VASCULAR ENDOTHELIAL GROWTH FACTORS EXPRESSION AND IMMUNE MICROENVIRONMENT IN COLONIC ADENOCARCINOMA AND ADENOMA
Cesare Ruffolo2, Melania Scarpa1, Elisabetta Trevellin2, Francesco Ferrara3, Ivana Cataldo3, Andromachi Kotsafti1, Marco Scarpa*2
1Oncological Surgery Unit, Veneto Institute of Oncology (IOV-IRCCS), Padova, Italy; 2University Hospital of Padua, Padova, Italy; 3Aulss2 Marca Trevigiana, Treviso, Italy
Background
In advanced colorectal cancer, VEGF-A and its receptors are the target of antiangiogenetic therapy for while immune checkpoints molecules are target of immunotherapy. Thus, the aim of the study was to evaluate the interplay among VEGF-A and VEGF-C and immune checkpoints in colonic adenocarcinoma and in the premalignant carcinogenesis steps.
Methods
The explorative series consisted of gene expression data from 331 colonic adenoarcinoma samples from the The Cancer Genome Atlas (TCGA) databank. In vitro supplementation of VEGF-A and VEGF-C on HTC116 and HT29 cell lines was performed and its effect on CD80, CD86, PDL-1, PDL-2, HLA-ABC expression were tested with FACS analysis. Moreover, 56 subjects with colonic adenoma who underwent screening colonoscopy were prospectively enrolled in the MICCE1 project (Treviso arm). Real time RT-PCR for VEGF-A, VEGF-C and CD34 mRNA expression and immunohistochemistry for CD8 and CD4 T cell infiltration and PD1 and PD-L1 expression were performed. Activated CD8 T cells (CD38 and CD28) and epithelial cells acting as antigen presenting cell (CD80 and HLA-ABC) were quantified with cytofluorimetry.
Results
Co-expression analysis on TCGA colon adenocarcinoma cohort showed an inverse correlation between and VEGF-A expression and HLA-ABC expression on tumour cells (r=-0.14, p=0.007) and direct correlation between VEGF-C and PD-L1 and PD-L2 (r=0.39, p<0.001 and r=0.66, p<0.001). An in vitro model on HTC116 and HT29 cell lines supplemented with recombinant VEGF-A and VEGF-C showed a decrease only of HLA-ABC protein expression (p=0.06 and p=0.15). In colonic adenoma with high grade dysplasia (HGD) high VEGF-A mRNA expression was associated to a high CD8+ lymphocyte infiltration (p=0.005). Similarly, in colonic adenoma with HGD, high VEGF-C mRNA expression was associated to a high CD8+ lymphocyte infiltration and a higher rate of epithelial cells expressing PD-L1 (p=0.01 and p=0.06, respectively).
Conclusions
This study demonstrated that in colonic adenocarcinoma VEGF-A and VEGF-C are associated to a decrease in HLA-ABC expression and this decrease was mechanistically confirmed in vitro. Despite being associated to high CD8 tumour infiltration VEGF-C expression was associated to an increase of PD-L1 and PD-L2 either in colonic adenocarcinoma or in HGD. Thus, VEGF-A and VEGF-C expression are involved in the tumour immune escape. Therefore, this study poses the basis of a possible combination of angiogenesis inhibitors and immune check point inhibitors in the treatment of colorectal cancer.
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