PROJECT SUMMARY
Multiple myeloma (MM), the 2nd most common hematologic malignancy, remains incurable with high rates of
relapse and drug resistance despite recent advances in treatment options such as autologous stem cell
transplantation, novel immunomodulatory agents, and proteasome inhibitors. Shorter duration of initial
response predicts poor prognosis even in the modern era of novel agents. Several clinical features and high-
risk cytogenetics are currently in use for stratifying patients by severity of disease. However, a substantial
number of patients continue to have poor outcomes without known negative prognostic factors. Aberrant
epigenetic modifications have been linked with more aggressive types and progression of MM. In addition to
DNA methylation and histone modifications, accumulating evidence suggests that post-transcriptional
regulation involving RNA modifications, particularly N6-methyladenosine (m6A), is associated with initiation and
progression hematological malignancies. Emerging evidence suggests that m6A is also involved in the
development and prognosis of MM. However, the role of m6A in outcomes of MM remains largely unknown due
partly to the lack of quantitative methods to map whole-transcriptome m6A at high resolution. Some notable
challenges of existing m6A profiling approaches include 1) low resolution; 2) not quantitative; and 3) the
requirement of a large amount of input materials that are typically not feasible in clinical settings. To address
these clinical and technical needs, we designed the m6A-selective allyl chemical labeling and sequencing
(m6A-SAC-seq), which features transcriptome-wide profiling of m6A at single-nucleotide resolution with
modification fraction information. The objective of this application is to use the m6A-SAC-seq to map
transcriptome-wide m6A in CD138+ tumor cells from MM patients to identify m6A modifications associated with
treatment outcomes. We will apply this novel technology in banked CD138+ cells collected at the time of
diagnosis for ~100 newly diagnosed patients with MM enrolled in our NCI-funded clinic-based survivorship
study as well as ~30 samples from a Phase II clinical trial. Our central hypothesis is that specific m6A
signatures in CD138+ tumor cells at the time of diagnosis correlate with outcomes in patients with MM. We will
identify altered m6A in transcripts associated with early relapse and response to first-line therapy (Aim 1) as
well as minimal residual disease (Aim 2). With respect to outcomes, we expect to 1) identify specific m6A
signatures and involved pathways that are associated with treatment outcomes among patients with MM; and
2) establish the m6A-SAC-seq for broader clinical applications. The proposed research is highly significant,
because it is expected to vertically advance understanding of the biological basis for treatment outcomes of
MM and promote the development of a new paradigm for prognostic risk stratification based on m6A
modifications that has broad translational importance in the personalized management of high-risk patients.