Granzyme B is expressed in urothelial carcinoma and promotes cancer cell invasion

Abstract

Granzyme B (GrB) is a serine proteinase known to be expressed by cytotoxic lymphocytes and to induce, in presence of perforin (Pf), apoptosis in target cells. Recently, GrB expression has been shown (often in absence of Pf) in nonlymphoid cells, but its function is not defined. In our study, we investigated GrB and Pf expression in bladder cancer cell lines and in urothelial carcinoma (UC) tissues by reverse transcription-polymerase chain reaction (RT-PCR), Western blot, ELISA, immunofluorescence and immunohistochemistry. We also assessed the function of GrB in UC cells; the in vitro function of GrB was examined by loss-of-function experiments. Our results revealed that GrB is expressed, in absence of Pf, in UC cells. Significant differences were found between GrB expression and both increasing pathological tumor spreading and high-grade vs. low-grade pTa tumors. Notably, GrB in UC tissues was concentrated at the cancer invasion front and was expressed in neoplastic cells undergoing epithelial-mesenchymal transition, a key event in carcinoma invasion. Indeed, GrB-positive cells also expressed Snail, N-cadherin or were negative for E-cadherin. GrB expressed in tumor cell lines was enzymatically active and capable of vitronectin cleavage, implying extracellular matrix (ECM) remodeling by GrB. Inhibition of GrB activity or Stealth RNA interference-mediated GrB gene silencing markedly suppressed bladder cancer cell invasion through matrigel. This data provides the first evidence for a role of GrB in promoting cancer cell invasion. Taken together, our findings suggest that GrB, via ECM degradation, contributes to the establishment of the UC invasive phenotype.

Expression of granzyme B sensitizes ALK+ ALCL tumour cells to apoptosis-inducing drugs

“…These substrates include extracellular matrix (ECM) proteins such as vitronectin, laminin, fibronectin, decorin, and biglycan [5–7]. Cleavage of these proteins by GzB has been implicated in detaching adherent cells bound to these proteins [5], promoting the migration of cells through matrigel [6], and releasing matrix-bound cytokines [7]. …”

“…GzB expression can also be found in cell lines and/or primary tumours of non-lymphoid origin including acute myeloid leukemia [17], prostate cancer [18] and some carcinomas of the breast [19] and urothelium [6]. While the significance, if any, of GzB expression in most of these malignancies has yet to be firmly established, in urothelial carcinoma GzB expression correlates with increased invasiveness and epithelial-to-mesenchymal transition of tumour cells [6]. More importantly, this study demonstrated that in GzB-expressing bladder cancer cell lines, GzB promotes the invasion of these cells through matrigel, presumably through the degradation of ECM proteins within the matrigel [6].…”

“…While the significance, if any, of GzB expression in most of these malignancies has yet to be firmly established, in urothelial carcinoma GzB expression correlates with increased invasiveness and epithelial-to-mesenchymal transition of tumour cells [6]. More importantly, this study demonstrated that in GzB-expressing bladder cancer cell lines, GzB promotes the invasion of these cells through matrigel, presumably through the degradation of ECM proteins within the matrigel [6]. In contrast, GzB expression in some cancers has been reported to be detrimental to tumour cells.…”

Granzyme B-induced apoptosis in cancer cells and its regulation (Review)

“…Compared to the wild-type QPY allele of GZMB gene encoding GrB preproprotein with Gln 48 , Pro 90 , and Tyr 247 in its sequence, the RAH allele of GZMB gene encodes GrB preproprotein containing Arg 48 , Ala 90 , and His 247 (67). The RAH allele occurs at a frequency rate of 25-30% in each of the racial groups studied and it was proven to represent a neutral GZMB gene polymorphism (68).…”

“…Interestingly, GrB has been detected in cancer cells of primary human breast carcinomas (88,89), lung carcinomas (88), urothelial carcinomas (90) and in nasal-type NK/T-cell lymphoma (91). Recently, GrB expression was revealed by immunohistochemistry in oral squamous cell carcinoma and it seems to be localized also in cancer cells (92).…”