In a study reported in The New England Journal of Medicine, Liu et al found that hypomethylating therapy was associated with upregulation of the oncogene SALL4 in a high proportion of patients with myelodysplastic syndrome. They also identified the gene region that is demethylated and responsible for upregulation.
As stated by the investigators, SALL4 was selected for use as a model to test the hypothesis that treatment with hypomethylating agents may result in demethylation of oncogenes, as well as demethylation of tumor-suppressor genes. SALL4 plays an essential role in myelodysplastic syndrome and acute myeloid leukemia leukemogenesis, as well as a role in tumorigenesis in other solid tumors, including germ cell tumors, hepatocellular carcinoma, breast cancer, and lung cancer.
Study Details
The study involved evaluation of paired bone marrow samples obtained from two cohorts of patients with myelodysplastic syndrome before and after treatment with a hypomethylating agent to assess changes in SALL4 expression and potential effect on treatment outcomes.
Leukemic cell lines with low/undetectable SALL4 expression were used to assess the association of SALL4 methylation and expression. A locus-specific demethylation technology, CRISPR–DNMT1-interacting RNA (CRISPR-DiR), was used to identify the CpG region of SALL4 involved in gene expression.
Key Findings
SALL4 up-regulation after treatment with hypomethylating agents was observed in 10 (40%) of 25 patients in cohort 1 and in 13 (30%) of 43 patients in cohort 2. A decrease in SALL4 expression was observed in 15 patients (60%) in cohort 1 and 30 patients (70%) in cohort 2.
Compared with patients with SALL4 downregulation, those with SALL4 upregulation had poorer overall survival in both cohort 1 (P = .03) and cohort 2 (P = .04). On multivariate analysis, SALL4 upregulation was associated with poorer overall survival in both cohort 1 (hazard ratio [HR] for death = 6.48, 95% confidence interval [CI] = 1.06–39.67) and cohort 2 (HR = 2.74, 95% CI = 0.93–8.03).
Assessment with CRISPR-DiR showed that demethylation of a CpG region within the 5’ untranslated region (5’UTR) of SALL4 was critical for SALL4 expression.
Analysis in low SALL4 cell lines showed that exposure to a hypomethylating agent resulted in upregulation of SALL4 expression and demethylation of the SALL4 5’UTR CpG region.
In analysis of six of the study patients who exhibited SALL4 upregulation, decreased methylation at the 5’UTR CpG region was observed, whereas no changes in methylation status in this region were observed in nine study patients who exhibited SALL4 downregulation.
The investigators concluded, “By combining analysis of patient samples with CRISPR-DiR technology, we found that demethylation and upregulation of an oncogene after treatment with a hypomethylating agent can indeed occur and should be further studied.”
Daniel G. Tenen, MD, of Harvard Medical School; Li Chai, MD, of Brigham and Women’s Hospital; and Maria T. Voso, MD, of University of Rome Tor Vergata, are the corresponding authors for The New England Journal of Medicine article.
Disclosure: The study was funded by the Associazione Italiana per la Ricerca sul Cancro and others. For full disclosures of the study authors, visit nejm.org.
