The Differential Expression Of MICA/B Affects The Immune Escape In NSCLC

NKG2D is a key homodimeric activation receptor expressed on the cell surface of almost all NK cells, CD8+T cells and γδ+T cells. NKG2D-NKG2DL signal play a key role in immune responses, especially trigger NK cell responses against tumors. In this study, we checked the expression of NKG2D ligands in non-small cell lung cancer (NSCLC) tissues and normal tissues, and we found that MICA and MICB were the main ligands expressed in NSCLC tissues and normal tissues. The expression of the MICA and MICB were related with the pathology types and gender, but not smoking or TNM stages. Interesting, the mRNA expression of MICA in normal tissues were much higher than in NSCLC tissues and the mRNA expression of MICB showed no significant different in both NSCLC tissues and normal tissues, but MICA/B protein level in NSCLC tissues were much higher than in normal tissues. Then we found that miRNA-20a was highly expressed in normal tissues, which can inhibit the translation of MICA/B and kept a low level of MICA/B in normal tissues. Moreover, the expressions of MICA and MICB were related with gender, and the MICA concentration had a positive correlation with the Estrogen level in NSCLC patients’ surem. In vitro, the mRNA expression of MICA and MICB were significant increased after treated with estrogen. What’s more, the secretion of MICA/B was also promoted by Estrogen, which can down-regulate the NKG2D receptor on the surface of NK92 cells. Further studies showed that Estrogen can enhance the expression of ADAM17, which associated the secretion of MICA/B.1. NKG2D ligands in lung cancer tissueEight ligands have been found to bind to the NKG2D, and trigger the immune response against tumor cells. There are MICA, MICB, ULBP1, ULBP2, ULBP3, ULBP4, RAET1G and RAET1L. NKG2D ligands are widely expressed in kinds of cancer cells, but rarely in normal cells.In this study, we found that MICA and MICB were the main NKG2D ligands expressing in NSCLC tissues and normal tissues, and ULBP1-4were rarely detected in the tissues. MICA and MICB mRNA can be detected in NSCLC tissues and normal tissues, while MICA/B protein in NSCLC tissues was much higher than in normal tissues. Then patients were classed into different groups, according to pathology type, gender, smoking or not, and TNM stages. Then we found that the expression of MICA and MICB in adenocarcinoma patients was much higher than in squamous cell carcinoma patients; and the mRNA expression of MICA in female patients is much higher than in male patients; however, the expression of MICA and MICB has little correlation with smoking and TNM stages.2. miRNA-20a regulates the expression of MICA/BPrevious study showed that MICA and MICB were the main NKG2D ligands expressing in NSCLC tissues and normal tissues. The mRNA expression of MICA in normal tissues were much higher than in NSCLC tissues and the mRNA expression of MICB showed no significant different in both NSCLC tissues and normal tissues. Then immunochemistry analysis showed that MICA/B expression is in a low level in normal tissue, but much higher in NSCLC tissues. Thus, we hypotheses there was a kind of post-transcript mechanism which could inhibit the translation of MICA/B in normal tissues.miRNA is a kind a no coding RNA, which is found and mature in eukaryotic cells. miRNAs are post-transcriptional regulators that bind to complementary sequences on target messenger RNA transcripts (mRNAs), usually resulting in translational repression or target degradation and gene silencing. The miRNA microarray was used to analyze the different expression of miRNAs in normal tissues and NSCLC tissues, and an online database-TargetScan demonstrated that there is a binding site on the3’-UTR of MICA and MICB by miRNA-20a. Then we constructed the luciferase plasmid with MICA3’-UTR and MICB3’-UTR. Con-transfected of the modified luciferase plasmid with miRNA-20a into293T cells, we found that miRNA-20a can directly bind to the MICA3’-UTR and MICB3’-UTR. Additionally, we over-expressed and low-expressed miRNA-20a in lung cancer cell lines. We found that the surface expression of MICA/B was significantly decreased when miRNA-20a was over-expressed, and the surface expression of MICA/B was increased when miRNA-20a was low-expressed. Thus, miRNA-20a might be one of the mechanisms which inhibit the translation of MICA and MICB in normal tissue.3. Estrogen promotes the immune escape mediated by MICA/BMICA and MICB were the main NKG2D ligands in NSCLC tissues and normal tissues, and their expression were related with gender. The mRNA expression of MICA in female patients is much higher than in male patients. Previous studies, we found that ER-β plays an important role in NSCLC, including proliferation, apoptosis and epithelial-mesenehymal transition. Whether estrogen can also play a role to help the lung cancer cells escape the attack from immune system.From the clinical data, we found that MICA/B levels were positive correlated with estrogen concentration in NSCLC patients, what’s more, the expression of Aromatase also have a positive correlated with MICA/B expression in NSCLC tissues. In vitro studies showed that the expression of MICA/B in lung adenocarcinoma cells lines was increased by estrogen treatment. What’s more, estrogen also regulated the secretion of the MICA/B. And this secretion of MICA/B down-regulated the NKG2D receptor on the surface of NK92cells, and impaired the cytotoxicity of NK92cells. Additionally, estrogen enhanced the expression of ADAM17, which associated the secretion of MICA/B. Thus estrogen can increase the expression of MICA/B and also help the secretion of MICA/B, which help the NSCLC cells escaping the immune response mediated by NKG2D-NKG2DL.