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

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.

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.

Long-term tumor regression induced by an antibody-drug conjugate that targets 5T4, an oncofetal antigen expressed on tumor-initiating cells.

Antibody-drug conjugates (ADC) represent a promising therapeutic modality for the clinical management of cancer. We sought to develop a novel ADC that targets 5T4, an oncofetal antigen expressed on tumor-initiating cells (TIC), which comprise the most aggressive cell population in the tumor. We optimized an anti-5T4 ADC (A1mcMMAF) by sulfydryl-based conjugation of the humanized A1 antibody to the tubulin inhibitor monomethylauristatin F (MMAF) via a maleimidocaproyl linker. A1mcMMAF exhibited potent in vivo antitumor activity in a variety of tumor models and induced long-term regressions for up to 100 days after the last dose. Strikingly, animals showed pathologic complete response in each model with doses as low as 3 mg antibody/kg dosed every 4 days. In a non-small cell lung cancer patient-derived xenograft model, in which 5T4 is preferentially expressed on the less differentiated tumor cells, A1mcMMAF treatment resulted in sustained tumor regressions and reduced TIC frequency. These results highlight the potential of ADCs that target the most aggressive cell populations within tumors, such as TICs. In exploratory safety studies, A1mcMMAF exhibited no overt toxicities when administered to cynomolgus monkeys at doses up to 10 mg antibody/kg/cycle × 2 and displayed a half-life of 5 days. The preclinical efficacy and safety data established a promising therapeutic index that supports clinical testing of A1mcMMAF.

Tbx3, the ulnar-mammary syndrome gene, and Tbx2 interact in mammary gland development through a p19Arf/p53-independent pathway.

The closely related T-box genes Tbx2 and Tbx3 are both expressed in the developing mammary glands of mouse embryos and both have been implicated in mammary carcinogenesis. Tbx3 is essential for induction of the mammary placodes in mice. In humans, mutations in TBX3 are responsible for ulnar-mammary syndrome. Here, we show a haploinsufficiency effect of Tbx3 on maintenance of the mammary placodes and on the extent of branching of the ductal tree in mice. Loss or heterozygosity for Tbx2, on the other hand, has no effect on either induction or maintenance of the placodes, although a small effect was seen on branching morphogenesis in adult heterozygotes. However, the deficiency in maintenance of the mammary placodes in Tbx2, Tbx3 double heterozygous mice is more marked than in Tbx3 single heterozygotes, indicating a genetic interaction between the two genes. In spite of a large body of evidence implicating these genes in cell cycle control through the p19(Arf)/p53 pathways, we find no evidence for involvement of these pathways either in embryonic lethality of homozygous mutants or in the mammary gland phenotype of Tbx3 heterozygous mice.