Abstract 966: Phase II clinical trial patient responses to the macrophage activating agent RRx-001 correlate to TGF- β pathway activation and markers for fibrosis
Authors: Jha Saheli , Summers Thomas A. , Zeman Karen , Brzezniak Christine , Carter Corey , Ferry Lindsey , Scicinski Jan , Oronsky Bryan , Caroen Scot , Trepel Jane B. , Cabrales Pedro , Day Regina 
Published: July 2017

Abstract
Introduction: RRx-001 is a novel systemically non-toxic small molecule macrophage stimulating agent with promising activity in an ongoing clinical trial in small cell and non-small cell lung cancer and in neuroendocrine tumors. In preclinical studies, RRx-001 activated normally immune-suppressive M2-like Tumor Associated Macrophages (TAMs) to express a series of pro-inflammatory cytokines and chemokines such as transforming growth factor-β1 (TGF-β1), tumor necrosis factor-α TNF-α), inducible nitric oxide synthase (iNOS), and interleukins-10 and -12, a profile that resembles the M1 activated macrophage state. Due to its chemotactic activity for macrophages, TGF-β1 was hypothesized to be involved in the antitumor mechanism of RRx-001. We therefore compared expression patterns of TGF-β1 and TGF-βRI in patient-derived biopsy samples obtained at screening and post RRx-001. As TGF-β1 is closely associated with the induction of fibrosis, we also examined key fibrosis markers.
Methods: Tumor biopsies before and after treatment with RRx-001 were obtained from consented patients with NSCLC, SCLC, and neuroendocrine tumors in the QUADRUPLE THREAT Phase II clinical trial (NCT02489903). Post treatment biopsies were obtained following 6 weeks of once-weekly RRx-001 treatment co-incidental with the first on-study CT scan. Tumor samples were evaluated immune-histochemically with putative markers for TGFβ pathway activation (TGF-β1, TGF-βR1), fibrosis (alpha-smooth muscle actin [α-SMA], Matrix metallopeptidase-9 [MMP-9], and Collagen III deposition), and macrophage activation. Patients were subsequently followed for tumor progression.
Results: Positive immune-histochemical staining for TGF-β1 and TGF-βR1 was seen in all responding patients, and in none of the non-responding patients to date. In responders we also found that MMP-9 and α-SMA activity was down regulated post RRx-001. The down-regulation of these fibrosis markers is suggestive of less invasive malignancy. The number of tumor associated macrophages and their activation was related to the activity of RRx-001.
Conclusions: Our pre and post dose patient derived data to date indicate a correlation between TGF-β signaling activation and a response to RRx-001 that may also correlate with increased numbers of macrophages, and their activation status, in the vicinity of the tumor. These data suggest that activation of the TGF-β1 pathway, as evident by expression of TGF-β1 and TGF-βR1 by tumor cells, could be a predictive biomarker for RRx-001 treatment.
Citation Format: Saheli Jha, Thomas A. Summers, Karen Zeman, Christine Brzezniak, Corey Carter, Lindsey Ferry, Jan Scicinski, Bryan Oronsky, Scot Caroen, Jane B. Trepel, Pedro Cabrales, Regina Day. Phase II clinical trial patient responses to the macrophage activating agent RRx-001 correlate to TGF- β pathway activation and markers for fibrosis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 966. doi:10.1158/1538-7445.AM2017-966