Prognostic and diagnostic value of metal metabolism in colorectal cancer

Colorectal cancer (CRC) ranks as the second leading cause of cancer-related mortality globally and is the third most prevalent cancer. The effective treatment of CRC is challenged by tumor drug resistance, underscoring the urgent need for novel therapeutic strategies. It is well recognized that epigenetic modifications, particularly DNA methylation, significantly contribute to the initiation and advancement of CRC. Tumors frequently exhibit hypermethylation of specific gene promoters alongside a general hypomethylation of genomic DNA. In normal tissues, gene promoters, particularly those of tumor suppressor genes, are typically unmethylated, whereas they are often hypermethylated in cancerous tissues. Previous studies have identified the methylation status of the CpG island within the SLC30A10 gene as a reliable biomarker for CRC. Notably, a hypermethylated phenotype is inversely correlated with SLC30A10 expression. However, the precise functional implications of SLC30A10 suppression in the context of CRC progression remain to be elucidated.). The investigation of molecular pathways involved in metal metabolism in CRC has not been conducted previously at the transcriptomic level. Furthermore, the relationship between the mutational characteristics of tumors—such as individual mutations, mutational burden, and microsatellite instability—and the activity of genes related to metal metabolic pathways has not been explored. This project aims to be the first to examine the antitumor potential of modulating the transport systems for manganese and zinc ions in CRC. The molecular function of SLC30A10 is to export zinc (Zn) and manganese (Mn) ions from the cell. Additionally, SLC30A10 plays a crucial role in mitigating Mn-induced cytotoxicity and facilitates Zn transport to early endosomes while promoting endosomal recirculation to avert Zn toxicity. Our findings indicate that overexpression of SLC30A10 is correlated with the knockout of its functional antagonist, SLC39A14 in the HuTu80 and HCT-15 cell lines. This study aimed to examine the involvement of the SLC39A8, SLC39A14, SLC30A3, and SLC30A10 genes in the progression of colorectal cancer (CRC). To achieve this, gene editing was performed in two cell lines: HCT-15, which harbors a KRAS mutation, and HuTu80. Biallelic mutations of the SLC39A8, SLC39A14, SLC30A3, and SLC30A10 genes were introduced using CRISPR/Cas9 technology in both cell lines. The genetic constructs were cloned into the pSpCas9(BB)-2A-GFP (PX458) plasmid (Addgene). Overexpression experiments in both cell lines were conducted using the pCDH-EF1-MCS-IRES-copGFP plasmid (SBI, USA), where GFP was replaced with a puromycin resistance selection marker. ORF clones for SLC39A8 (NM_022154.5), SLC30A3 (NM_003459.5), SLC30A10 (NM_018713.3), and SLC39A14 (NM_015359.6) were obtained from Origene for the construction of lentiviral vectors.