Abstract
BackgroundWhile programmed death 1 (PD-1) and programmed death-ligand 1 (PD-L1) checkpoint inhibitors have activity in a proportion of patients with advanced bladder cancer, strongly predictive and prognostic biomarkers are still lacking. In this study, we evaluated PD-L1 protein expression on circulating tumor cells (CTCs) isolated from patients with muscle invasive (MIBC) and metastatic (mBCa) bladder cancer and explore the prognostic value of CTC PD-L1 expression on clinical outcomes.MethodsBlood samples from 25 patients with MIBC or mBCa were collected at UCSF and shipped to Epic Sciences. All nucleated cells were subjected to immunofluorescent (IF) staining and CTC identification by fluorescent scanners using algorithmic analysis. Cytokeratin expressing (CK)+ and (CK)−CTCs (CD45−, intact nuclei, morphologically distinct from WBCs) were enumerated. A subset of patient samples underwent genetic characterization by fluorescence in situ hybridization (FISH) and copy number variation (CNV) analysis.ResultsCTCs were detected in 20/25 (80 %) patients, inclusive of CK+ CTCs (13/25, 52 %), CK−CTCs (14/25, 56 %), CK+ CTC Clusters (6/25, 24 %), and apoptotic CTCs (13/25, 52 %). Seven of 25 (28 %) patients had PD-L1+ CTCs; 4 of these patients had exclusively CK−/CD45−/PD-L1+ CTCs. A subset of CTCs were secondarily confirmed as bladder cancer via FISH and CNV analysis, which revealed marked genomic instability. Although this study was not powered to evaluate survival, exploratory analyses demonstrated that patients with high PD-L1+/CD45−CTC burden and low burden of apoptotic CTCs had worse overall survival.ConclusionsCTCs are detectable in both MIBC and mBCa patients. PD-L1 expression is demonstrated in both CK+ and CK−CTCs in patients with mBCa, and genomic analysis of these cells supports their tumor origin. Here we demonstrate the ability to identify CTCs in patients with advanced bladder cancer through a minimally invasive process. This may have the potential to guide checkpoint inhibitor immune therapies that have been established to have activity, often with durable responses, in a proportion of these patients.Electronic supplementary materialThe online version of this article (doi:10.1186/s12885-016-2758-3) contains supplementary material, which is available to authorized users.
Highlights
While programmed death 1 (PD-1) and programmed death-ligand 1 (PD-L1) checkpoint inhibitors have activity in a proportion of patients with advanced bladder cancer, strongly predictive and prognostic biomarkers are still lacking
Assessment of PD-L1 antibody specificity To evaluate the specificity of the PD-L1 circulating tumor cell (CTC) assay, anti-PD-L1 antibody and species-matched isotype controls were tested on high PD-L1 expressing and negative control cell lines (H820 and Colo205, respectively; Fig. 1a)
Membrane-localized staining was observed in H820 cells stained with anti-PD-L1, whereas no staining
Summary
While programmed death 1 (PD-1) and programmed death-ligand 1 (PD-L1) checkpoint inhibitors have activity in a proportion of patients with advanced bladder cancer, strongly predictive and prognostic biomarkers are still lacking. We evaluated PD-L1 protein expression on circulating tumor cells (CTCs) isolated from patients with muscle invasive (MIBC) and metastatic (mBCa) bladder cancer and explore the prognostic value of CTC PD-L1 expression on clinical outcomes. Expression of PD-1 and PD-L1 on cancer cells is hypothesized to allow cancers to evade immune surveillance and eradication. The discovery of this mechanism of resistance has provided the rationale for the development of PD-1 and PD-L1 checkpoint immunotherapy. PD-1 checkpoint immunotherapy is rapidly emerging as a promising option for pre-treated patients with advanced tumors [8,9,10,11]. As of 2016, immunotherapies targeting PD-1 or PD-L1 have been approved by the FDA for the treatment of relapsed/refractory melanoma [19], squamous cell lung cancer [20, 21], non-small cell lung cancer [22], renal cell carcinoma [23], and most recently bladder cancer [24]
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