Abstract
Death Receptor 5 (DR5) agonists demonstrate anti-tumor activity in preclinical models but have yet to demonstrate robust clinical responses. A key limitation may be the lack of patient selection strategies to identify those most likely to respond to treatment. To overcome this limitation, we screened a DR5 agonist Nanobody across >600 cell lines representing 21 tumor lineages and assessed molecular features associated with response. High expression of DR5 and Casp8 were significantly associated with sensitivity, but their expression thresholds were difficult to translate due to low dynamic ranges. To address the translational challenge of establishing thresholds of gene expression, we developed a classifier based on ratios of genes that predicted response across lineages. The ratio classifier outperformed the DR5+Casp8 classifier, as well as standard approaches for feature selection and classification using genes, instead of ratios. This classifier was independently validated using 11 primary patient-derived pancreatic xenograft models showing perfect predictions as well as a striking linearity between prediction probability and anti-tumor response. A network analysis of the genes in the ratio classifier captured important biological relationships mediating drug response, specifically identifying key positive and negative regulators of DR5 mediated apoptosis, including DR5, CASP8, BID, cFLIP, XIAP and PEA15. Importantly, the ratio classifier shows translatability across gene expression platforms (from Affymetrix microarrays to RNA-seq) and across model systems (in vitro to in vivo). Our approach of using gene expression ratios presents a robust and novel method for constructing translatable biomarkers of compound response, which can also probe the underlying biology of treatment response.
Highlights
Death Receptor 5 (DR5, TNFRSF10B), a receptor for Apo2L ligand or Tumor Necrosis Factor (TNF)–Related Apoptosis Inducing Ligand (Apo2L/TRAIL), signals through apoptotic pathways to induce cell death [1]
We have developed a method termed Gene Ratio Expression Prediction (GREP), to generate classifiers built from gene expression ratios, and applied it to predict responses to DR5Nb1-tetra
To limit computational costs and multiple hypothesis false discovery rates, we focused on 173 genes with reported relevance to DR5 signaling or sensitivity to a DR5 agonist, or involvement in death domain caspase signaling (S2 Table)
Summary
Death Receptor 5 (DR5, TNFRSF10B), a receptor for Apo2L ligand or Tumor Necrosis Factor (TNF)–Related Apoptosis Inducing Ligand (Apo2L/TRAIL), signals through apoptotic pathways to induce cell death [1]. Despite significant anti-tumor activity in preclinical models, efficacy in clinical settings has been disappointing. While these agents have been generally well tolerated, durable responses to monotherapy have been reported in only a few patients [3,4]. In so-called type II cells, caspase-8 cleaves BID to induce mitochondrial-dependent, intrinsic apoptotic signaling [5]. Several inhibitory proteins, such as c-FLIP, which negatively regulates caspase-8, and the IAP and Bcl-2 protein families keep the apoptotic program in check [1]. The heterogeneous expression of these and other pro-survival signaling factors suggests that multiple molecular features might contribute to mediating the response to DR5-mediated apoptosis [6], highlighting the complexity of predicting response
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