouse models of several tumor types and confirmed that it reduces tumor-initiating cells and outperforms standard chemotherapy. The antibody-drug conjugate also had some unexpected benefits, reducing tumor angiogenesis and promoting antitumor immunity, all of which may contribute to its effectiveness. Disease relapse after treatment is common in triple-negative breast cancer (TNBC), ovarian cancer (OVCA), and non–small cell lung cancer (NSCLC). Therapies that target tumor-initiating cells (TICs) should improve patient survival by eliminating the cells that can drive tumor recurrence and metastasis. We demonstrate that protein tyrosine kinase 7 (PTK7), a highly conserved but catalytically inactive receptor tyrosine kinase in the Wnt signaling pathway, is enriched on TICs in low-passage TNBC, OVCA, and NSCLC patient–derived xenografts (PDXs). To deliver a potent anticancer drug to PTK7-expressing TICs, we generated a targeted antibody-drug conjugate (ADC) composed of a humanized anti-PTK7 monoclonal antibody, a cleavable valine-citrulline–based linker, and Aur0101, an auristatin microtubule inhibitor. The PTK7-targeted ADC induced sustained tumor regressions and outperformed standard-of-care chemotherapy. Moreover, the ADC specifically reduced the frequency of TICs, as determined by serial transplantation experiments. In addition to reducing the TIC frequency, the PTK7-targeted ADC may have additional antitumor mechanisms of action, including the inhibition of angiogenesis and the stimulation of immune cells. Together, these preclinical data demonstrate the potential for the PTK7-targeted ADC to improve the long-term survival of cancer patients.

Development of a new fully human anti-CD20 monoclonal antibody for the treatment of B-cell malignancies

SummaryDespite the widespread use of rituximab, a chimeric monoclonal antibody with demonstrated efficacy in the treatment of non-Hodgkin’s lymphomas, there is a recognized need to develop new agents with improved efficacy. Towards this end, using XenoMouse® technology, a fully human IgG1 anti-CD20 monoclonal antibody was generated. This antibody, denoted mAb 1.5.3, evoked enhanced pro-apoptotic activity in vitro, as compared to rituximab, in the Ramos lymphoma cell line. Also, mAb 1.5.3 mediated both complement-dependent cytotoxicity (CDC) and antibody-dependent cellular cytotoxicity (ADCC) similar to rituximab in human B-lymphoma lines. Interestingly, mAb 1.5.3 demonstrated superior ADCC compared to rituiximab when FcγRIIIa F/F allotype donors were profiled and superior cytolytic activity across multiple human B-lymphoma and chronic B-cell leukemia lines in an in vitro whole blood assay. Furthermore, mAb 1.5.3 exhibited enhanced anti-tumor activity in Ramos, Daudi, and Namalwa tumour xenograft models. Lastly, mAb 1.5.3 produced a superior B-cell depletion profile in lymph node organs and bone marrow as compared to rituximab in a primate pharmacodynamic (PD) model. These findings underscore the potential of mAb 1.5.3 to exhibit improved clinical activity in the treatment of B-cell malignancies compared to rituximab.

Abstract 1220: A novel PTK7-targeted antibody-drug conjugate eliminates tumor-initiating cells and induces sustained tumor regressions

Disease relapse after treatment is common in triple-negative breast cancer (TNBC), ovarian cancer and non-small cell lung cancer (NSCLC). Therapies that target tumor-initiating cells (TICs) should improve patient survival by eliminating the cells that can drive tumor regrowth and metastasis. Here we identify Protein Tyrosine Kinase 7 (PTK7), a highly conserved but catalytically inactive receptor tyrosine kinase, as an antigen that is enriched on TICs in low-passage patient-derived xenografts (PDX) of TNBC, NSCLC and other tumor types. An anti-PTK7 antibody-drug conjugate (ADC) was generated from a humanized anti-PTK7 monoclonal antibody, a cleavable valine-citrulline-based linker and the Aur0101 auristatin microtubule inhibitor. The anti-PTK7 ADC induced sustained regressions of TNBC, NSCLC and ovarian cancer PDX, with improved activity over standard-of-care chemotherapy, and reduced the frequency of TICs as determined by serial transplantation experiments. Moreover, the ADC may have additional mechanisms of action, including an anti-angiogenic effect, that promote anti-tumor immune responses. Together these preclinical results indicate the potential of the anti-PTK7 ADC to improve the long-term survival of cancer patients. The ADC is currently being tested in a Phase 1 clinical trial, from which interim results will be presented. Citation Format: Marc Isaac Damelin, Alex Bankovich, Jeff Bernstein, Justin Lucas, Liang Chen, Sam Williams, Albert Park, Jorge Aguilar, Elana Ernstoff, Manoj Charati, Russell Dushin, Amy Jackson-Fisher, Monette Aujay, Christina Lee, Hanna Ramoth, Milly Milton, Johannes Hampl, Sasha Lazetic, Virginia Pulito, Douglas Armellino, Edward Rosfjord, Magali Guffroy, Hadi Falahatpisheh, Lindsay King, Frank Barletta, Robert Stull, Marybeth Pysz, Paul Escarpe, David Liu, Orit Foord, Brenda Gibson, Eric Powell, Christopher O’Donnell, Xiaohua Xin, Hans Peter Gerber, Puja Sapra, Scott Dylla. A novel PTK7-targeted antibody-drug conjugate eliminates tumor-initiating cells and induces sustained tumor regressions. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1220.

Abstract 5425: An anti-Ephrin-A4 calicheamicin conjugate effectively targets triple-negative breast and ovarian tumor-initiating cells to result in sustained tumor regression

Triple-negative breast cancer (TNBC) and ovarian cancer comprise heterogeneous tumors, and neither targeted therapies nor traditional chemotherapies have provided consistent clinical benefit. Novel therapies that target and actively eradicate the subpopulation of tumor cells that mediate drug resistance and tumor relapse could significantly improve patient survival. Tumor-initiating cells (TIC) are functionally defined as the subpopulation of cells that drive long-term tumor growth, resistance to therapy and disease relapse. We herein identified CD324 as a surface antigen able to reproducibly enrich for TIC in well annotated, low passage TNBC and ovarian cancer patient-derived xenografts (PDXs). Gene expression analysis of TIC led to the identification of Ephrin-A4 as a prospective therapeutic TIC target. Humanized Ephrin-A4-specific monoclonal antibodies (mAbs) were generated and demonstrated to internalize to mediate the delivery of potent cytotoxins. An antibody-drug conjugate (ADC) comprising a humanized anti-Ephrin-A4 mAb conjugated to the DNA damaging agent calicheamicin achieved sustained tumor regressions in vivo in both TNBC and ovarian cancer PDX. Anti-Ephrin-A4-ADC (PF-06647263) actively reduced TIC frequency as evidenced by limiting dilution analysis in serial transplantation assays. Unexpectedly, TNBC tumors of the non-Claudin low molecular subtype exhibited higher Ephrin-A4 expression and more robust responses to the ADC than other breast cancer subtypes, which suggests a specific translational application for breast tumor subtype classification. Together these findings demonstrate the potential of the Ephrin-A4-targeted calicheamicin conjugate as a first-in-class compound designed to eradicate TIC and improve long-term survival of cancer patients. PF-06647263 is currently being evaluated in a Phase I clinical trial. Citation Format: Marc Damelin, Alex Bankovich, Albert Park, Jorge Aguilar, Wade Anderson, Marianne Santaguida, Sarah Fong, Monette Aujay, Kiran Khandke, Virginia Pulito, Elana Ernstoff, Paul Escarpe, Jeff Bernstein, Marybeth A. Pysz, Wenyan Zhong, Erik Upeslacis, Judy Lucas, Justin Lucas, Timothy Nichols, Kathryn Loving, Orit Foord, Johannes Hampl, Robert Stull, Frank Barletta, Hadi Falahatpisheh, Puja Sapra, Hans Peter Gerber, Scott J. Dylla. An anti-Ephrin-A4 calicheamicin conjugate effectively targets triple-negative breast and ovarian tumor-initiating cells to result in sustained tumor regression. [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 5425. doi:10.1158/1538-7445.AM2015-5425