Mien Sho

Antibodies to TWEAK Receptor Inhibit Human Tumor Growth through Dual Mechanisms

Purpose: Targeted therapeutics have significantly changed the outcome for patients diagnosed with cancer. Still, effective therapeutic intervention does not exist for many cancers and much remains to be done. The objective of this study was to identify novel genes that potentially regulate tumor growth, to target these gene products with monoclonal antibodies, and to examine the therapeutic potential of these antibodies. Experimental Design: Using cDNA microarray analysis, we identified genes overexpressed in several solid malignancies. We generated a mouse monoclonal antibody, 19.2.1, and its humanized counterpart, PDL192, to one such target, TweakR (TWEAK receptor, Fn14, TNFRSF12A, CD266), and characterized the antitumor activities in vitro and in mouse xenograft models. Results: Both 19.2.1 (mouse IgG2a) and PDL192 (human IgG1), like TWEAK, the natural ligand of TweakR, inhibited the growth of several TweakR-expressing cancer cell lines in anchorage-dependent and anchorage-independent assays in vitro. Both antibodies showed significant antitumor activity in multiple mouse xenograft models. PDL192 and 19.2.1 also induced antibody-dependent cellular cytotoxicity (ADCC) of cancer cell lines in vitro. A chimeric version of 19.2.1 containing the mouse IgG1 Fc region (19.2.1×G1) exhibited significantly less ADCC than 19.2.1. However, 19.2.1×G1 showed differential activity in vivo, with activity equivalent to 19.2.1 in one model, but significantly less efficacy than 19.2.1 in a second model. These results indicate that PDL192 and 19.2.1 mediate their antitumor effects by signaling through TweakR, resulting in reduced tumor cell proliferation, and by ADCC. Clin Cancer Res; 16(2); 497–508

Expression of TweakR in breast cancer and preclinical activity of enavatuzumab, a humanized anti-TweakR mAb

BackgroundThe receptor for the cytokine TWEAK (TweakR) is a cell surface member of the tumor necrosis factor receptor superfamily with diverse biological roles. TNFRSF family members are appealing therapeutic targets in oncology due to their aberrant expression and function in tumor cells. The goal of the current study was to examine the potential of TweakR as a therapeutic target in breast cancer.MethodsExpression of TweakR in primary breast cancer tissues and metastases was characterized using immunohistochemistry. To determine the functional relevance of TweakR, breast cancer cell lines were treated in vitro and in vivo with enavatuzumab, a humanized mAb against TweakR.ResultsOverexpression of TweakR was observed in infiltrating tumors compared to normal adjacent breast tissues, and strong staining of TweakR was observed in all subtypes of invasive ductal breast cancer. In addition, a positive correlation of TweakR and HER2 expression and co-localization were observed, irrespective of ER status. TweakR expression was also observed in bone metastasis samples from primary breast cancer but rarely in benign tumors. Enavatuzumab inhibited the in vitro growth of TweakR-expressing breast cancer cell lines, and this activity was augmented by cross-linking the mAb. In addition, enavatuzumab significantly inhibited the in vivo growth of multiple breast cancer xenograft models including a model of metastasis.ConclusionsTweakR is highly expressed in all subtypes of invasive ductal breast cancer, and enavatuzumab administration exhibited a dose-dependent inhibition of primary tumor growth and lung metastasis and enhanced the antitumor activity of several chemotherapy agents currently used to treat breast cancer. These data provide the rationale to evaluate enavatuzumab as a potential therapy for the treatment of breast cancer.

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 TGFβ 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-085s 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.

Abstract 2722: Active recruitment of immune effector cells mediates in vivo tumor growth inhibition by enavatuzumab, an antibody to human TWEAK receptor

Proceedings: AACR 103rd Annual Meeting 2012‐‐ Mar 31‐Apr 4, 2012; Chicago, IL Enavatuzumab (PDL192) is a humanized anti-TWEAK receptor antibody currently under clinical evaluation for the treatment of solid malignancies. Enavatuzumab is postulated to exert its potent anti-tumor activity in xenograft models through two distinct mechanisms of action: direct inhibition of tumor cell growth and Fc-mediated function. The goal of this study was to elucidate the role of immune effector cells in Fc-mediated function. The initial evidence for a role of effector cells was obtained in a subset of tumor xenograft mouse models which responded to enavatuzumab but did not respond to a version of enavatuzumab containing a mutation in the FcαRIII binding region. Subsequent analysis of circulating effector cells showed increased expression of activation markers DX5 and CD27 on monocyte-like cells (CD45+CD11bHigh) and DX5 on NK-like cells (CD45+CD11bLow) after enavatuzumab treatment in xenograft models dependent on Fc-FcR interactions for enavatuzumab efficacy. The involvement of effector cells was further confirmed by immunohistochemistry, which revealed strong infiltration of CD45+ effector cells into tumor xenografts in responding models, but minimal infiltration in non-responders. Consistent with the in vivo xenograft studies, human effector cells migrated towards enavatuzumab-treated tumor cells in vitro. Effector cells preferentially migrated toward in vivo responsive tumor cells, and the majority of migratory cells were monocytes and NK cells. Conditioned media from enavatuzumab-treated tumor cells also induced migration, and contained elevated levels of cytokines/chemokines, including IL8 and MCP-1. The presence of an anti-MCP-1 neutralizing antibody in enavatuzumab-treated tumor cell conditioned medium significantly blocked effector cell migration, suggesting that MCP-1 might be responsible for effector cell migration triggered by enavatuzumab. We also observed elevated levels of human MCP-1 in the serum of both enavatuzumab-treated xenograft mouse models and in a subset of patients from the enavatuzumab Phase I clinical study. In summary, in vivo and in vitro studies suggest that enavatuzumab exerts its potent anti-tumor activity, in part, by actively recruiting effector cells and activating effectors to kill tumor cells. In addition, enavatuzumab-induced chemokines may be potential pharmacodynamic biomarkers and warrant further evaluation in clinical studies. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2722. doi:1538-7445.AM2012-2722

Enavatuzumab, a Humanized Anti-TWEAK Receptor Monoclonal Antibody, Exerts Antitumor Activity through Attracting and Activating Innate Immune Effector Cells

Enavatuzumab is a humanized IgG1 anti-TWEAK receptor monoclonal antibody that was evaluated in a phase I clinical study for the treatment of solid malignancies. The current study was to determine whether and how myeloid effector cells were involved in postulated mechanisms for its potent antitumor activity in xenograft models. The initial evidence for a role of effector cells was obtained in a subset of tumor xenograft mouse models whose response to enavatuzumab relied on the binding of Fc of the antibody to Fcγ receptor. The involvement of effector cells was further confirmed by immunohistochemistry, which revealed strong infiltration of CD45+ effector cells into tumor xenografts in responding models, but minimal infiltration in nonresponders. Consistent with the xenograft studies, human effector cells preferentially migrated toward in vivo-responsive tumor cells treated by enavatuzumab in vitro, with the majority of migratory cells being monocytes. Conditioned media from enavatuzumab-treated tumor cells contained elevated levels of chemokines, which might be responsible for enavatuzumab-triggered effector cell migration. These preclinical studies demonstrate that enavatuzumab can exert its potent antitumor activity by actively recruiting and activating myeloid effectors to kill tumor cells. Enavatuzumab-induced chemokines warrant further evaluation in clinical studies as potential biomarkers for such activity.

Abstract B17: PDL192, a humanized antibody to TweakR exhibits potent antitumor activity in pancreatic cancer models

Approximately 42,000 new cases of pancreatic cancer were diagnosed in the United States in 2009, and the 5-year survival rate for this disease is exceedingly low (approximately 5%). Given the lack of therapeutic success with current standards of care, new therapies are clearly needed to treat this disease. We have generated PDL192, a humanized IgG1 antibody to TweakR (Fn14, TNFRSF12A, CD266), a cell surface protein and member of the TNF receptor superfamily. PDL192 has been shown to exhibit antitumor activity in xenograft models on a range of solid tumor types via both direct tumor cell growth inhibition and by antibody-dependent cellular cytotoxicity. In this study, we explored the activity of PDL192 in pancreatic cancer models. Approximately 60% of primary pancreatic cancers express TweakR protein at high levels, as determined by immunohistochemistry. PDL192 exhibited potent anti-tumor activity in two cell line xenograft models, Panc1 and MiaPaCa2. PDL192 also exhibited significant anti-tumor activity against 3 of 6 primary tumor xenograft models derived from pancreatic patients. PDL192 was also tested in combination with gemcitabine, the most common treatment for pancreatic cancer patients. In the Panc1 and MiaPaCa2 xenograft models, the combination of PDL192 and gemcitabine exhibited more potent activity than either agent alone. In three of five primary tumor xenograft models tested, combining PDL192 and gemcitabine also resulted in greater anti-tumor activity than either agent alone. In one of the primary tumor models, where neither PDL192 nor gemcitabine alone exhibited significant anti-tumor activity, the combination completely inhibited tumor growth. The activity of PDL192 in multiple pancreatic xenograft models using both cell lines and primary tumors is supportive of the use of PDL192 for the treatment of pancreatic cancer. PDL192 is currently being evaluated in a phase1 safety study in patients with solid tumors. Citation Information: Clin Cancer Res 2010;16(14 Suppl):B17.

Abstract B231: Targeting Her2+ ductal breast cancer with PDL192, a novel humanized anti‐TweakR monoclonal antibody

TWEAK receptor (TweakR), also known as Fn14, is a cell surface protein and member of the tumor necrosis factor receptor superfamily. TNFRSF family members are appealing therapeutic targets due to their expression and function in tumor cells. The goal of the current study was to examine the potential of TweakR as a therapeutic target in various subtypes of breast cancer by 1) determining expression of TweakR using immunohistochemistry (IHC) and 2) treating cells derived from breast cancer in vitro and in vivo with PDL192, a novel, humanized anti‐TweakR mAb. Overexpression of TweakR protein in breast cancer was confirmed by IHC on formalin fixed paraffin embedded (FFPE) tissues. TweakR was elevated in infiltrating tumors when compared with normal adjacent tissues. 30 to 40% of various subtypes of invasive ductal breast cancer samples stained highly for TweakR; however, fewer than 5% of lobular breast cancer samples had high levels of TweakR. In addition, a strong positive correlation of TweakR and Her2 expression was observed, irrespective of ER status. More than 60% of Her2+ invasive ductal cancer exhibited elevated levels of TweakR. Strong TweakR expression was also observed in bone metastasis samples from primary breast cancer but rarely in benign tumors, such as DCIS. PDL192 inhibited the in vitro growth of various subtypes of TweakR‐expressing breast cancer cell lines and this activity was augmented by crosslinking the mAb. In addition, PDL192 significantly inhibited the in vivo growth of a variant of the breast cancer xenograft model MDA‐MB231 that exhibits metastatic potential. PDL192 administration exhibited a dose‐dependent inhibition of primary tumor growth and lung metastasis and enhanced the anti‐tumor activity of several chemotherapy agents currently used to treat breast cancer. These data provide the rationale to evaluate PDL192 as a potential therapy for the treatment of breast cancer. Currently, PDL192 is being tested in a phase I safety study for the treatment of patients with solid tumors. Citation Information: Mol Cancer Ther 2009;8(12 Suppl):B231.