HIPK3 Inhibition by Exosomal hsa-miR-101-3p Is Related to Metabolic Reprogramming in Colorectal Cancer

BackgroundExosomes are extracellular vesicles secreted by most cells to deliver functional cargoes to recipient cells. MicroRNAs (miRNAs) constitute a significant part of exosomal contents. The ease of diffusion of exosomes renders them speedy and highly efficient vehicles to deliver functional molecules. Cancer cells secrete more exosomes than normal cells. Reports have showed that exosomal miRNAs of cancer cells facilitate cancer progression. Yet the complexity of cancer dictates that many more functional exosomal miRNAs remain to be discovered.MethodsIn this study, we analyzed miRNA expression profiles of tissue and plasma exosome samples collected from 10 colorectal cancer (CRC) patients and 10 healthy individuals. We focused on hsa-miR-101-3p (101-3p), a profoundly up-regulated miRNA enriched in plasma exosomes of patients bearing CRC. We performed target analysis of 101-3p and pursued functional studies of this microRNA in two colorectal cancer cell lines, namely HCT116 and SW480.ResultsOur results indicated that inhibiting 101-3p slowed cell growth and retarded cell migration in vivo in two colorectal cancer cell lines. Target analysis showed that Homeodomain-interacting protein kinase (HIPK3) is a target of miR-101-3p. HCT116 and SW480 cells stably overexpressing HIPK3 showed increased level of phosphorylated FADD, as well as retarded cell growth, migration, and increased sensitivity to 5-FU. In-depth analysis revealed increased mitochondrial membrane potential upon HIPK3 overexpression along with increased production of reactive oxygen species, number of mitochondria, and expression of respiratory complexes. Measurements of glycolytic parameters and enzymes revealed decreased level of glycolysis upon HIPK3 overexpression in these two cell lines. Xenograft model further confirmed a profoundly improved potency of the synergistic treatment combining both 5-FU and 101-3p inhibitor compared to 5-FU alone.ConclusionThis study unraveled an oncogenic nature of the exosomal 101-3p and suggested a relationship between the 101-3p-HIPK3 axis and metabolic homeostasis in colorectal cancer. Expression level of 101-3p is positively correlated with glycolytic capacity in CRC and therefore 101-3p itself is an oncomiR. Combining 101-3p inhibitor with chemotherapeutic agents is an effective strategy against CRC.

Secondary Metabolites from Marine-Derived Fungi and Actinobacteria as Potential Sources of Novel Colorectal Cancer Drugs

Colorectal cancer is one of the most common cancers diagnosed in the world. Chemotheraphy is one of the most common methods used for the pharmacological treatment of this cancer patients. Nevertheless, the adverse effect of chemotherapy is not optimized for improving the quality of life of people who are older, who are the most vulnerable subpopulation. This review presents recent updates regarding secondary metabolites derived from marine fungi and actinobacteria as novel alternatives for cytotoxic agents against colorectal cancer cell lines HCT116, HT29, HCT15, RKO, Caco-2, and SW480. The observed marine-derived fungi were from the species Aspergillus sp., Penicillium sp., Neosartorya sp., Dichotomomyces sp., Paradendryphiella sp., and Westerdykella sp. Additionally, Streptomyces sp. and Nocardiopsis sp. are actinobacteria discussed in this study. Seventy one compounds reviewed in this study were grouped on the basis of their chemical structures. Indole alkaloids and diketopiperazines made up most compounds with higher potencies when compared with other groups. The potency of indole alkaloids and diketopiperazines was most probably due to halogen-based functional groups and sulfide groups, respectively.

Mutant KRAS-associated proteome is mainly controlled by exogenous factors

KRAS signaling has been extensively studied, yet the clarification between KRAS-autonomous and non-autonomous mechanisms are still less explored. Understanding how KRAS signaling and effects are affected by exogenous stimuli can provide valuable insights not only to understand resistance mechanisms that justify pathway inhibition failure, but also to uncover novel therapeutic targets for mutant KRAS patients. Hence, aiming at understanding KRAS-autonomous versus non autonomous mechanisms, we studied the response of two mutant KRAS colorectal cancer cell lines (HCT116 and LS174T) - control and KRAS silenced- to TGFβ1-activated fibroblasts secretome. By performing a total proteome analysis, we observed that TGFβ1-activated fibroblast-secreted factors triggered cell line-specific proteome alterations and that mutant KRAS governs approximately 1/3 of those alterations. Moreover, the analysis of the impact of exogenous factors on the modulation of KRAS proteome revealed that, in both cell lines, more than 2/3 of the KRAS-associated proteome is controlled in a KRAS-non-autonomous manner and dependent on the exogenous factors. This work highlights the context-dependency of KRAS-associated signaling and reinforces the importance of establishing more integrative models resembling the complexity of the tumor microenvironment to study KRAS-associated signals.

Antiproliferative and Apoptotic Effects of Red Beetroot and Betanin on Human Colorectal Cancer Cell Lines

Background: Colorectal cancer (CRC) is the third most common type of cancer worldwide. Fruit and vegetables have some active compounds such as flavonoids and polyphenols that protect against malignancies through their antioxidative, anti-inflammatory, anti-proliferative, neuro, and hepatoprotective properties. Red beetroot (Beta vulgaris) contains red (betacyanins) and yellow (betaxanthins) pigments known as betalains. Betanin makes up 75-95% of the total betacyanins, possessed a wide range of favorable biological effects such as chemopreventive, anticarcinogenic, anti-tumorogenic, antiangiogenic, and proapoptotic effects. Methods: Red beetroot hydro-alcoholic extract and betanin were used to treat Caco-2 and HT-29 colorectal cancer cells, as well as KDR/293 normal epithelial cells. The half-maximal inhibitory concentration (IC50) was determined by prescreening MTT tests in the range of 20 to 140 µg/ml at 24 and 48 h. The cytotoxicity and apoptosis-inducing evaluations were performed via MTT assay, DAPI staining, and FACS-flow cytometry tests using determined times and doses. Moreover, the expression level of six important genes involving in the apoptosis pathway (Bcl-2, BAD, Caspase-3, Caspase-8, Caspase-9, and Fas-R) were determined using the real-time polymerase chain reaction (RT-PCR) method.Results: The IC50 doses for HT-29 and Caco-2 cell lines were determined to be about 92 μg/mL, 107 μg/mL for beetroot hydro-alcoholic extract, and 64 μg/mL, 90 μg/mL for betanin at 48 h, respectively. Our findings showed that beetroot extract and betanin significantly inhibit the growth of HT-29 and Caco-2 cell lines, time and dose-dependently, without considerable adverse effects on KDR/293 normal cells. Moreover, DAPI staining and flow cytometry results revealed significant apoptosis symptoms in treated cancerous cell lines. The expression level of pro-apoptotic genes involved in intrinsic and extrinsic apoptosis pathways (BAD, Caspase-3, Caspase-8, Caspase-9, and Fas-R) in treated HT-29 and Caco-2 cells was higher than untreated and normal cells, whereas the anti-apoptotic gene (Bcl-2) was downregulated. Conclusion: Beetroot hydro-alcoholic extract and betanin significantly inhibited cell proliferation and induced cell apoptosis (intrinsic and extrinsic pathways) via modification of effective genes in both colorectal cancer cell lines with no significant cytotoxic effects on KDR/293 normal cells. The mechanism of the anticancer effects of red beetroot extract and betanin needs to be further studied.