DNA methylation plays an important role in gene expression, chromatin stability, and genetic imprinting. In mammals, DNA methylation patterns are written and regulated by DNA methyltransferases (DNMTs), including DNMT1, DNMT3A and DNMT3B. Recent emerging evidence shows that defects in DNMTs are involved in tumor transformation and progression, thus indicating that epigenetic disruptions caused by DNMT abnormalities are associated with tumorigenesis. Herein, we review the latest findings related to DNMT alterations in cancer cells and discuss the contributions of these effects to oncogenic phenotypes.

DNMT3A mutations are frequently discovered in acute myeloid leukemia (AML), associated with poor outcome. Recently, a relapse case report of AML extramedullary disease has showed that AML cells harboring DNMT3A variation were detected in the cerebral spinal fluid. However, whether a causal relationship exists between DNMT3A mutation (D3Amut) and extramedullary infiltration (EMI) is unclear. We took advantage of DNMT3A (R882C) mutation-carrying AML cell strain, that is, OCI-AML3, assessing its migration ability in vitro and in vivo. By RNA interfering technology and a xenograft mouse model, we evaluated the effect of DNMT3A mutation on cell mobility and explored the possible mechanism. OCI-AML3 displayed extraordinary migration ability in vitro and infiltrated into meninges of NOD/SCID mice after intravenous transfusion. We found that this leukemic migration or infiltration capacity was significantly compromised by the knockdown of DNMT3A mutant. Notably, TWIST1, a critical inducer of epithelial–mesenchymal transition, which underlies the metastasis of carcinomas, was highly expressed in association with R882 mutations. Abrogation of TWIST1 in DNMT3A mutated cells considerably weakened their mobility or infiltration. Our results demonstrate that D3Amut in OCI-AML3 strain enhances leukemic aggressiveness by promoting EMI process, which is partially through upregulating TWIST1.

Purpose: The resistance to differentiation therapy and early death caused by fatal bleeding endangers the health of a significant proportion of APL patients. This study aims to investigate the molecular mechanisms of ATRA resistance and uncover new potential therapeutic strategies to block the rapid progression of early death. Experimental Design: The important role of TWIST1 in APL leukemogenesis was first determined by gain- and loss-of-function assays. We then performed in vivo and in vitro experiments to explore the interaction of TWIST1 and TRIB3 and develop a potential peptide-initiated therapeutic opportunity to protect against early death and induction therapy resistance in patients with APL. Results: We found that the epithelial-mesenchymal transition (EMT)-inducing transcription factor TWIST1 is highly expressed in APL cells and is critical for leukemic cell survival. TWIST1 and TRIB3 were highly co-expressed in APL cells compared with other subtypes of acute myeloid leukemia (AML) cells. We subsequently demonstrated that TRIB3 could bind to the WR domain of TWIST1 and contribute to its stabilization by inhibiting its ubiquitination. TRIB3 depletion promoting TWIST1 degradation reverses resistance to induction therapy and improves sensitivity to ATRA. Based on a detailed functional screen of synthetic peptides, we discovered a peptide analogous to the TWIST1 WR domain that specifically represses APL cell survival by disrupting the TRIB3/TWIST1 interaction. Conclusions: Our data not only define the essential role of TWIST1 as an EMT-TF in APL patients but also suggest that disrupting the TRIB3/TWIST1 interaction reverses induction therapy resistance and blocks rapid progression of APL early death.

Antitumor activity of CD8⁺ T cells is potentially restrained by a variety of negative regulatory pathways that are triggered in the tumor microenvironment, yet, the exact mechanisms remain incompletely defined. Here, we report that intrinsic RIG-I in CD8⁺ T cells represents such a factor, as evidenced by observations that the tumor-restricting effect of endogenous or adoptively transferred CD8⁺ T cells was enhanced by intrinsic Rig-I deficiency or inhibition, with the increased accumulation, survival, and cytotoxicity of tumor-infiltrating CD8⁺ T cells. Mechanistically, T cell activation–induced RIG-I upregulation restrained STAT5 activation via competitive sequestering of HSP90. In accordance with this, the frequency of RIG-I⁺ tumor-infiltrating CD8⁺ T cells in human colon cancer positively correlated with attenuated survival and effector signatures of CD8⁺ T cells as well as poor prognosis. Collectively, these results implicate RIG-I as a potentially druggable factor for improving CD8⁺ T cell–based tumor immunotherapy.

Retinoic acid inducible gene-I (Rig-I) has been well documented as a cytosolic pattern recognition receptor that can sense viral RNA ligands to initiate the interferon-mediated antiviral immunity. However, little is known about the biological behaviors of Rig-I devoid of viral infection. Herein, we investigated the roles of Rig-I in the regulation of cellular senescence. In comparison to wild-type counterparts, Rig-I^-/- mice displayed the accelerated loss of hair, less responsiveness to gentle physical stimuli and shorten survival time. Likewise, Rig-I deficiency rendered mouse embryonic fibroblasts (MEFs) more susceptible to the serial passages-associated replicative senescence. By performing a transcriptome analysis, we identified integrins at the intersections of biological pathways affected by Rig-I. Among these, integrin β3 was negatively regulated by Rig-I, and significantly upregulated with the occurrence of senescence. Gene silencing of Itgb3 (encoding integrin β3) retarded the progression of cellular senescence in both WT and Rig-I^-/- MEFs. Notably, this effect was more prominent in Rig-I^-/- MEFs. Furthermore, p38 MAPK was a key downstream molecule for integrin β3-mediated senescence, and overactivated in senescent Rig-I^-/- MEFs. Taken together, Rig-I deficiency contributes to cellular senescence through amplifying integrin β3/p38 MAPK signaling. Our findings provide the evidence that Rig-I is a key regulator of cellular senescence, which will be helpful in better understanding its function without viral infection. Abbreviations: Rig-I: retinoic acid inducible gene-I; SASP: senescence-associated secretory phenotype; ECM: extracellular matrix; Itgb3: integrin beta 3; PRR: pattern recognition receptor; MEFs: mouse embryonic fibroblasts; Il-1β: interleukin-1 beta; Il-6: interleukin-6; Il-8: interleukin-8; Cxcl1: chemokine (C-X-C motif) ligand 1; Ccl2: chemokine (C-C motif) ligand 2; WT, wild type; BM: bone marrow; MAPK: mitogen-activated protein kinase; ERK: extracellular signal-regulated kinases; JNK: Jun N-terminal kinases; SA-β-gal: senescence-associated β-galactosidase; qPCR: quantitative reverse-transcription PCR; PBS: phosphate-buffered saline

Multiple myeloma (MM) relapse still occurs after a durable response to anti-BCMA chimeric antigen receptor-engineered T (CAR-T) cell therapy with less-defined factors. Herein, we investigated a CAR-T-exposed MM patient who relapsed after 12 months of remission by single-cell transcriptome sequencing. The bone marrow CAR-T population at relapse exhibited exhaustion and proliferation attenuation. The recurrent myeloma cells were deficient in or weakly expressed TNFRSF17 (BCMA) but possessed an identical immunoglobulin clonality to the baseline tumor. Interestingly, combined with the transcriptome profile of the myeloma strains, MM cells with BCMA negativity featured high ANXA1 expression that was identified as an inferior prognostic indicator for MM patients. At a single-cell resolution, BCMA-negative myeloma could be present in the MM patients without CAR-T cell exposure and displayed an increased level of intrinsic ANXA1 transcripts. In vitro assays unveiled that ANXA1 elevation conferred growth capacity to BCMA-negative myeloma cells via AMPKα signaling activation and disturbed CAR-T cell fitness. Blockade of ANXA1 reduced BCMA-negative myeloma cell proliferation. Murine models further demonstrated that ANXA1 inhibition could effectively diminish BCMA-negative myeloma that escaped from CAR-T's attack. Together, our data identified ANXA1 as a potential target for BCMA-negative myeloma clearance. The ANXA1-targeting strategy might be helpful to CAR-T treatment optimization.