Impact of radiotherapy on the immune landscape in oesophageal adenocarcinoma

Abstract

BACKGROUND In the contemporary era of cancer immunotherapy, an abundance of clinical and translational studies have reported radiotherapy (RT) and immunotherapies as a viable option for immunomodulation of many cancer subtypes, with many related clinical trials ongoing. In locally advanced disease, chemotherapy or chemoradiotherapy followed by surgical excision of the tumour remain the principal treatment strategy in oesophageal adenocarcinoma (OAC), however, the use of the host immune system to improve anti-tumour immunity is rapidly garnering increased support in the curative setting. AIM To immunophenotype OAC patients’ immune checkpoint (IC) expression with and without radiation and evaluate the effects of checkpoint blockade on cell viability. METHODS In the contemporary era of cancer immunotherapy, an abundance of studies have demonstrated that combination RT and IC inhibitors (ICIs) are effective in the immunomodulation of many cancer subtypes, with many related clinical trials ongoing. Although surgical excision and elimination of tumour cells by chemotherapy or chemoradiotherapy remains the gold standard approach in OAC, the propagation of anti-tumour immune responses is rapidly garnering increased support in the curative setting. The aim of this body of work was to immunophenotype OAC patients’ IC expression with and without radiation and to establish the impact of checkpoint blockade on cell viability. This study was a hybrid combination of in vitro and ex vivo models. Quantification of serum immune proteins was performed by enzyme-linked immunosorbent assay. Flow cytometry staining was performed to evaluate IC expression for in vitro OAC cell lines and ex vivo OAC biopsies. Cell viability in the presence of radiation with and without IC blockade was assessed by a cell counting kit-8 assay. RESULTS We identified that conventional dosing and hypofractionated approaches resulted in increased IC expression (PD-1, PD-L1, TIM3, TIGIT) in vitro and ex vivo in OAC. There were two distinct subcohorts with one demonstrating significant upregulation of ICs and the contrary in the other cohort. Increasing IC expression post RT was associated with a more aggressive tumour phenotype and adverse features of tumour biology. The use of anti-PD-1 and anti-PD-L1 immunotherapies in combination with radiation resulted in a significant and synergistic reduction in viability of both radiosensitive and radioresistant OAC cells in vitro. Interleukin-21 (IL-21) and IL-31 significantly increased, with a concomitant reduction in IL-23 as a consequence of 4 Gray radiation. Similarly, radiation induced an anti-angiogenic tumour milieu with reduced expression of vascular endothelial growth factor-A, basic fibroblast growth factor, Flt-1 and pl...

Analysing the Combined Effects of Radiotherapy and Chemokine Receptor 5 Antagonism: Complementary Approaches to Promote T Cell Function and Migration in Oesophageal Adenocarcinoma

“…Additional studies also identified that irradiation stimulates tumour cells and immune cells to secrete RANTES and MIP-1α [20], which suggests that irradiation might promote the trafficking of T cells to OAC tumours. In a previous study, we observed that irradiating OAC tumour biopsy explants ex vivo had favourable effects on the secretome, significantly increasing the secretion of anti-tumour cytokines IL-21 and IL-31 [21] and decreasing the production of a tumour-promoting cytokine IL-23 [21]. In addition, radiation induced an anti-angiogenic tumour milieu by reducing the secretion of VEGF-A, BFGF, Flt-1, and PIGF pro-angiogenic factors [21].…”

“…In a previous study, we observed that irradiating OAC tumour biopsy explants ex vivo had favourable effects on the secretome, significantly increasing the secretion of anti-tumour cytokines IL-21 and IL-31 [21] and decreasing the production of a tumour-promoting cytokine IL-23 [21]. In addition, radiation induced an anti-angiogenic tumour milieu by reducing the secretion of VEGF-A, BFGF, Flt-1, and PIGF pro-angiogenic factors [21].…”

Tumour microenvironment influences response to treatment in oesophageal adenocarcinoma

“…Several studies found immune checkpoints, such as PD-1, CTLA-4, TIGIT, TIM-3, LAG-3 or ICOS, to be significantly upregulated in infiltrating T cells following NAC ( 46 , 47 ). An OAC cell line study validated ex-vivo also suggested increased expression of several immune checkpoint expression following radiotherapy ( 48 ). Soeratram et al.…”

Radiotherapy remodels the tumor microenvironment for enhancing immunotherapeutic sensitivity

“…RNA sequencing and flow cytometry analysis showed that TIGIT of CD8 + T cells increased on day 7 after 8 Gy × 3 RT while 2 Gy × 18 irradiation treatment resulted in a drop in TIGIT expression over time (day 7, day 14 and day 30) [ 120 ]. In two cohorts of esophageal cancer, the expression of PD-1, PD-L1, TIGIT and TIM-3 was downregulated in CD3 + CD4 + T and CD3 + CD8 + T cells in patients well responding to 2 and 4 Gy RT while PD-1, PD-L1 and TIGIT were elevated in those poorly [ 127 ]. 10 Gy RT combined with αPD-L1 led to a 2- and 2.4-fold upsurge in the expression of TIM-3 in CD8 + T cells and Tregs, respectively, indicating upregulation of TIM-3 mediated resistance to PD-L1 blockade [ 122 ].…”