Caveolin-1 (CAV1) acts as a growth suppressor in various human malignancies, but its expression is elevated in many advanced cancers, suggesting the oncogenic switch of its role during tumor progression. To understand the molecular basis for the growth-promoting function of CAV1, we characterized its expression status, differential roles for tumor growth, and effect on glucose metabolism in colorectal cancers. Abnormal elevation of CAV1 was detected in a substantial fraction of primary tumors and cell lines and tightly correlated with promoter CpG sites hypomethylation. Depletion of elevated CAV1 led to AMPK activation followed by a p53-dependent G₁ cell-cycle arrest and autophagy, suggesting that elevated CAV1 may contribute to ATP generation. Furthermore, CAV1 depletion downregulated glucose uptake, lactate accumulation, and intracellular ATP level, supporting that aerobic glycolysis is enhanced by CAV1. Consistently, CAV1 was shown to stimulate GLUT3 transcription via an HMGA1-binding site within the GLUT3 promoter. HMGA1 was found to interact with and activate the GLUT3 promoter and CAV1 increased the HMGA1 activity by enhancing its nuclear localization. Ectopic expression of HMGA1 increased glucose uptake, whereas its knockdown caused AMPK activation. In addition, GLUT3 expression was strongly induced by cotransfection of CAV1 and HMGA1, and its overexpression was observed predominantly in tumors harboring high levels of CAV1 and HMGA1. Together, these data show that elevated CAV1 upregulates glucose uptake and ATP production through HMGA1-mediated GLUT3 transcription, suggesting that CAV1 may render tumor cells growth advantages by enhancing aerobic glycolysis. Cancer Res; 72(16); 4097–109. ©2012 AACR.

Epigenetic inactivation of X-linked inhibitor of apoptosis (XIAP)-associated factor 1 (XAF1) is frequently observed in multiple human malignancies. However, the mechanisms underlying its tumor-suppression function remain largely undefined. Here, we identify XAF1 as a positive feedback regulator of p53, which directs the apoptotic switch of p53 signaling. As a unique transcription target of p53 in signaling apoptosis, XAF1 acts as a competitor of E3 ubiquitin ligase MDM2 in binding to p53 and thus disrupts the p53–MDM2 regulatory loop. Moreover, XAF1 appears to promote homeodomain-interacting protein kinase 2 (HIPK2)-mediated p53 phosphorylation by interrupting HIPK2-targeting function of E3 ubiquitin ligase Siah2 and promotes zinc finger protein 313 (ZNF313)-induced p21WAF1 ubiquitination. XAF1 thus represents one critical regulator of p53’s cell-fate decision function, suggesting that restoring the p53–XAF1 feedback loop could be an attractive avenue for the therapeutic intervention of malignant tumor progression.

Testing new ways to identify untapped opportunities for glioblastoma therapies remains highly significant. Amplification and overexpression of MDM2 gene is frequent in glioblastoma and disrupting the MDM2−p53 interaction is a promising strategy to treat the cancer. RG7112 is the first-in class inhibitor and recently discovered AMG232 is the most potent MDM2 inhibitor known to date. Here, we compared the effects of these two clinical MDM2 inhibitors in six glioblastoma cell lines and ten patient-derived glioblastoma stem cells. Targeted sequencing of the TP53, MDM2 genes and whole transcriptome analysis were conducted to verify genetic status associated with sensitivity and resistance to the drugs. Although TP53 wild-type glioblastoma cell lines are similarly sensitive to AMG232 and RG7112, we found that four TP53 wild-type out of ten patient-derived glioblastoma cells are much more sensitive to AMG232 than RG7112 (average IC₅₀ of 76 nM vs. 720 nM). Among these, 464T stem cells containing MDM2 gene amplification were most sensitive to AMG232 with IC₅₀ of 5.3 nM. Moreover, AMG232 exhibited higher selectivity against p53 wild-type cells over p53 mutant stem cells compared to RG7112 (average selectivity of 512-fold vs. 16.5-fold). Importantly, we also found that AMG232 is highly efficacious in three-dimensional (3D) tumor spheroids growth and effectively inhibits the stemness-related factors, Nestin and ZEB1. Our data provide new evidence that glioblastoma stem cells have high susceptibility to AMG232 suggesting the potential clinical implications of MDM2 inhibition for glioblastoma treatment. These will facilitate additional preclinical and clinical studies evaluating MDM2 inhibitors in glioblastoma and direct further efforts towards developing better MDM2-targeted therapeutics.

Radiotherapy represents the most effective non-surgical modality in cancer treatment. MicroRNAs (miRNAs) are small non-coding RNAs that regulate gene expression, and are involved in many biological processes and diseases. To identify miRNAs that influence the radiation response, we performed miRNA array analysis using MCF7 cells at 2, 8, and 24 h post irradiation. We demonstrated that miR-770-5p is a novel radiation-inducible miRNA. When miR-770-5p was overexpressed, relative cell number was reduced due to increased apoptosis in MCF7 and A549 cells. Transcriptomic and bioinformatic analyses revealed that PDZ-binding kinase (PBK) might be a possible target of miR-770-5p for regulation of radiosensitivity. PBK regulation mediated by direct targeting of miR-770-5p was demonstrated using luciferase reporter assays along with wild-type and mutant PBK-3′untranslated region constructs. Radiation sensitivity increased and decreased in miR-770-5p- and anti-miR-770-5p-transfected cells, respectively. Consistent with this result, transfection of short interfering RNA against PBK inhibited cell proliferation, while ectopic expression of PBK restored cell survival from miR-770-5p-induced cell death. In addition, miR-770-5p suppressed tumor growth, and miR-770-5p and PBK levels were inversely correlated in xenograft model mice. Altogether, these data demonstrated that miR-770-5p might be a useful therapeutic target miRNA that sensitizes tumors to radiation via negative regulation of PBK.