3.1.Characteristics of the of the Patients
537 patients’ clinical data were acquired from TCGA, including the age, gender, Histological grade, TNM classification of KIRC(Table 1).
3.2.High HSPA7 mRNA Expression in KIRC
First, we assessed the differences in HSPA7 expression between KIRC tumor tissues and adjacent tissues via differential expression scatter plots and paired difference analyses. We find that the expression level of HSPA7 was significantlly higher in KIRC tumor tissues (p=6.183e-35) and in paired cancer tissues(p=3.311e-18) compared with adjacent tissues(Figure 1A and 1B). Then, the expression level of HSPA7 in KIRC tumor tissues and adjacent tissues were verified by GEPIA (Figures 1C) database, UALCAN database(Figures 1D) and qRT-PCR analysis(Figures 1E) .
3.3. Correlation Between HSPA7 Expression level and Clinico-pathological featuresin KIRC tumors
As the Table 2 shown the expression of HSPA7 was highly statistically significantly correlated with clinical stage (p = 0.044) and distant metastasis (positive vs. negative, p = 0.049) .
3.4.Correlation Between KIRC patients Survival and HSPA7 Expression
To evaluate the effect of HSPA7 expression on KIRC patients survival, the log-rank test and Kaplan-Meier survival analysis were used to estimate the correlation between HSPA7 expression and KIRC patients prognosis. The patients with high HSPA7 expression level displayed relatively poor survival (p=1.176e−04; Figure 2A). The clinical subgroup analysis implied that the patients in Histological grade (G1–2 vs. G3–4) , clinical stage(Stage I vs Stage IV), M classification and T classification (the T1–2 vs. T3–4) with HSPA7 expression also had significantly poor overall survival(OS) (Figure 2B-E) , whereas not in the N classification (Figures 2F). We performed the univariate analysis with the variables and listed in the Table 3. We also performed Multivariate analysis with the Cox proportional hazards model and the results implied that the expression of HSPA7 (HR =1.304605, p =0.005187) is a potential prognostic factor for KIRC patients (Table 4). Then we performed the forest plot analysis (Figure 3), the outcome of KIRC patients are statistically significant correlation with age (p <0.001), histological grade(p=0.002), clinical stage (p=0.019) and the expression of HSPA7 (p <0.001). In conclusion, HSPA7 is a reliable and effective independent prognostic biomarker of KIRC patients.
3.5. HSPA7-Related Signaling Pathway was Identified by GSEA
The differentially regulated pathways between high and low expression of HSPA7 groups were identified by GSEA and then the activated signaling pathways in KIRC were founded. The results with significant differences in enrichment (FDR < 0.25, NOM p < 0.05) in the MSigDB gene set (c2.cp.kegg.v6.2.symbols.gmt) were selected based on the NES and listed in Table 5. Figure 4 showed that renin angiotensin system, primary immunodeficiency,O-glycan biosynthesis, JAK-STAT signaling pathway, hematopoietic cell lineage, intestinal immune network for IgA production, glycerophospholipid metabolism, cytokine-cytokine receptor interaction, cytosolic DNA sensing pathway,autoimmune thyroid disease and asthma. GSEA analysis displayed the high expression of HSPA7 in KIRC were related to several tumor-related and immune-related pathways.
3.6.HSPA7 expression correlated with immune cell infiltration in KIRC
Previous studies showed that lymph node metastasis and survival are independently predicted by the frequency of lymphocytes infiltrating in cancer patients. Also GSEA analysis displayed the high expression of HSPA7 in KIRC were related to immune-related pathways.Using TIMER database we investigated whether HSPA7 expression was correlated with six main infiltrating immune cells in KIRC. The result implied that expression of HSPA7 associated with CD4+ T cells (r=0.395, p-value=1.24e−18), Macrophage (r = 0.216, p-value = 3.97e-06), neutrophils (r= 0.335, p-value=1.88e−13) and Dendritic Cell（DC）(r= 0.212, p-value =4.86e−06) (Figure 5). The HSPA7 expression levels was also correlated with tumor purity (cor=0.125, p-value=6.98e−03).These results suggested that immune infiltration may searve as a important role in KIRC patient outcomes, and HSPA7 could modulate immune infiltrating cells into KIRC tissues.