N6-methyladenosine (m6A) is the most abundant dynamic mark of messenger RNA where it may affect most posttranscriptional steps in gene expression, including splicing, stability and translation. This modification is indispensable for cell viability and development, yet its role in cancer is still poorly understood.
In mammals, the writer of m6A is a nuclear multicomponent complex composed of two methyltransferases METTL3 and METTL14, and the regulatory protein WTAP. METTL3 is the catalytic component while METTL14 functions in structural stabilization and RNA substrate recognition. Notably, WTAP has been recently described as an oncogenic factor in acute myeloid leukemia (AML). Moreover, we have observed a general increase of METTL3 and METTL4 expression in AML, suggesting that their alteration might contribute to leukemogenesis.
Deregulation of the m6A complex in AML cell lines (our preliminary data), strongly induces apoptosis. Thus, a major aim of this proposal is to prove whether the inhibition of the formation of m6A modification could have therapeutic value in AML. Our preliminary data show that in AML cells the METTL3 catalytic component is mis-localized in the cytoplasm where it associates with translating ribosomes. Thus, indicating additional function for METTL3 independent from the nuclear methylation complex. Therein, it will be critical to establish if the phenotypes observed in AML cells are indeed methylation dependent and not due to secondary functions of the associated enzymes.
To address these questions, we propose the following aims: 1) To address the contribution of METTL3 catalytic activity in AML and normal CD34+ hematopoietic stem cells (HSC) proliferation and, 2) to assess the role of cytoplasmic METTL3 in AML cells.
Epigenetic alterations are potentially reversible, unlike genetic mutations. Therefore, such alterations have become an attractive target for cancer therapy.
Following the identification of the contribution of epigenetic programs to the regulation of gene expression during normal and pathological hematopoietic cell fate determination, many efforts have been focused on the development of chromatin modifying-based therapeutics strategies for human hematological cancers. Growing evidence indicates that the targeting of the post-transcriptional reversible mRNA modification (RNA epigenetics), which regulates the genomic output and mRNA function, could also represent a central tool for the development of novel RNA-based drugs and therapies. In this context, different strategies focused to alter the post-transcriptional RNA modification status, targeting the RNA methyltransferase complex, may be relevant to disclose innovative and significant molecular pathways at the basis of the differentiation block and aberrant proliferation that characterized the Acute Myeloid Leukemia (AML) phenotype.
Results obtained from our proposal will contribute to important advances in the search for novel therapeutic strategies for human AML. The ability to identify and modify the post-transcriptional RNA modification status by the use of RNA interference technology or selective compounds targeting the RNA methyltransferase complex may reveal possible future therapeutic applications for patient treatments. In addition, a potential outcome of the project may be the identification of specific RNAs modified at posttranscriptional level as selective diagnostic markers for the characterization of a specific human AML subtype. Therefore, based on results from this proposal, a patent might be filed for: i) therapeutic use of post-transcriptional RNA modification status (through the use of RNA interference technology or a selective compound targeting the RNA methyltransferase complex), and ii) diagnostic/prognostic markers specific for a specific AML subtype by the characterization of the methylation status of each molecule of RNA. In line with this, results of our studies may lead to possible commercial applications of the modification status of the transcriptome in the clinical setting.
The advancement of knowledge in AML RNA epigenetics will provide useful information to develop a higher level of knowledge on AML tumorigenesis for the identification of innovative tools or targets to design novel therapeutic strategies. Our results will promise a significant reinforcement of the capability to address not only a specific therapeutic strategy for AML, but also to enhance the diagnosis and prognosis of this haematological malignancies.
The activities conducted in the context of this research project will be the starting point for the creation of significant intellectual property (patenting of new biological target for the design of compound potentially interesting for therapy). Along this line, the main results expected from this project are:
- Identification of novel therapeutic targets for pharmacological therapy (targets for drug screening and design) and biological therapy (small molecules-based therapy) for AML cells.
- Identification of new molecular pathways that could be explored as markers for diagnosis and/or prognosis of AML cells (diagnostic, prognostic array for the evaluation of the methylation status of specific RNA molecules).
Our technological advancement will be also shared with the international scientific community, to provide a molecular and biological advance that is strongly needed in this field of cancer research, and to guarantee important improvements in the management and effectiveness of treatments of leukemia.