LRRC15 Is a Novel Mesenchymal Protein and Stromal Target for Antibody-Drug Conjugates.

Progress in understanding tumor stromal biology has been constrained in part because cancer-associated fibroblasts (CAF) are a heterogeneous population with limited cell-type-specific protein markers. Using RNA expression profiling, we identified the membrane protein leucine-rich repeat containing 15 (LRRC15) as highly expressed in multiple solid tumor indications with limited normal tissue expression. LRRC15 was expressed on stromal fibroblasts in many solid tumors (e.g., breast, head and neck, lung, pancreatic) as well as directly on a subset of cancer cells of mesenchymal origin (e.g., sarcoma, melanoma, glioblastoma). LRRC15 expression was induced by TGFß on activated fibroblasts (αSMA+) and on mesenchymal stem cells. These collective findings suggested LRRC15 as a novel CAF and mesenchymal marker with utility as a therapeutic target for the treatment of cancers with LRRC15-positive stromal desmoplasia or cancers of mesenchymal origin. ABBV-085 is a monomethyl auristatin E (MMAE)-containing antibody-drug conjugate (ADC) directed against LRRC15, and it demonstrated robust preclinical efficacy against LRRC15 stromal-positive/cancer-negative, and LRRC15 cancer-positive models as a monotherapy, or in combination with standard-of-care therapies. ABBV-085's unique mechanism of action relied upon the cell-permeable properties of MMAE to preferentially kill cancer cells over LRRC15-positive CAF while also increasing immune infiltrate (e.g., F4/80+ macrophages) in the tumor microenvironment. In summary, these findings validate LRRC15 as a novel therapeutic target in multiple solid tumor indications and support the ongoing clinical development of the LRRC15-targeted ADC ABBV-085.Significance: These findings identify LRRC15 as a new marker of cancer-associated fibroblasts and cancers of mesenchymal origin and provide preclinical evidence for the efficacy of an antibody-drug conjugate targeting the tumor stroma. Cancer Res; 78(14); 4059-72. ©2018 AACR.

Preclinical evaluation of an anti-alpha5beta1 integrin antibody as a novel anti-angiogenic agent.

Integrin alpha5beta1, the principal fibronectin receptor, is an important survival factor, playing a key role in angiogenesis. Angiogenesis is critical for tumor growth, and anti-angiogenic therapies have met clinical success. To validate the therapeutic potential of an anti-alpha5beta1 strategy, we generated volociximab (M200) a chimeric human IgG4 version of the alpha5beta1 function-blocking murine antibody IIA1; and F200, the Fab derivative. Volociximab, F200 and IIA1 showed similar activity by ELISA (EC50= 0.2nM), Biacore (Kd= 0.1-0.4nM) and inhibition of fibronectin binding (IC50= 2-3nM). The inhibitory potential of alpha5beta1 antibodies was compared to HuMV833, an anti-VEGF antibody. Both volociximab and HuMV833 inhibited HUVEC proliferation (IC50 of volociximab = 0.2-0.5nM; IC50 of HuMV833 = 45nM). However, IIA1, volociximab and F200 were also potent inhibitors of an in vitro model of angiogenesis (HUVEC tube formation assay), unlike HuMV833. Additionally, volociximab inhibited in vitro tube formation induced by VEGF and/or bFGF, suggesting a mechanism of action independent of growth factor stimulus. In fact, inhibition of alpha5beta1 function by volociximab induced apoptosis of actively proliferating, but not resting, endothelial cells. Volociximab does not cross-react with rodent alpha5beta1, therefore in vivo validation of an anti-alpha5beta1 approach was conducted in a cynomolgus model of choroidal revascularization. Volociximab and F200 were potent inhibitors of neovessel formation in this model. These data demonstrate that volociximab has therapeutic potential in diseases in which new vessel formation is a component of the pathology.

E-selectin up-regulation allows for targeted drug delivery in prostate cancer.

We have used the Eos Hu03 GeneChip array, which represents over 92% of the transcribed human genome, to measure gene expression in a panel of normal and diseased human tissues. This analysis revealed that E-selectin mRNA is selectively overexpressed in prostate cancer epithelium, a finding that correlated strongly with E-selectin protein expression as assessed by immunohistochemistry. Antibodies against E-selectin that blocked function failed to impede cancer cell growth, suggesting that overexpression of E-selectin was not essential for cell growth. However, a novel auristatin E-based antibody drug conjugate (ADC), E-selectin antibody valine-citrulline monomethyl-auristatin E, was a potent and selective agent against E-selectin-expressing cancer cell lines in vitro, with the degree of cytotoxicity varying with surface antigen density. Interestingly, sensitivity to the ADC differed among cell lines from different tissues expressing similar amounts of E-selectin and was found to correlate with sensitivity to free auristatin E. Furthermore, E-selectin-expressing tumors grown as xenografts in severe combined immunodeficient mice were responsive to treatment with E-selectin antibody valine-citrulline monomethyl-auristatin E in vivo, with more than 85% inhibition of tumor growth observed in treated mice. These findings demonstrate that an E-selectin-targeting ADC has potential as a prostate cancer therapy and validates a genomics-based paradigm for the identification of cancer-specific antigens suitable for targeted therapy.

Preclinical validation of anti-TMEFF2-auristatin E-conjugated antibodies in the treatment of prostate cancer.

Current treatments for advanced stage, hormone-resistant prostate cancer are largely ineffective, leading to high patient mortality and morbidity. To fulfill this unmet medical need, we used global gene expression profiling to identify new potential antibody-drug conjugate (ADC) targets that showed maximal prostate cancer-specific expression. TMEFF2, a gene encoding a plasma membrane protein with two follistatin-like domains and one epidermal growth factor-like domain, had limited normal tissue distribution and was highly overexpressed in prostate cancer. Immunohistochemistry analysis using a specific monoclonal antibody (mAb) to human TMEFF2 showed significant protein expression in 74% of primary prostate cancers and 42% of metastatic lesions from lymph nodes and bone that represented both hormone-naïve and hormone-resistant disease. To evaluate anti-TMEFF2 mAbs as potential ADCs, one mAb was conjugated to the cytotoxic agent auristatin E via a cathepsin B-sensitive valine-citrulline linker. This ADC, Pr1-vcMMAE, was used to treat male severe combined immunodeficient mice bearing xenografted LNCaP and CWR22 prostate cancers expressing TMEFF2. Doses of 3 to 10 mg/kg of this specific ADC resulted in significant and sustained tumor growth inhibition, whereas an isotype control ADC had no significant effect. Similar efficacy and specificity was shown with huPr1-vcMMAE, a humanized anti-TMEFF2 ADC. No overt in vivo toxicity was observed with either murine or human ADC, despite significant cross-reactivity of anti-TMEFF2 mAb with the murine TMEFF2 protein, implying minimal toxicity to other body tissues. These data support the further evaluation and clinical testing of huPr1-vcMMAE as a novel therapeutic for the treatment of metastatic and hormone-resistant prostate cancer.