Differential expression of MCM2 and SKP2 as biomarkers of therapy resistance in prostate cancer

Background Prostate cancer (PCa) is the second most incident cancer and the sixth cause of cancer death among man worldwide. Patients diagnosed with advanced PCa are often treated with androgen deprivation therapy. Although tumors initially respond, they may progress to a lethal and drug-resistant form of PCa. Recent works from our team demonstrated that a promising anti-cancer agent is associated with reduced MCM2 and SKP2 expression in PCa cell lines. MCM2 and SKP2 play an important role in cell cycle progression with overexpression observed in several cancers with association with poor prognostic. Thus, to test its potential as predictive biomarker, MCM2 and SKP2 expression was evaluated in PCa tissues. Methods A total of 57 cases (25 resistant and 33 non-resistant) were used to explore MCM2 and SKP2 expression by immunohistochemistry. Percentage of positive tumor cells, intensity of immunostaining, and immunoexpression were blinded evaluated. Comparisons between therapy conditions and proteins expression were assessed by Chi square test (statistical significance considered when P < 0.05). Results MCM2 expression is associated to PCa patients’ therapy resistance, with significant differences found for percentage of positive tumor cells (P = 0.001) and immunostaining intensity (P = 0.002). SKP2 expression is also associated with resistance to therapy, with differences observed referred to immunostaining intensity (P = 0.001) and immunoexpression pattern (P = 0.02). Specifically, cytoplasmatic and nuclear expression was apparent in most cases that developed therapy-resistance, whereas responsive PCa patients showed only SKP2 cytoplasmatic expression. Conclusions In PCa patients, MCM2 and SKP2 expression is significantly associated with therapy resistance. SKP2 cellular localization is also indicative of patients’ resistance to treatment.

Mint3 depletion restricts tumor malignancy of pancreatic cancer cells by decreasing SKP2 expression via HIF-1

Pancreatic cancer is one of the most fatal cancers without druggable molecular targets. Hypoxia inducible factor-1 (HIF-1) is a heterodimeric transcriptional factor that promotes malignancy in various cancers including pancreatic cancer. Herein, we found that HIF-1 is accumulated in normoxic or moderate hypoxic areas of pancreatic cancer xenografts in vivo and is active even during normoxia in pancreatic cancer cells in vitro. This prompted us to analyze whether the HIF-1 activator Mint3 contributes to malignant features of pancreatic cancer. Mint3 depletion by shRNAs attenuated HIF-1 activity during normoxia and cell proliferation concomitantly with accumulated p21 and p27 protein in pancreatic cancer cells. Further analyses revealed that Mint3 increased transcription of the oncogenic ubiquitin ligase SKP2 in pancreatic cancer cells via HIF-1. This Mint3-HIF-1-SKP2 axis also promoted partial epithelial-mesenchymal transition, stemness features, and chemoresistance in pancreatic cancer cells. Even in vivo, Mint3 depletion attenuated tumor growth of orthotopically inoculated human pancreatic cancer AsPC-1 cells. Database and tissue microarray analyses showed that Mint3 expression is correlated with SKP2 expression in human pancreatic cancer specimens and high Mint3 expression is correlated with poor prognosis of pancreatic cancer patients. Thus, targeting Mint3 may be useful for attenuating the malignant features of pancreatic cancer.

Fbxw7 and Skp2 Regulate Stem Cell Switch between Quiescence and Mitotic Division in Lung Adenocarcinoma

Background/Aims. The molecular mechanism of dormancy initiation of cancer stem cells (CSCs) is not clear. This study was to explore the molecular mechanism by which CSCs switch from mitotic division to quiescence. Methods. MTT assays, flow cytometry, Western blotting, qRT-PCR, and immunofluorescence staining were used to test cell viability, cell cycle and expression of F-box and WD repeat domain-containing 7 (Fbxw7), c-myc, S phase kinase associated protein-2 (Skp2), cyclin-dependent kinase inhibitor 1B (p27), octamer-binding transcription factor 3/4 (Oct3/4), and β catenin gene in 5-fluorouracil (5-FU)-treated A549 cells. Lung adenocarcinoma xenograft models were employed to detect the effects of Fbxw7 on tumor growth. Results. 5-FU inhibited the proliferation of A549 cells, with a median inhibitory concentration (IC50) of 200 μg/ml after 24 h treatment. 5-FU treatment increased the expressions of Oct3/4, Fbxw7, and p27 and increased the number of A549 cells at G0/G1. 5-FU treatment triggered nuclear translocation of β-catenin, decreased the expression levels of c-myc and Skp2, and decreased the number of A549 cells at S phase. Release from 5-FU decreased the expressions of Oct3/4, Fbxw7 and p27; decreased the percentage of cells in the G0/G1 phase; increased the expressions of Skp2 and c-myc; and increased the proportion of cells in S phase. 5-FU treatment led to high expressions of Oct3/4, c-myc, and p27, with low expressions of Fbxw7 and Skp2. Knockdown of Fbxw7 augmented the expression of c-myc and decreased the proportion of A549 cells in Go/G1 phase. Skp2 siRNA increased the expression of p27 and the percentage of G0/G1 phase cells and reduced the proportion of S phase cells. Fbxw7 overexpression inhibited tumor growth in mouse lung adenocarcinoma xenograft models. When Fbxw7 expression was low, Skp2 expression was higher in lung adenocarcinoma tissues and associated with the differentiation of lung adenocarcinoma. Conclusion. 5-FU enriches the CSCs in lung adenocarcinoma cells via increasing Fbxw7 and decreasing Skp2 expression, followed by downregulation of c-myc and upregulation of p27, which switches cells to quiescence.