Acute myeloid leukemia (AML) is most common in older adults. However, it is rarely cured by standard chemotherapy alone in older patients. According to NCI, it is critical that complete remission occurs, or there is no survival benefit. There are many clinically active therapies, including epigenetic drugs. A big translational research goal is to develop effective therapeutic strategies to demonstrate how and when oncologists should use these therapies. A clinical combinational treatment regime could be guided in part by genome-wide methylation profiling. Genes in CpG islands and their promoters have established aberrant methylation patterns in cancers. Perhaps also genome-wide methylation will help reveal new mechanistic details leading to new drug discoveries.
Reduced representation bisulfite sequencing (RRBS) was introduced by scientists at the Whitehead institute in Cambridge, Massachusetts in 2005. Recently RRBS was published as a Nature protocol. The regular Bisulfite sequencing method is considered the gold standard in methylation profiling. It provides single base pair resolution. The challenge of Bisulfite-seq is how laborious and expensive it is to sequence an entire genome. RRBS meets this challenge by targeting CpG-rich areas of the genome, via a shot gun cloning type library construction. Very little genomic DNA is required for the work flow. You digest it by the methylation-insensitive restriction enzyme MspI, which recognizes CCGG. You then end repair, Poly-A tail, and ligate into next generation sequencing adapters. You excise your fragments from a gel, convert with bisulfite, amplify by PCR, and sequence. The RRBS technique is well suited for characterising precious AML human bone marrow samples. Oncologists now use cytological AML subtyping. RRBS represents an epigenetic view, more finely discriminating AML cancer subtypes.
In their PloS Genetics paper, Altuna Akalin et al. cleverly expanded their fragment size selection to account for CpG shores and intragentic methylation sites, thereby enriching the methylation profiling information collected. They also reduced their bisulfite conversion to one step, preserving sample quantities. To quote their impressive stats, increases of 75% more CpG sites, 54% more CpG shores, 11.9% more promoter regions and 58% more introns. Essentially they demonstrated AML sub-type methylome diversity by comparing more of the CpG Islands and their promoters, but also a greater portion of the CpG shores and distal intergenic DNA. The group saw a wide variance in divergent methylation profiles overall. But they also defined a core set of shared leukemic differentially methylated sites. With this richer information in hand about AML subtype methylomes, the authors suggest a future goal of looking at AML transcriptomes and non-coding RNA.
I hope this work will contribute to effective medical treatment especially for older and elderly AML patients, as soon as possible. The title of the paper states “…profoundly divergent Epigenetic Landscapes in Acute Myeloid Leukemia“. This kind of data represents the promise of epigenetics and cancer research. That promise is definitely profound.
Akalin A, Garrett-Bakelman FE, Kormaksson M, Busuttil J, Zhang L, Khrebtukova I, Milne TA, Huang Y, Biswas D, Hess JL, Allis CD, Roeder RG, Valk PJ, Löwenberg B, Delwel R, Fernandez HF, Paietta E, Tallman MS, Schroth GP, Mason CE, Melnick A, & Figueroa ME (2012). Base-pair resolution DNA methylation sequencing reveals profoundly divergent epigenetic landscapes in acute myeloid leukemia. PLoS genetics, 8 (6) PMID: 22737091