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Background

Targeting AKT suppresses tumor growth through inducing apoptosis, however, during which whether other forms of cell death occurring is poorly understood.

Methods

The effects of increasing PARP1 dependent cell death (parthanatos) induced by inhibiting AKT on cell proliferation were determined by CCK-8 assay, colony formation assay, Hoechst 33,258 staining and analysis of apoptotic cells by flow cytometry. For the detailed mechanisms during this process, Western blot analysis, qRT-PCR analysis, immunofluorescence and co-immunoprecipitation were performed. Moreover, the inhibition of tumor growth by inducing p53/SIRT6/PARP1-dependent parthanatos was further verified in the xenograft mouse model.

Results

For the first time, we identified that inhibiting AKT triggered parthanatos, a new form of regulated cell death, leading to colon cancer growth suppression. For the mechanism investigation, we found that after pharmacological or genetic AKT inhibition, p53 interacted with SIRT6 and PARP1 directly to activate it, and promoted the formation of PAR polymer. Subsequently, PAR polymer transported to outer membrane of mitochondria and resulted in AIF releasing and translocating to nucleus thus promoting cell death. While, blocking PARP1 activity significantly rescued colon cancer from death. Furthermore, p53 deletion or mutation eliminated PAR polymer formation, AIF translocation, and PARP1 dependent cell death, which was promoted by overexpression of SIRT6. Meanwhile, reactive oxygen species production was elevated after inhibition of AKT, which might also play a role in the occurrence of parthanatos. In addition, inhibiting AKT initiated protective autophagy simultaneously, which advanced tumor survival and growth.

Conclusion

Our findings demonstrated that AKT inhibition induced p53-SIRT6-PARP1 complex formation and the activation of parthanatos, which can be recognized as a novel potential therapeutic strategy for cancer. Video Abstract. Show less

Colorectal cancer (CRC) is the third most prevalent cancer type worldwide with a mortality rate of approximately 50%. Elevated cell-surface expression of truncated carbohydrate structures such as Tn antigen (GalNAcα-Ser/Thr) is frequently observed during tumor progression. We have previously demonstrated that the C-type lectin macrophage galactose-type lectin (MGL), expressed by human antigen presenting cells, can distinguish healthy tissue from CRC through its specific recognition of Tn antigen. Both MGL binding and oncogenic BRAF mutations have been implicated in establishing an immunosuppressive microenvironment. Here we aimed to evaluate whether MGL ligand expression has prognostic value and whether this was correlated to BRAF(V600E) mutation status. Using a cohort of 386 colon cancer patients we demonstrate that high MGL binding to stage III tumors is associated with poor disease-free survival, independent of microsatellite instability or adjuvant chemotherapy. In vitro studies using CRC cell lines showed an association between MGL ligand expression and the presence of BRAF(V600E). Administration of specific BRAF(V600E) inhibitors resulted in decreased expression of MGL-binding glycans. Moreover, a positive correlation between induction of BRAF(V600E) and MGL binding to epithelial cells of the gastrointestinal tract was found in vivo using an inducible BRAF(V600E) mouse model. We conclude that the BRAF(V600E) mutation induces MGL ligand expression, thereby providing a direct link between oncogenic transformation and aberrant expression of immunosuppressive glycans. The strong prognostic value of MGL ligands in stage III colon cancer patients, i.e. when tumor cells disseminate to lymph nodes, further supports the putative immune evasive role of MGL ligands in metastatic disease. Show less

Cell lines are invaluable biomedical research tools, and recent literature has emphasized the importance of genotype authentication and characterization. In the present study, 24 out of 27 cell line identities were confirmed by short tandem repeat profiling. The molecular phenotypes of the 24 colon cancer cell lines were examined, and microsatellite instability (MSI) and CpG island methylator phenotype (CIMP) were determined, using the Bethesda panel mononucleotide repeat loci and two epimarker panels, respectively. Furthermore, the BRAF, KRAS and PIK3CA oncogenes were analyzed for mutations in known hotspots, while the entire coding sequences of the PTEN and TP53 tumor suppressors were investigated. Nine cell lines showed MSI. Thirteen and nine cell lines were found to be CIMP positive, using the Issa panel and the Weisenberger et al. panel, respectively. The latter was found to be superior for CIMP classification of colon cancer cell lines. Seventeen cell lines harbored disrupting TP53 mutations. Altogether, 20/24 cell lines had the mitogen-activated protein kinase pathway activating mutually exclusive KRAS or BRAF mutations. PIK3CA and PTEN mutations leading to hyperactivation of the phosphoinositide 3-kinase/AKT pathway were observed in 13/24 cell lines. Interestingly, in four cell lines there were no mutations in neither BRAF, KRAS, PIK3CA nor in PTEN. In conclusion, this study presents molecular features of a large number of colon cancer cell lines to aid the selection of suitable in vitro models for descriptive and functional research. Show less

Key words: Mitochondrial uncoupling protein 2; Colon cancer; Uncoupling protein 2; Clinicopathologic characteristics AIM: To detect the expression of mitochondrial uncoupling protein 2 (UCP2) in colon cancer and analyze the relation between UCP2 expression and clinical pathological features of colon cancer. Peer reviewer: Guangcun Huang, MD, PhD, Research Institute at Nationwide Children’s Hospital, Center for Clinical and Translational Research, Columbus, OH 43205, United States METHODS: Fifteen colon tissue samples and 15 its adjacent tissue samples were obtained from colon cancer Show less