Bladder cancer (BlCa) represents the sixth most commonly diagnosed type of male
malignancy. Due to the clinical heterogeneity of BlCa, novel markers would optimize treatment efficacy and improve prognosis. The
small heat shock proteins (sHSP) family is one of the major groups of
molecular chaperones responsible for the maintenance of
proteome functionality and stability. However, the role of sHSPs in BlCa remains largely unknown. The present study aimed to examine the association between HSPB2 and HSPB3 expression and BlCa progression in patients, and to investigate their role in BlCa cells. For this purpose, a series of experiments including reverse transcription-quantitative PCR, Western blotting, MTT assay and flow cytometry were performed. Initial analyses revealed increased vs. human transitional
carcinoma cells, expression levels of the HSPB2 and HSPB3 genes and
proteins in high grade BlCa cell lines. Therefore, we then evaluated the clinical significance of the HSPB2 and HSPB3 genes expression levels in
bladder tumor samples and matched adjusted normal bladder specimens. Total
RNA from 100
bladder tumor samples and 49 paired non-cancerous bladder specimens were isolated, and an accurate
SYBR-Green based real-time quantitative polymerase chain reaction (qPCR) protocol was developed to quantify HSPB2 and HSPB3
mRNA levels in the two cohorts of specimens. A significant downregulation of the HSPB2 and HSPB3 genes expression was observed in
bladder tumors as compared to matched normal urothelium; yet, increased HSPB2 and HSPB3 levels were noted in muscle-invasive (T2-T4) vs. superficial
tumors (TaT1), as well as in high-grade vs. low-grade
tumors. Survival analyses highlighted the significantly higher risk for post-treatment disease relapse in TaT1 patients poorly expressing HSPB2 and HSPB3 genes; this effect tended to be inverted in advanced disease stages (muscle-invasive
tumors) indicating the biphasic impact of HSPB2, HSPB3 genes in BlCa progression. The pro-survival role of HSPB2 and HSPB3 in advanced
tumor cells was also evident by our finding that HSPB2, HSPB3 genes expression silencing in high grade BlCa cells enhanced
doxorubicin toxicity. These findings indicate that the HSPB2, HSPB3 chaperone genes have a likely pro-survival role in advanced BlCa; thus, they can be targeted as novel molecular markers to optimize treatment efficacy in BlCa and to limit unnecessary interventions.