Predicting candidate biomarkers for COVID-19 associated with leukemia in children.

Am J Clin Exp Immunol 2024;13(6):246-258 www.ajcei.us /ISSN:2164-7712/AJCEI0162141 Original Article Predicting candidate biomarkers for COVID-19 associated with leukemia in children Judy Bai1, Qing Li2 Greenhills School, Ann Arbor, MI 48105, USA; 2Department of Internal Medicine, University of Michigan, Ann Arbor, MI 48109, USA 1 Received November 19, 2024; Accepted December 1, 2024; Epub December 25, 2024; Published December 30, 2024 Abstract: Since the COVID-19 pandemic, a significant number of pediatric leukemia patients have shown to have also contracted COVID-19 several weeks or months prior to the development of their cancer. Current research indicates the expression of MDA5, encoded by IFIH1, is associated with increased immunity to COVID-19 in children. Children are also known to have a much lower risk of developing leukemia. Our hypothesis is that IFIH1 and its regulatory miRNAs are biomarkers associated with pediatric leukemia; the objective of our study is to identify genes, through miRNA targeting mechanisms, which may be biomarkers associated with COVID-19 infection and leukemia. The database TarBase was analyzed to identify miRNAs that target IFIH1, followed by the identification of other genes regulated by IFIH1’s targeting miRNAs, to construct a gene-miRNA targeting network. Protein-Protein Interaction (PPI) analysis and DAVID/KEGG pathway analysis were conducted to identify genes with meaningful biological interactions and pathways. We identified two significant miRNAs, hsa-196a-5p and hsa-196b-5p, and 51 of their targeted and highly expressed genes reported in the Acute Myeloid Leukemia (AML) samples from The Cancer Genome Atlas (TCGA) RNA sequencing database. When conducting additional analysis using the Gene Constellation module of the Immunological Genome Project for the top three candidate genes, several other genes were identified to be highly correlated with STAT3 and IFIH1 in our study. Based on our investigation into co-expression analysis, we found that IFIH1 is a potential biomarker for AML. We are expanding our work to create a machine learning model to identify other biomarkers, examine the significance of various parameters (age, race, etc.), and perform comorbidity network analysis for other potential genes/miRNAs. Keywords: Leukemia, COVID-19, RNA-Seq, differential expression Introduction Acute Myeloid Leukemia (AML) has a higher mortality rate compared to Acute Lymphoblastic Leukemia (ALL) in children, with most cases occurring in children who are either younger than 2 years old or teenagers. In AML, myeloid stem cells differentiate into unhealthy white blood cells called “blasts”. These blasts, instead of fighting off infection and helping to protect the body, build up in bone marrow and leave less room for healthy blood cells. This type of leukemia also does not always stay in the blood or bone marrow and may travel to the skin, spine, or other organs. Many studies have consistently reported that many children with AML have experienced a previous episode of COVID infection weeks to months prior to the onset of their cancer, suggesting an association between COVID-19 infection and AML development. In one study, the researchers discovered that the correlation between leukemia and COVID-19 infection is the strongest when compared to other cancers [1]. This is consistent with results from another study, which showed that COVID-19 infection is associated with acute leukemia and severe bone marrow involvement [2]. COVID-19 has also been associated with other types of blood cancers. For example, the diagnosis of Hair Cell Leukemia (HCL) in COVID-19 patients has been a reported relationship [3]. In addition, children who have blood malignancy are more likely to contract significant COVID-19 infection. Furthermore, patients with COVID-19 have an increased risk of developing a life-threatening illness if they are receiving one or more treathttps://doi.org/10.62347/ULTA9461 Candidate biomarkers for COVID-19 in pediatric leukemia ments intended to treat their blood cancer [4]. These studies highlight the possible association between COVID-19 infection and AML. The goal of our project is to identify genomewide candidate genes which could serve, in the context of miRNA targeting mechanisms, as biomarkers of COVID-19 and leukemia. Many studies show that children are less susceptible to contracting COVID-19 and are also less susceptible to developing AML compared to adults. Children’s heightened immunity to COVID-19 has been proposed to stem from their high expression of the IFIH1 gene, which encodes the MDA5 protein. It is known that MDA5 recognizes double-stranded RNA and prompts the innate immune response [5]. We used IFIH1 as an entry point of the study, analyzed it using multiple databases, and identified potential upstream regulatory miRNAs of IFIH1 and other potential target genes of IFIH1 that are dysregulated in AML. To examine the evident relationship between COVID-19 and AML, we used data from The Cancer Genome Atlas (TCGA), which has over 20,000 original tumor and matched normal samples from 33 different studied cancer types and is a part of a significant cancer genomics initiative project [6] (https://www.cancer.gov/ ccg/research/genome-sequencing/tcga). The Level 3 (de-identified) RNA expression data generated from TCGA are publicly available to the community. diseases and potential influences have also been reported previously in other studies [9]. Our study has identified several candidate genes and miRNAs in the context of miRNA targeting mechanisms, based on TCGA Acute Myeloid Leukemia (LAML) RNA sequencing data, that could serve as comorbidity biomarkers of diseases (i.e., Coronavirus disease and leukemia cancer). Material and methods Identification of miRNAs that regulate the hub gene IFIH1 and their other target genes Because previous studies have shown that high expression of IFIH1 in children plays a role in their resistance to COVID-19, we first decided to search for miRNAs that target IFIH1. Using TarBase 7.0 [10], we inputted IFIH1 and selected “Homo sapiens” as the species. Two upstream miRNAs (hsa-miR-196a-5p and hsamiR-196b-5p) from the same family that had a prediction score greater than 0.7 were identified. Since miRNAs usually target multiple genes, we also decided to search for other genes targeted by the two miRNAs using TarBase. There were 77 targeted genes reported for hsa-miR-196b-5p and 101 targeted genes reported for hsa-miR-196a-5p. Evaluation of expression of the target genes of miRNAs from TCGA RNA-Seq data We also wanted to incorporate microRNAs (or miRNAs) into our study, since miRNAs are important regulators of cell function and gene expression and they have also not been widely examined in COVID-19 and leukemia. MicroRNAs are small (~17-22 bp) non-coding RNAs that can regulate gene expression through many mechanisms, including binding to the 3’ Untranslated Region (UTR) of an mRNA and thus reduc (...truncated)