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MiR-146a deletion is a driving molecular event in del(5q) myeloid malignancies, which acquire dependency on a nuclear factor kappa B (NF-κB) regulatory network. p62, a neighboring haploid 5q gene, is induced by NF-κB and required to sustain TRAF6-mediated NF-κB activation. Interfering with p62/TRAF6 binding may have a therapeutic benefit in miR-146a–deficient leukemic cells.
Myelodysplastic syndromes (MDS) are hematopoietic stem cell (HSC) disorders that can evolve into aggressive forms of acute myeloid leukemia (AML). MDS and AML occur as a result of genomic instability and somatic mutations within disease-initiating and/or disease-propagating hematopoietic stem/progenitor cells (HSPCs). A common genetic alteration in MDS and secondary AML involves deletion of a single copy of the long arm of chromosome 5 (del[5q]). Although patients with isolated del(5q) and few blasts in the bone marrow (BM) have a favorable prognosis, the majority of MDS/AML patients with del(5q) have elevated BM blasts (>5%), complex cytogenetics, and poor prognosis. Treatment failure is often observed in this cohort of higher-risk (HR) patients after standard chemotherapy and allogeneic HSC transplantation. Recently, hypomethylating agents, such as 5-azacitidine, and immunomodulators, such as lenalidomide/revlamid, have demonstrated modest responses in HR del(5q) MDS/AML patients, either in combination or as single-agent therapies. Despite encouraging responses to hypomethylating and immunomodulatory therapy, progression of the disease invariably advances. Therefore, strategies to improve the outcomes of HR del(5q) MDS/AML patients are needed. Understanding the molecular circuitry contributing to the pathogenesis, disease evolution, and maintenance of del(5q) MDS/AML will likely uncover novel therapeutic opportunities. Commonly deleted regions have been mapped on chr 5q: a distal region at 5q33.1 and a proximal region at 5q31.1. Over the last 2 decades, multiple genes have been identified that contribute to the pathogenesis of del(5q) MDS/AML. One such gene, microRNA-146a (miR-146a), is located adjacent to the distal deleted region and is implicated in the pathogenesis of human MDS. Knockout of miR-146a in mice results in features of an MDS-like disease including an early onset of myeloid expansion in the BM, and progression to more aggressive diseases such as lymphomas, BM failure, and myeloid leukemia.1-3 At the molecular level, tumor necrosis factor (TNF) receptor-associated factor 6 (TRAF6), a signal transducer of nuclear factor (NF)-κB in response to Toll-like receptor-family activation, is a well-characterized target of miR-146a and important for the hematopoietic phenotype in miR-146a–deficient cells.1,3
In our recent study, we show that miR-146a is significantly downregulated in advanced del(5q) MDS and AML, and that deletion of miR-146a modifies the cellular function of MDS/AML-HSPC in del(5q) disease, rendering the cells more aggressive.4 As expected, this occurs by derepression of TRAF6 and activation of NF-κB. Therefore, we postulated that inhibiting the TRAF6/NF-κB axis may represent a novel therapeutic opportunity for aggressive forms of del(5q) MDS/AML, particularly for patients with low miR-146a expression. Unfortunately, the success of specific TRAF6 and NF-κB inhibitors as agents against human cancers has yet to be proven. To overcome the limitations of directly targeting the TRAF6/NF-κB hub, we explored alternate possibilities. Inactivation of tumor suppressors as a result of large chromosome deletions, as observed with loss of miR-146a in del(5q) MDS/AML, often involves deletion of innocuous neighboring genes. However, haplodeficiency of neighboring genes can also create therapeutic vulnerabilities unique to the tumor cells.5,6 By applying this concept to del(5q) MDS/AML, we reasoned that dysregulation of neighboring chr 5q genes may modify the TRAF6/NF-κB pathway in such a way that exposes a genetic vulnerability unique to MDS/AML with haploid/reduced miR-146a levels. To test this hypothesis, we applied an integrative genomics and bioinformatics approach and identified a subset of genes that are overexpressed from the intact 5q allele. Interestingly, the overexpressed (i.e., compensated) genes in del(5q) MDS/AML collectively establish an NF-κB signaling network (Fig. 1A). Furthermore, detailed promoter analysis confirmed enrichment of NF-κB binding sites within the promoter of the compensated genes, suggesting that a NF-κB feed-forward loop exists and may be necessary to sustain the NF-κB signaling pathway following deletion of miR-146a. As one example, sequestosome 1 (SQSTM1; best known as p62), a neighboring haploid gene that is expressed from the intact 5q allele (5q35), is required to sustain miR-146a–deficient MDS/AML cells (Fig. 1B). p62 regulates several cellular functions, including bone remodeling, metabolism, and cancer.7 One of the most-studied functions of p62 is its role in regulating NF-κB activation; p62 recruits TRAF6, which then initiates NF-κB signaling. An essential role of p62 in TRAF6/NF-κB activation has been described in solid tumors,8 and also recently in the survival of AML cells during all-trans retinoic acid-induced differentiation.9 Given that concomitant loss of miR-146a and overexpression of p62 cooperate to enhance TRAF6/NF-κB signaling by derepressing and activating TRAF6, respectively, we hypothesize that del(5q) MDS/AML cells are vulnerable to disruption of the p62-TRAF6 complex. As expected, knocking down p62 or disrupting p62-TRAF6 binding results in cell cycle arrest and apoptosis in MDS/AML cell lines and patient samples. Importantly, depletion of p62 has minimal effects on survival, proliferation, or function of normal HSPCs. These findings suggest that, compared to normal cells, del(5q) myeloid malignancies acquire a dependency on an intrachromosomal NF-κB gene network, which can be targeted for therapeutic purposes.
Deletions involving chr 5q in MDS/AML span several megabases and involve multiple neighboring genes. In fact, very few del(5q) MDS/AML patients are reported to have microdeletions, suggesting that multiple genes on chr 5q are important for the malignant phenotype. In this study, we found that concomitant loss of miR-146a and overexpression of p62 sustain survival and proliferation of del(5q) malignant cells through TRAF6/NF-κB, thus providing a therapeutic window for HR del(5q) MDS/AML. Collectively, these observations establish novel paradigms in targeting cancer, and underscore the genetic and molecular complexity of these malignancies. In addition, we provide a proof-of-concept that intra- and/or inter-chromosomal gene networks may expose therapeutic vulnerabilities in cancers with genomic instability.
No potential conflicts of interest were disclosed.
The manuscript was written by DTS and JF.