Circulating RNAs in advanced colorectal cancer patients reveal increased expression of PD-1 with conventional CRC therapy

Abstract

Background: Colorectal cancer (CRC) is the third most diagnosed cancer among men and second among women globally. Generally, with age, the risk of this cancer grows and is happened by various genetic alterations. The immune system plays an important role against CRC and that provides a new means to CRC therapy. Our research is focused on the relative expression level of ten different immune genes namely IFNc CD 273, CD274, PD-1, b2M, CD3e, CD28, HLA-A, ICAM1, and CTLA 4 which are known to play important role in immune regulation during different cancers.We hypothesize that the expression of the above-mentioned genes will be altered during CRC and therapy will promote the expression of genes involved in the activation of immune system. Methods: RNA was extracted from the whole blood sample of CRC patient (n=50) and normal (n=34) volunteers. Then cDNA was synthesized, and RNase inhibitor was used. Specific primers for all these genes were designed. Later, RT-qPCR has been done using SYBRVR green master mix. Patient demographics were also recorded. All tested genes have been normalized to a housekeeping gene, RPL11. Values of relative expression for all genes were calculated using REST 2009 software. Results: Among screened genes, CD 273, CD274, and CTLA 4 were not expressed while, IFNc, PD-1, b2M, CD3e, CD28, HLA-A, and ICAM 1 found expressed in CRC. After comparing the gene expression of advanced CRC patients with normal control PD-1 found upregulated (P<0.045). Clinicopathological correlation revealed PD-1 is highly expressed (P<0.001) in advance stage of CRC patients upon treatment with both chemotherapy and immunotherapy (capecitabine, oxaliplatin, irinotecan hydrochloride folinic acid, fluorouracil, and bevacizumab). Conclusions: Our study indicates significant induction of PD-1 at the advanced stage of CRC and during therapy. It suggests that conventional therapies for CRC must be modulated and/or a combination of therapy with anti PD-1 drug should be conducted for the better clinical outcome of CRC.