title: A genome-wide RNAi screen identifies FOXO4 as a metastasis-suppressor through counteracting PI3K/AKT signal pathway in prostate cancer.
abstract: Activation of the PI3K/AKT signal pathway is a known driving force for the progression to castration-recurrent prostate cancer (CR-CaP), which constitutes the major lethal phenotype of CaP. Here, we identify using a genomic shRNA screen the PI3K/AKT-inactivating downstream target, FOXO4, as a potential CaP metastasis suppressor. FOXO4 protein levels inversely correlate with the invasive potential of a panel of human CaP cell lines, with decreased mRNA levels correlating with increased incidence of clinical metastasis. Knockdown (KD) of FOXO4 in human LNCaP cells causes increased invasion in vitro and lymph node (LN) metastasis in vivo without affecting indices of proliferation or apoptosis. Increased Matrigel invasiveness was found by KD of FOXO1 but not FOXO3. Comparison of differentially expressed genes affected by FOXO4-KD in LNCaP cells in culture, in primary tumors and in LN metastases identified a panel of upregulated genes, including PIP, CAMK2N1, PLA2G16 and PGC, which, if knocked down by siRNA, could decrease the increased invasiveness associated with FOXO4 deficiency. Although only some of these genes encode FOXO promoter binding sites, they are all RUNX2-inducible, and RUNX2 binding to the PIP promoter is increased in FOXO4-KD cells. Indeed, the forced expression of FOXO4 reversed the increased invasiveness of LNCaP/shFOXO4 cells; the forced expression of FOXO4 did not alter RUNX2 protein levels, yet it decreased RUNX2 binding to the PIP promoter, resulting in PIP downregulation. Finally, there was a correlation between FOXO4, but not FOXO1 or FOXO3, downregulation and decreased metastasis-free survival in human CaP patients. Our data strongly suggest that increased PI3K/AKT-mediated metastatic invasiveness in CaP is associated with FOXO4 loss, and that mechanisms to induce FOXO4 re-expression might suppress CaP metastatic aggressiveness.
authors.affiliation: Biomedical Research Institute, Shenzhen-PKU-HKUST Medical Center, Shenzhen, Guangdong, China; Department of Cancer Genetics, Roswell Park Cancer Institute, Buffalo, New York, United States of America.
authors.forename: Bing
authors.initials: B
authors.lastname: Su
authors.affiliation: Department of Cancer Genetics, Roswell Park Cancer Institute, Buffalo, New York, United States of America.
authors.forename: Lingqiu
authors.initials: L
authors.lastname: Gao
authors.affiliation: Department of Cancer Genetics, Roswell Park Cancer Institute, Buffalo, New York, United States of America.
authors.forename: Catherine
authors.initials: C
authors.lastname: Baranowski
authors.affiliation: Department of Pharmacology and Therapeutics, Roswell Park Cancer Institute, Buffalo, New York, United States of America.
authors.forename: Bryan
authors.initials: B
authors.lastname: Gillard
authors.affiliation: Department of Biostatistics and Bioinformatics, Roswell Park Cancer Institute, Buffalo, New York, United States of America.
authors.forename: Jianmin
authors.initials: J
authors.lastname: Wang
authors.affiliation: Department of Cancer Genetics, Roswell Park Cancer Institute, Buffalo, New York, United States of America.
authors.forename: Ryan
authors.initials: R
authors.lastname: Ransom
authors.affiliation: Department of Pharmacology, Yale University School of Medicine, New Haven, Connecticut, United States of America.
authors.forename: Hyun-Kyung
authors.initials: HK
authors.lastname: Ko
authors.affiliation: Department of Cancer Genetics, Roswell Park Cancer Institute, Buffalo, New York, United States of America.
authors.forename: Irwin H
authors.initials: IH
authors.lastname: Gelman
mesh_terms: D000818:Animals; D018797:Cell Cycle Proteins; D045744:Cell Line, Tumor; D051858:Forkhead Transcription Factors; D015972:Gene Expression Regulation, Neoplastic; D055106:Genome-Wide Association Study; D064593:Heterografts; D006801:Humans; D008297:Male; D051379:Mice; D008819:Mice, Nude; D009362:Neoplasm Metastasis; D009368:Neoplasm Transplantation; D019869:Phosphatidylinositol 3-Kinases / Q000235:genetics / Q000378:metabolism; D011471:Prostatic Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D051057:Proto-Oncogene Proteins c-akt; D034622:RNA Interference; D020218:Response Elements; D015398:Signal Transduction / Q000235:genetics*; D014157:Transcription Factors / Q000096:biosynthesis / Q000235:genetics; D025521:Tumor Suppressor Proteins / Q000235:genetics / Q000378:metabolism
chemical_list: D018797:Cell Cycle Proteins; D051858:Forkhead Transcription Factors; D019869:Phosphatidylinositol 3-Kinases; D051057:Proto-Oncogene Proteins c-akt; D014157:Transcription Factors; D025521:Tumor Suppressor Proteins; C108803:FOXO4 protein, human