Stage 5 (January 2022- November 2022)
In stage 5, the main changes induced at the level of signaling pathways were analyzed in co-cultures of gastric cancer cells with increased expression of PD-L1 with human T lymphocytes with PD1 expression, in the presence of inhibitors of the PD1/PD- L1 interaction and VEGFR2.
The results of the experiments indicated a reduction in signal transmission on the Ras/MAPK/ERK, Ras/MAPK/JNK, PI3K/AKT/mTOR/p70 S6 kinase, STAT3 and STAT6 signaling pathways, especially in the case of using the PD1/PDL1 interaction inhibitor which attests to the beneficial effect of immunotherapy in gastric cancer with PDL1 positive cells.
Treatment with VEGFR2 inhibitor indicated an increase in the inhibitory effect especially on the PI3K/AKT/mTOR/p70 S6 kinase pathway. This functional model can be used in further studies to validate other immune-checkpoints (LAG3, TIM3, IDO, BTLA) as targets for therapy in gastric tumor cells.
At the same time, the recruitment of patients in the prospective group and the biobanking of normal and tumor tissue, plasma, serum and buffy coat continued. In stage 5, 24 patients with gastric adenocarcinoma (ADK) were included in the study, so up to now a biobank has been created with a prospective group of 121 patients in total.
Until stage 5, from the entire prospective group, 62 patients with ADK of the stomach were classified into the 5 molecular subtypes based on the expression level of the 14 specific markers, being evaluated by in situ hybridization to determine the EBER status and by immunohistochemistry to determine the level of protein expression. In addition, in the prospective group, the expression level of the PDL1 gene was evaluated in 71 patients with ADK of the stomach by qRT-PCR, and the results showed that the expression level of PD-L1 is significantly increased in the tumor compared to the non-adjacent tissue – tumor.
Post-translational studies of vesicular traffic in cancer, through the colocalization of E-cadherin and LAMP-1 at the tissue level, revealed a dynamic profile of the epithelial-mesenchymal transition process, thus highlighting the progression and metastasis of gastric cancer.
The following immune targets were analyzed by IHC in the prospective group, which includes a number of 62 patients with stomach adenocarcinoma: LAG-3, TIM-3, CTLA4, PD-L1, PD1, CD4 and CD8. The results show that by analyzing the prognostic role of IHC checkpoint immune markers, the group of patients with LAG-3 with a negative IHC status have a significantly lower survival rate than the group with positive LAG-3 IHC. Based on the results obtained in the study, LAG-3 could represent another important immune target in gastric cancer, the IHC results confirming the presence of this protein in 80% of the analyzed samples.