PDL1 test to assess the dynamic range of mRNA expression from platelet enriched plasma in patients with NSCLC.

Abstract

e23063 Background: Therapeutic options for patients with non-small cell lung cancer (NSCLC) continue to expand with the advent of immunotherapies. Lack of tissue and drawbacks with IHC have increased the need for blood-based diagnostics that measure continuous variables. Thus these tests are of increasing relevance for clinical testing. Methods: We focused on mechanisms of blood-based testing for the sensitive measurement of circulating RNA using ddPCR. Specifically, we optimized methods for the detection of PD-L1 and cytokeratin 19 (CK19) transcripts recovered from plasma. Specimens included tumor derived cell-lines, activated and resting immune cells, normal donor plasma and NSCLC (stages 1-4) donor plasma. Results: Assessing PD-L1 in circulation is complicated by its expression in both immune and cancer cells. Analytic performance was initially evaluated with cancer cell lines and activated lymphocytes and monocytes expressing variable levels of cytokeratins and PD-L1. A preliminary threshold was defined based on CK19 expression levels in normal donor plasma (n = 35). Of the 79 NSCLC donors tested 49% expressed CK19 above the threshold, indicating they contained sufficient circulating RNA derived from tumor. These donor samples tested either positive for EGFR del19 E746-A750 or L858R (n = 13), negative for EGFR (n = 10) or positive for KRAS G12C/V (n = 16). We found no differentiation in PD-L1 levels on the basis of EGFR sensitizing and EGFR wild-type status. We observed only three donors that expressed PD-L1 at high levels, and this was regardless of EGFR mutation status. In contrast, 30% of KRAS mutation-positive donor samples were above the pre-defined PDL1 threshold (n = 6 of 16 samples tested). These data are consistent with tissue-based studies that report trends of higher PD-L1 expression in KRAS positive NSCLC patients. Conclusions: We have developed sensitive and specific methods to measure the dynamic range of PD-L1 in circulation and have shown utility of these methods by evaluating key immune and cancer-specific RNAs.