A study on programmed death ligand 1 expression after radiation in colorectal cancer human tissue and in vitro cell line model

Abstract

Purpose: The aim of this study was to evaluate the programmed death-ligand 1 (PD-L1) expression status before and after radiation in colorectal cancer human tissue and in vitro cell line model. Materials and methods: In this study, formalin-fixed paraffin-embedded tissue specimens from pre-operative biopsy via sigmoidoscopy and surgical resection of the primary tumor were obtained from 24 rectal adenocarcinoma patients who underwent neoadjuvant CRT between August 2016 and December 2017. Pre- and post-RT PD-L1 expression level on tumor cell and immune cell was assessed by immunohistochemical (IHC) analysis. The human colorectal cancer cell lines, DLD 1, HT-29, and HCT116 were exposed to graded doses of radiation 2, 4, and 8 Gy for 24 h and PD-L1 expression was analyzed using fluorescence-activated cell sorting analysis. Results: Patients were classified, according to their tumor regression grade, as responders (grade 2; 9 patients, 37.5%) and non-responders (grade 3, 4, or 5; 15 patients, 62.5%). In the non-responder group, low PD-L1 expression in tumor cells was observed in 13 patients, whereas the reverse pattern was noted in 7 patients (53.9%) after CRT (McNemar test, p = 0.034). There were more patients with advanced T stage in the non-responder group (p = 0.033), and the relative risk of responder after CRT was 2.99 times higher in patients with microsatellite instability (MSI)-high than in patients with microsatellite stable or MSI-low status (relative risk = 2.99, 95% CI = 1.510–5.921). After irradiation at 8 Gy, PD-L1 expression increased 1.5- and 1.7-fold in DLD-1 and HCT-116 cells, respectively. There was no significant change in the low basal level of PD-L1 expression in HT-29 cells. Conclusion: This prospective study verified the CRT-induced immune-oncologic shift toward increased PD-L1 expression in tumor cells and provided the theoretical framework for a combined treatment strategy that involves CRT with immunomodulation followed by administering an immune checkpoint inhibitor for locally advanced rectal cancer. 목적: 방사선 치료는 직접적으로 종양에 방사선을 가함으로써, 종양의 크기를 줄이고, 종양의 국소 치료 효과를 높인다. 본 연구는 진행성 직장암 환자들의 암조직과 대장암 세포주에서 방사선 치료 조사 전과 후의 세포사멸 수용체-1의 발현에 대해 알아보고자 하였다. 재료 및 방법: 2016년 8월부터 2017년 12월까지 국소성 진행성 직장암으로 진단되어 수술 전 항암방사선 치료를 시행 받은 환자들의 치료 전후 암조직을 채취하여 종양세포 및 면역세포들에 대한 면역조직화학염색을 시행하여 세포사멸 수용체-1 발현의 정도를 분석하였다. 대장암 세포주 DLD 1, HT-29, HCT116에 대하여 2,4,8 Gy의 방사선을 조사한 24시 간 후 형광이용세포분류를 이용하여 단일세포 표면의 세포사멸 수용체-1발현 분석하였다. 결과: 환자들을 종양의 사멸정도에 따라 분류하였을 때 반응군은 37.5%였고 비 반응군는 62.5%였다. 항암방사선 치료 전 비 반응군의 종양에서 낮은 세포사멸 수용체-1 발현을 보였던 13명의 환자 중 7명 (53.9%) 의 환자에서 치료 후 높은 발현으로 역전되었다 (McNemar test, p = 0.034).