Negative Regulation of ULK1 by microRNA-106a in Autophagy Induced by a Triple Drug Combination in Colorectal Cancer Cells In Vitro

Abstract

Colorectal cancer (CRC) is among the top three most deadly cancers worldwide. The survival rate for this disease has not been reduced despite the treatments, the reason why the search for therapeutic alternatives continues to be a priority issue in oncology. In this research work, we tested our successful pharmacological combination of three drugs, metformin, doxorubicin, and sodium oxamate (triple therapy, or TT), as an autophagy inducer. Firstly, we employed western blot (WB) assays, where we observed that after 8 h of stimulation with TT, the proteins Unc-51 like autophagy activating kinase 1(ULK1), becline-1, autophagy related 1 protein (Atg4), and LC3 increased in the CRC cell lines HCT116 and SW480 in contrast to monotherapy with doxorubicin. The overexpression of these proteins indicated the beginning of autophagy flow through the activation of ULK1 and the hyperlipidation of LC3 at the beginning of this process. Moreover, we confirm that ULK1 is a bona fide target of hsa-miR-106a-5p (referred to from here on as miR-106a) in HCT116. We also observed through the GFP-LC3 fusion protein that in the presence of miR-106a, the accumulation of autophagy vesicles in cells stimulated with TT is inhibited. These results show that the TT triggered autophagy to modulate miR-106a/ULK1 expression, probably affecting different cellular pathways involved in cellular proliferation, survivance, metabolic maintenance, and cell death. Therefore, considering the importance of autophagy in cancer biology, the study of miRNAs that regulate autophagy in cancer will allow a better understanding of malignant tumors and lead to the development of new disease markers and therapeutic strategies.

Controlling the confounding effect of metabolic gene expression to identify actual metabolite targets in microsatellite instability cancers

“…Many metabolic drugs have been investigated as metabolic therapeutic targets in MSI cancers, such as glycolysis inhibitors (oxamate, lonidamine (LND)) [40,41], glutamate inhibitors (6-diazo-5-oxo-Lnorleucine, CB839) [42,43], and lipid metabolism inhibitors (cerulenin, TVB-3664, TOFA) [37]. In our study, dysregulated metabolic pathways resulted in aberrant glycolysis and nucleotide, glutamate, and lipid pathways in MSI cancer cell lines [44].…”

Experimental validation of in silico analysis estimated the reverse effect of upregulated hsa‐miR‐106a‐5p and hsa‐miR‐223‐3p on SLC4A4 gene expression in Iranian patients with colorectal adenocarcinoma by RT‐qPCR

“…Gene Ontology and signaling pathway analysis indicate that the miR‐106 family plays a significant role in the onset and progression of CRC. Therefore, recent publications suggest the prognostic role of hsa‐miR‐106a in CRC progression 40,41 . Furthermore, we could propose that the hsa‐miR‐106a/ SLC4A4 regulatory network plays a role in CRC pathogenicity.…”

“…Therefore, recent publications suggest the prognostic role of hsa‐miR‐106a in CRC progression. 40 , 41 Furthermore, we could propose that the hsa‐miR‐106a/ SLC4A4 regulatory network plays a role in CRC pathogenicity.…”

Autophagy and beyond: Unraveling the complexity of UNC-51-like kinase 1 (ULK1) from biological functions to therapeutic implications

“…Uridine at the 5′ end of miRNA, which can partially complement the 3′ untranslated region of mRNA, leading to in translation repression of the mRNA, thus, playing a vital role in post-transcriptional regulation of gene expression 60 . Numerous studies have unveiled that multiple mi-RNAs, including miR-372 61 , miR-93 62 , miR-142-5p 63 , miR-20a 64 , 65 , miR-106b 64 , miR-106a 66 , 67 , 68 , miR-26a/b 69 , miR-489 70 , miR-1262 71 , miR-214 72 , miR-214-3p 46 , miR-125b 72 , and miR-155 73 inhibit ULK1 gene expression by directly targeting the mRNA of ULK1 74 , 75 , 76 , 77 , 78 , 79 , 80 , 81 , 82 , 83 , 84 , 85 , 86 , 87 , 88 . In addition, microRNAs could also play an indirect role by regulating ULK1-related pathways.…”