N6-methyladenosine (N6-甲基腺苷, m6A) is a modification formed by adding a methyl group (-CH3) to the sixth nitrogen atom of adenosine. This modification is widely found in the RNA molecules of various organisms and is the most common internal (non-capping) modification in messenger RNA (mRNA) in all higher eukaryotes. It can also modify viral RNA and exhibits antiviral activity. N6-methyladenosine antibodies, known for their high specificity and sensitivity, have broad application value in areas such as epigenomics, oncology, neuroscience, and stem cell regenerative medicine. This article reviews the biological functions of N6-methyladenosine antibodies, their applications in scientific research and clinical settings, and explores future research directions.

M6A was first discovered in mammalian cells by Desrosiers.R et al. in the 1970s.

However, the research on the functions and mechanisms of m6A has not been sufficiently in-depth. It was not until the discovery of the m6A demethylase FTO that it was revealed that m6A modifications are reversible. Since then, more enzymes related to m6A modifications have been discovered, and their mechanisms of action have been studied in depth. Research indicates that m6A influences RNA stability, translation efficiency, splicing, and nuclear export, thereby regulating various biological functions, playing a crucial role in important biological processes such as embryonic development, neural cell development, and stem cell fate determination.

M6A antibody is the core tool for studying RNA epigenetic modification. The core function of M6A antibody is to enrich m6A-modified RNA fragments containing m6A through immunoprecipitation (such as m6A-seq, technology), so as to realize high-throughput detection of m6A sites in the whole transcriptome, which is a key tool for analyzing m6A epigenetic transcriptome.

▌ N6-methyladenosine antibody for scientific research

• Neurodegenerative diseases

In neurodegenerative diseases, m6A antibodies have helped identify FTO, and the abnormal m6A modifications can regulate the mRNA stability of the β-amyloid precursor protein (APP). By using m6A immunoprecipitation sequencing (MeRIP-seq) technology, m6A antibodies can be enriched and sequenced to identify m6A-modified RNA at the whole transcriptome level, thereby elucidating the regulatory mechanisms of these modifications on neurogenic gene expression. Furthermore, single-cell sequencing techniques can analyze the heterogeneity of m6A modifications at the single-cell level across different types of neurons, such as neurons, astrocytes, and microglia.

• Metabolic disease research

m6A modification regulates the expression of genes involved in metabolism, impacting glucose and lipid metabolism as well as organ function. In diabetes, m6A modification in adipocytes, mediated by YTHDF1, enhances the translation of GLUT4 mRNA, thereby improving insulin sensitivity. In obesity models, a decrease in m6A levels is associated with insulin resistance. In non-alcoholic fatty liver disease (NAFLD), METTL3-mediated m6A modification enhances the stability of PPARγ mRNA, promoting fat production. Inhibiting METTL3 can improve hepatic steatosis in mice.

• Cardiovascular Disease Study

Cardiomyocyte apoptosis and remodeling: In the myocardial ischemia model, m6A modification affects cell apoptosis by regulating the stability of mRNA of Bcl-2, a family gene; while overexpression of ALKBH5 demethylase can reduce myocardial injury.

Vascular smooth muscle cell proliferation: m6A antibody detection revealed that after vascular injury, the level of m6A modification increased, and the degradation of mRNA of proliferative related genes (such as Cyclin D1) was reduced by YTHDF2, thus promoting the process of atherosclerosis.

▌ Clinical application of N6-methyladenosine antibody

• Tumor marker development organization

Detection of m6A levels in blood: Using immunohistochemistry (IHC) or ELISA techniques, the overall m6A modification level in tumor tissues or the expression of key regulatory enzymes (such as METTL3 and FTO) is assessed. This aids in tumor classification: In liver cancer, high m6A levels are associated with overexpression of METTL3, indicating a poor prognosis. In lymphoma, high FTO expression leads to reduced m6A levels, which is linked to chemotherapy resistance and an increased risk of recurrence. 

• Exploration of biomarkers for neurodegenerative diseases

Cerebrospinal fluid (CSF)/brain tissue m6A detection: In Alzheimer's disease (AD) patients, the level of m6A modification in CSF is reduced, which is associated with Aβ deposition and Tau protein phosphorylation. Using m6A antibodies to locate abnormally modified mRNA (e.g., APP and PSEN1) in neurons aids in validating the pathological mechanisms.

• Small molecule compounds target m6A enzyme

Methyltransferase inhibitors, such as METTL3 inhibitors (e.g., STM2457 and MI-503), can reduce the m6A level in tumor cells, leading to cell cycle arrest or apoptosis. These inhibitors have shown efficacy in animal models of leukemia and lymphoma. Demethylase inhibitors, like FTO inhibitors (e.g., MA2), can increase the m6A modification level, thereby inhibiting the self-renewal and metastasis capabilities of breast cancer stem cells. ALKBH5 inhibitors enhance the stability of IFN-γ mRNA in T cells, improving the response to tumor immunotherapy

▌Future clinical transformation exploration

• Disease diagnosis and biomarker development: The application scenarios of liquid biopsy involve developing non-invasive diagnostic tools by analyzing m6A-modified RNA (e.g., exosomal RNA) in blood, cerebrospinal fluid, or urine. Potential disease areas include: Cancer —— The unique m6A modification patterns in tumors (such as the METTL3-mediated increase in m6A in lung cancer) could serve as biomarkers for early diagnosis. Neurodegenerative diseases —— In Alzheimer's disease (AD), the abnormal m6A modifications are associated with pathological changes in tau protein. Metabolic diseases —— In obesity or diabetes, the dynamic changes in m6A levels regulated by FTO. By combining tissue pathology classification, immunohistochemistry (IHC) or in situ hybridization techniques using m6A antibodies can quantify m6A levels in tumor tissues, aiding in cancer grading (e.g., glioma, breast cancer).

• Target development and drug screening for m6A regulation proteins. Inhibitor development: FTO inhibitors, such as FB23-2, exert anti-tumor effects in acute myeloid leukemia (AML) by inhibiting m6A demethylation. METTL3 inhibitors, such as STM2457, target m6A-dependent tumors and are currently in clinical trials. Antibody-drug conjugates (ADCs) use m6A antibodies to deliver chemotherapy drugs to cancer cells with high m6A expression. RNA modification therapy involves using oligonucleotide drugs (such as ASOs or siRNAs) to specifically regulate the m6A modification levels of disease-related genes.

▌Reference Documentation

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