Inhibition of Topors Ubiquitin Ligase Augments the Efficacy of DNA Hypomethylating Agents through DNMT1 Stabilization

作者：Satoshi Kaito, Kazumasa Aoyama, Motohiko Oshima, Karl Agger, Kristian Helin, Yasuhito Nannya, Seishi Ogawa, Atsuya Nishiyama, Makoto Nakanishi, Atsushi Iwama · 发表于：Blood · 年份：2023 · DOI：10.1182/blood-2023-179684 · 被引用次数：2 · 研究领域：Epigenetics and DNA Methylation

DNA hypomethylating agents (HMAs) are commonly used to treat myeloid malignancies including myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML). However, their therapeutic effects are not satisfactory. We performed CRISPR-Cas9 knockout (KO) screening to identify molecular targets that enhance the efficacy of HMAs. MDS-L and MOLM-13 cells expressing Cas9 were infected with an sgRNA lentiviral library containing 12,409 sgRNAs targeting 1,383 epigenetic factors and exposed to low-dose HMAs, decitabine (DAC) or azacitidine (AZA), for 14 days. The read counts of sgRNA against TOPORS which encodes a ubiquitin/SUMO E3 ligase, were reproducibly decreased during culture specifically in the presence of HMAs in both screening. To validate the screening results, we performed competitive growth assays by co-culturing parental cells and TOPORS-KO cells using MOLM-13, MDS-L, SKK-1, and SKM-1 cells. In all cell lines, GFP-positive TOPORS-KO cells decreased over time only in the presence of HMAs. We established single-cell clones of TOPORS-KO MOLM-13 and MDS-L cells using two different sgRNAs. The cell growth of all TOPORS-KO clones was dramatically suppressed only in the presence of DAC. Mice transplanted with TOPORS-KO cells showed significantly longer survival when treated with DAC. Next, we analyzed Topors-deficient mice and no significant changes were observed in the peripheral blood or in the percentage of bone marrow (BM) hematopoietic stem and progenitor cells. Topors−/− B...