Genome-wide in vivo RNAi screen identifies ITIH5 as a metastasis suppressor in pancreatic cancer

Abstract

The overwhelming majority of pancreatic ductal adenocarcinoma (PDAC) is not diagnosed until the cancer has metastasized, leading to an abysmal average life expectancy (3-6 months post-diagnosis). Earlier detection and more effective treatments have been hampered by inadequate understanding of the underlying molecular mechanisms controlling metastasis. We hypothesized that metastasis suppressors are involved in controlling metastasis in pancreatic cancer. Using an unbiased genome-wide shRNA screen, an shRNA library was transduced into the non-metastatic PDAC line S2-028 followed by intrasplenic injection. Resulting liver metastases were individually isolated from these mice. One liver metastatic nodule contained shRNA for ITIH5 (Inter-alpha-trypsin inhibitor heavy chain 5), suggesting that ITIH5 may act as a metastasis suppressor. Consistent with this notion, metastatic PDAC cell lines had significantly lower protein expression of ITIH5 compared to immortalized pancreatic ductal epithelial cells and non-/poorly-metastatic PDAC cell lines. By manipulating expression of ITIH5 in different PDAC cell lines (over-expression in metastatic, knockdown in non-metastatic) functional and selective regulation of metastasis was observed for ITIH5. Orthotopic tumor growth of PDAC cells was not blocked following orthotopic injection. In vitro ITIH5 over-expression inhibited motility and invasion. Immunohistochemical analysis of a human PDAC tissue microarray revealed that ITIH5 expression inversely correlated with both survival and invasion/metastasis. ITIH5 is, therefore, functionally validated as a PDAC metastasis suppressor and shows promise as a prognostic biomarker.

ITIH5 induces a shift in TGF‐β superfamily signaling involving Endoglin and reduces risk for breast cancer metastasis and tumor death

“…Recently, the working group of Danny R. Welch who had characterized the metastasis suppressor gene KISS1 in breast cancer, proposed ITIH5 as novel metastasis suppressor in pancreatic ductal carcinoma. By performing a genome‐wide shRNA library screen consisting of 74 468 shRNA directed against 21 416 human genes, ITIH5 was identified as one of two genes whose knockdown enabled the non‐metastatic PDAC cell line S2‐028 to metastasize to the liver using an intrasplenic injection mouse model …”

“…By performing a genome-wide shRNA library screen consisting of 74 468 shRNA directed against 21 416 human genes, ITIH5 was identified as one of two genes whose knockdown enabled the nonmetastatic PDAC cell line S2-028 to metastasize to the liver using an intrasplenic injection mouse model. 21,56 In the current study we aimed at specifying the role of ITIH5 in 33 in vitro. In addition, genes known to promote metastasis like S100A4 48 were inversely regulated with ITIH5.…”

“…In aggressive MDA‐MB‐231 breast cancer cells ITIH5 expression induces epigenetic reprogramming and abrogates mesenchymal migration, a hallmark of metastatic tumor cells, involving the tumor suppressor gene “death associated protein kinase 1” ( DAPK1 ) . In addition, ITIH5 was recently identified as a metastasis suppressor gene in in pancreatic ductal carcinoma …”

“…20 In addition, ITIH5 was recently identified as a metastasis suppressor gene in in pancreatic ductal carcinoma. 21 Apart from that the role of ITIH5 in metastatic processes, its downstream targets and signaling pathways are still unknown.…”

Uncovering the signaling landscape controlling breast cancer cell migration identifies splicing factor PRPF4B as a metastasis driver

“…Independent functional genomics screens have assessed the role of kinases and adhesion-related molecules in cell migration of lung carcinoma H1299 cells (24) and breast cancer MCF10A cells (25). Also recent in vivo RNAi screens have identified several novel modulators of cancer metastasis directly (26)(27)(28). However, a systematic analysis on the role of the complete spectrum of individual signaling molecules in tumor cell migration is lacking.…”