The characteristic signature of cancer stem cells (CSCs) is their association with enriched populations of cells that express the CD44 protein (CD44+). CD44 is a cell-adhesion molecule associated with branching during normal prostate gland morphogenesis and, when not expressed, leads to reduced tumor growth and metastasis. CDCs have been shown to play a key role in the growth and metastasis of many cancers, including breast, pancreatic, head and neck, colon, small intestine, liver, stomach, bladder and ovarian cancers. They also appear to be resistant to conventional anti-cancer therapies and the reason why many cancers recur. Their functions in many cancers are known to be regulated by microRNAs (miRNAs). However, the ability of miRNAs to inhibit prostate CSC function was unknown until a research group led by Professor Dean Tang, Department of Molecular Carcinogenesis, the University of Texas, Houston, reported that a specific miRNA, miR-34a, inhibits prostate CSC functions (Liu et al. 2011). These findings indicated that miR-34a could be a new weapon in the fight against prostate cancer.
miRNAs are short RNAs (averaging only 22 nucleotides long) found in all cells. They are post-transcriptional regulators that "silence" genes by binding to complementary sequences on mRNAs. The human genome may encode more than 1,000 miRNAs, which may target about 60% of mammalian genes. They are abundant in many human cell types. In 2011Professor Tang's team found that miR-34a, identified from a pool of 324 sequence-validated human miRNAs, is expressed significantly less in all CD44+ prostate cancer cell populations than in corresponding CD44− cells from the same tumors—in both prostate cancer cell lines and primary human prostate tumors. Additionally, they found that miR-34a levels in human prostate cancers and cell lines positively correlate with the functional status of p53, a known transcriptional regulator of miR-34a.
miR-34a protects mice from cancer
To determine if miR-34a protects mice from prostate cancer, Tang's team implanted NOD.CB17-Prkdcscid/J (NOD scid, 001303) mice either subcutaneously or orthotopically with prostate cancer cell lines transfected with either miR-34a or vectors encoding pre-miR-34a. They found that these mice develop significantly smaller tumors than controls.
miR-34a acts on prostate CSCs
To determine if miR-35a inhibits prostate cancer growth and metastasis by targeting prostate CSCs, Tang's team implanted NOD scid mice with purified CSCs from prostate cancer cell lines transfected with vectors encoding either miR-34a, pre-miR-34a, or miR34-a inhibitors. They found that mice implanted with CSCs transfected with miR34-a inhibitors develop significantly more tumors and lung metastases. Additionally, miR-34a inhibits the clonogenic, holoclone-forming, and sphere-forming capacity of prostate CD44+ CSCs, and miR34-a inhibitors increase the inherently low sphere-forming capacity of CD44− CSCs. Taken together, these results indicate that miR-34a inhibits prostate CSC properties.
miR-34a thwarts growth of established tumors
Tang and his team demonstrated that miR-34a thwarts the growth and metastasis of established prostate cancers in NOD scid mice: repeated injections of miR-34a into subcutaneously implanted prostate tumors halts their growth; tail vein injections of liposome encapsulated miR-34a reduces tumor burden by 50%; and systemic delivery of miR-34a into orthotopically implanted tumors reduces lung metastasis and extends mouse survival.
CD44/miR-34a binding sites identified
Tang and his team wanted to know how miR-34a suppresses CD44 expression levels. By implementing a target-prediction program and performing experiments involving miR-34a oligos, a luciferase reporter and several prostate cancer lines, they found strong evidence that miR-34a, surprisingly, regulates CD44 expression directly through two binding sites in the 3′ UTR of the gene that encodes CD44.
miR-34a directly targets CD44
Tang and his team produced several lines of evidence that miR-34a directly targets CD44: inactivating CD44 in several prostate cancer lines inhibits orthotopic and/or subcutaneous tumorigenesis and/or lung metastasis in NOD scid mice. Although CD44+ prostate cancer cells have higher migratory and invasive capacities than CD44− cells, these capacities are partially inhibited by miR-34a. CD44 cDNA that lacks the 3′ UTR miR-34a binding site thwarts miR-34a's ability to inhibit the migratory and invasive capacities of cultured prostate cancer cells, though over-expression of CD44 does not significantly reduce miR-34a's ability to inhibit the proliferation of these cells.
In summary, Tang and his team showed for the first time that miR-34a, a microRNA regulated by p53, significantly inhibits prostate cancer growth and metastasis by suppressing the migratory, invasive and regenerative properties of CD44+ prostate CSCs. This previously unknown miR-34a signaling pathway reveals promising targets for fighting prostate and other cancers, such as those in the lung and pancreas.
References
Liu C, Kelnar K, Liu B, Chen X, Calhoun-Davis T, Li H, Patrawala L, Yan H, Jeter C, Honorio S, Wiggins JF, Bader AG, Fagin R, Brown D, Tang DG. 2011. The microRNA miR-34a inhibits prostate cancer stem cells and metastasis by directly repressing CD44. Nat Med Jan16.