title: Silencing KRAS Overexpression in Cadmium-Transformed Prostate Epithelial Cells Mitigates Malignant Phenotype.
abstract: Cadmium (Cd) is a potential human prostate carcinogen. Chronic Cd exposure malignantly transforms RWPE-1 human prostate epithelial cells into CTPE cells by an unclear mechanism. Previous studies show that RWPE-1 can also be malignantly transformed by arsenic, and KRAS activation is key to causation and maintenance of this phenotype. Although Cd and arsenic can both transform prostate epithelial cells, it is uncertain whether their mechanisms are similar. Thus, here we determined whether KRAS activation is critical in causing and maintaining Cd-induced malignant transformation in CTPE cells. Expression of KRAS, miRNAs, and other genes of interest was analyzed by Western blot and RT-PCR. Following stable KRAS knockdown (KD) by RNA interference using shRNAmir, the malignant phenotype was assessed by various physical and genetic parameters. CTPE cells greatly overexpressed KRAS by 20-fold, indicating a likely role in Cd transformation. Thus, we attempted to reverse the malignant phenotype via KRAS KD. Two weeks after shRNAmir transduction, KRAS protein was undetectable in CTPE KD cells, confirming stable KD. KRAS KD reduced stimulated RAS/ERK and PI3K/AKT signaling pathways and markedly mitigated multiple physical and molecular malignant cell characteristics including: hypersecretion of MMP-2, colony formation, cell survival, and expression of cancer-relevant genes (reduced proliferation and cell cycle-related genes; activated tumor suppressor PTEN). However, KRAS KD did not reverse miRNA expression originally down-regulated by Cd transformation. These data strongly suggest KRAS is a key gene in development and maintenance of the Cd-induced malignant phenotype, at least in the prostate. It is not, however, the only genetic factor sustaining this phenotype.
authors: Stem Cell Toxicology Group, National Toxicology Program Laboratory, Division of the National Toxicology Program, National Institute of Environmental Health Sciences , Research Triangle Park, North Carolina 27709, United States.
authors: Ntube N O
authors: NN
authors: Ngalame
authors: Stem Cell Toxicology Group, National Toxicology Program Laboratory, Division of the National Toxicology Program, National Institute of Environmental Health Sciences , Research Triangle Park, North Carolina 27709, United States.
authors: Michael P
authors: MP
authors: Waalkes
authors: Stem Cell Toxicology Group, National Toxicology Program Laboratory, Division of the National Toxicology Program, National Institute of Environmental Health Sciences , Research Triangle Park, North Carolina 27709, United States.
authors: Erik J
authors: EJ
authors: Tokar
mesh_terms: D002104:Cadmium / Q000737:chemistry / Q000633:toxicity; D002460:Cell Line; D049109:Cell Proliferation / Q000235:genetics; D002471:Cell Transformation, Neoplastic / Q000187:drug effects / Q000235:genetics*; D004847:Epithelial Cells / Q000473:pathology / Q000502:physiology*; D015870:Gene Expression / Q000235:genetics; D055785:Gene Knockdown Techniques; D020868:Gene Silencing; D006801:Humans; D008297:Male; D011467:Prostate / Q000503:physiopathology*; D011471:Prostatic Neoplasms / Q000503:physiopathology*; D016283:Proto-Oncogene Proteins p21(ras) / Q000235:genetics* / Q000378:metabolism*; D015398:Signal Transduction / Q000235:genetics
chemical_list: C117307:KRAS protein, human; D002104:Cadmium; D016283:Proto-Oncogene Proteins p21(ras)