TET2 deletion

In order to gain insight on the TET mediated molecular events that safeguard T cells from aberrant proliferation we performed serial adoptive transfers of murine CD4 T cells that lack concomitantly TET2 and TET3 to fully immunocompetent congenic mice. Here we show a progressive acquisition of malignant traits upon loss of TET2 and TET3 that is characterized by loss of genomic integrity, acquisition of aneuploidy and upregulation of the protooncogene Myc.

Classically, cytogenetic aberrations are not common except in cases of SM associated with another haematological neoplasm. We highlight here an unusual clinical presentation of SM and demonstrate the utility of advanced cytogenetic analysis (optical genome mapping, OGM) in detecting a novel cytogenetic abnormality resulting in an unusual mechanism of DNMT3A and TET2 loss of function.

Our findings underscore that Tet2 in macrophages negatively regulates liver regeneration by interacting with Stat1. Targeting Tet2 in macrophages promotes liver regeneration and function after a hepatectomy, presenting a novel target to promote liver regeneration and function.

Using aged mouse models and human progenitors, we demonstrate that targeting IL1 signaling could represent an early intervention strategy in preleukemic disorders. In summary, our results show that Tet2 is an important mediator of an IL1β-promoted epigenetic program to maintain the fine balance between self-renewal and lineage differentiation during hematopoiesis.

To study pharmacologic Brd4 inhibition in Tet2 KO mice, we utilized PLX51107, a structurally distinct BET inhibitor previously reported on by our laboratory...Our novel inducible double knockout mouse model of CH achieved simultaneous knockout of Brd4 and Tet2, which has never been investigated. Collectively, our data provides rationale for further pre-clinical investigation of BET inhibitors to prevent progression of Tet2 CH.

The prognostic influence of bone marrow blasts varied in the individual genetic subgroups, suggesting that the clinical impact of increased blasts depends on the genetic context. The molecular taxonomy derived in this study is clinically relevant and will inform future classification schemas.

To explore whether TET2-CH might identify patients primed for ICB response, we screened for CH in the prospectively collected baseline PBMCs from 90 solid tumour patients treated in a Phase 2 trial of the PD-1 inhibitor Pembrolizumab (NCT02644369)...Mechanistically, this occurs through lineage-specific promoter hypermethylation and gene silencing. Further study of TET2-CH as a potential biomarker of ICB response is warranted and targeting TET2-regulated pathways may yield novel strategies to improve immunotherapy outcomes.

We performed competitive bone marrow transplants (bmt) from tamoxifen‐treated C57BL/6J CD45... CH‐mutations lead to resistance to dab/tram in murine ATC. Our work opens new avenues for the selective targeting of tumor‐promoting inflammation in ATC.

MC-38 colon adenocarcinoma cells were subcutaneously implanted and mice were treated with anti-PD-1 or control antibodies +/- cell depleting antibodies or clodronate...Tet2-het TILs exhibited hypermethylation of genetic pathways regulating leukocyte differentiation.ConclusionTET2-CH is associated with increased odds of 6 month clinical benefit in immunotherapy-treated melanoma patients. In a mouse TET2-CH model, Tet2-loss of function reshapes the TIL epigenome and enhances effector/memory fate while relieving exhaustion and inhibiting regulatory T cell fate, to drive immunotherapy response.

Palliative drugs commonly used are AML-like chemotherapy, hypomethylating agents, hydroxyurea (HU), or PEG– IFN–α, ruxolotinib and trametinib. Atypical Chronic Myeloid Leukemia, BCR::ABL1 negative (aCML) is a rare hematological entity, with a dismal prognosis. Molecular landscape is heterogenous. Further comprensive molecular profiling may allow to find new targeted treatments in the next future.

p38 inhibition in combination with chemotherapy plus TKI treatment leads to greater depletion of Flt3-ITD AML LSC compared with CXCL12 deletion. Our studies support roles for CXCL12 and p38 signaling in microenvironmental protection of AML LSC and provide a rationale for inhibiting p38 signaling to enhance Flt3-ITD AML targeting.

Cancer cell. 2018 Jan 8; 33(1):44-59.e8.

Given that these mechanisms are also relevant in FLT3-ITD AML, we investigated the efficacy of CFZ in targeting FLT3-ITD mutant AML cells by itself and in combination with TKI.Method : FLT3-ITD mutant MV4-11 and MOLM13 cell lines and primary AML CD34+ cells were treated with CFZ as a single agent or in combination with FLT3 TKIs, Quizartinib (AC220) and Giltertinib followed by determination of cell cytotoxicity using CellTiter-Glo, apoptosis using Annexin V labeling, and differentiation using cell surface markers. We are currently investigating the effect of CFZ alone and with TKI in a large panel of FLT3-ITD AML samples and analyzing the role of STAT5 inhibition and increased oxidative stress in mediating the observed effects.Conclusion : Our studies indicate that the FDA-approved anti-leprosy agent CFZ demonstrates activity against FLT3-ITD+ AML stem and progenitor cells and significantly enhances FLT3 TKI targeting of these populations. These results support further investigation of CFZ as a potential therapeutic agent in FLT3-ITD+ AML.

The TET2KO/KD cells were 2-10X more sensitive to olaparib and rukaparib compared to parental TET2 proficient cells, both under steady state and under conditions of oxidative stress as observed in gH2AX assays and various apoptotic readouts...Loss of TET2 impairs this process, explaining the accumulation of secondary hits in mutant clones, which by this virtue, progressively increases their leukemogenic fitness. Given that MSH6 is known to interact 5hmC, TET2 may guide MMR to TET2-dependent DNA loci directly linking TET2MT to genomic instability a biologic feature that can be therapeutically exploited.

The combination of GSK-591 or LLY-283 with either of the FLT3 TKIs Quizartinib or Giltertinib resulted in synergistically enhanced inhibition of FLT3-ITD+ MOLM-13 and MV4-11 AML cells, compared to TKI or PRMT5 inhibitor alone. An epigenetic probe screen identified PRMT5 as a key regulator of FLT3-ITD AML cell viability. We show an important role for PRMT5 in maintenance of murine and human FLT3-ITD AML stem cells and in their persistence following FLT3 TKI treatment. Treatment with a PRMT5 inhibitor in combination with a FLT3 TKI could be a promising approach to enhance elimination of FLT3-ITD AML stem cells.

We reproduce Srsf2/Tet2 co-operativity in vivo, yielding a disease with core characteristics of CMML, unlike single Srsf2 or Tet2 mutation. This model represents a significant step toward building high fidelity and genetically tractable models of CMML.

Finally, we profiled the epigenomes of in vivo JQ1 treated AML-derived CD8 + T cells by single-cell ATAC seq and find that JQ1 increases Tcf7 accessibility specifically in Tex cells, suggesting that BETi likely mechanistically acts by relieving repression of progenitor programs in Tex CD8+ T cells and maintaining a pool of anti-PD1 responsive CD8 T cells (figure 3). Conclusions Using an AML mouse model that exhibits leukemia-induced immune exhaustion, we demonstrate the pre-clinical efficacy of combining BETi and anti-PD1 therapy in the treatment of AML.

Azacitidine and ascorbate restored 5hmC abundance and slowed or reverted the expansion of TET2-mutant clones in vivo. These results demonstrate the key role of 5hmC in normal hematopoiesis and TET2-mutant phenotypes and raise the possibility of utilizing these agents to further our understanding of pre-leukemia/clonal hematopoiesis.

Response rates to azacitidine in PTCL of follicular helper cell origin are promising. Deletion of the histone methyltransferase SETD2 sensitised to HMA but TET2 deletion did not. Guadecitabine conveyed an acceptable ORR and toxicity profile; decitabine analogues may provide a backbone for future combinatorial regimens co-targeting histone methyltransferases.

In addition, TET2-deficient lymphomas select for high expression of the pro-survival BCL2 family protein BCLX L , potentially providing protection from MYC-driven apoptosis. Conclusion Altogether, our data concerning the impact of TET2 loss-of-function on DNA repair and apoptosis mechanisms creates understanding of how TET2 deficiency nudges transformation, and reveals testable therapeutic vulnerabilities.

In these studies, we investigated the effects of the BET inhibitor (BETi) JQ1 on T cell exhaustion and checkpoint responsiveness in a murine model of AML and AML patient samples...Finally, we show that BETi rescues anti-PD1 resistance in vivo and reduces tumor burden in multiple organ sites and enriches CD8+ T cells in the bone marrow. In total, we demonstrate that combining BETi and anti-PD1 therapy in the treatment of AML is a rational strategy to overcome anti-PD1 resistance.

Clustered regularly interspaced short palindromic repeats (CRISPR)-mediated depletion of nucleases and helicases that regulate G-quadruplexes and R-loops decreased the viability of TET-deficient B cells. Our studies suggest a molecular mechanism by which TET loss of function might predispose to the development of B cell malignancies.

We have generated primary murine AML-ETO9a+ and MLL-AF9+ leukemia models with Tet2 +/+, Tet2 +/- and Tet2 -/- alleles to determine the Tet2-dependent efficacy of PARPi treatment when combined with vitamin C. Furthermore, we have performed CRISPR gene knockout and drug library screening in human AML cell lines in combination with vitamin C treatment, and tested a panel of 10 AML cell lines with titrating concentrations of PARPi (Olaparib, Talazoparib, Veliparib and Rucaparib) alone or in combination with vitamin C (L-ascorbic acid) mimicking physiological to pharmacological in vivo doses. Our findings confirm that vitamin C can act synergistically with PARPi to block AML cell viability, reduce colony-forming capacity, and decrease leukemia burden in PDX and primary murine leukemia models in a TET2 allelic dose-dependent manner. The combinatorial effect works at clinically relevant concentrations of PARPi, and low-pharmacological doses of vitamin C. These studies suggest that vitamin C can be used as a non-toxic therapeutic adjuvant to PARPi therapy for the treatment of AML.

Additionally, we show that chemical inhibition of p300 acetyltransferase activity phenocopied Ep300 deletion in Tet2-deficient HSPCs, whereas activation of p300 activity with a small molecule impaired the self-renewal and leukemogenicity of Tet2-deficient cells. This suggests a potential therapeutic application of p300 activators in the treatment of MDS with TET2 inactivating mutations.

Methods 104 NSCLC patients who received platinum-based chemotherapy were selected from TCGA: platinum-etoposide (N=18), platinum-gemcitabine (N=20), platinum-vinorelbine (N=31), platinum-paclitaxel (N=21), and platinum-docetaxel (N=14). By using novel biomarkers, a CBM-informed decision tree can be employed to identify the optimal drug combination for platinum-based therapy. We suggest that this approach be validated prospectively in a larger patient cohort.

SIGNIFICANCE: Loss-of-function somatic TET2 mutations are among the most frequent lesions in myeloid neoplasms and associated disorders. Here we report a strategy for selective targeting of residual TET dioxygenase activity in TET-deficient clones that results in restriction of clonal evolution in vitro and in vivo.

Cell and animal models with concurrent mutations in TET2 and DNMT3A were sensitive to T-dCyd treatment. The mutations were detectable in human solid tumors and frequently occur in some hematological malignancies.

Transcriptome analysis revealed an important dysregulation in gene expression accompanied by aberrant mRNA splicing. This study provides new insights into the biology of this MDS subtype and opens the possibility to more in-depth interrogate its molecular disease mechanism.

Unlike ARIH2-WT, ectopic expression of ARIH2K441N or ARIH2R484I blocked FGFR1 ubiquitination and degradation, activating STAT5, AKT and ERK1/2 signaling, suggesting that TET2 loss leads to secondly hits, which activate proliferation/survival signaling. Taken together, our finding provides a rationale for enhancing TET2 function to block MDS-malignant transformation.

NeoTCR-T cells with concomitant deletion of TET2-expression exhibited enhanced cytotoxicity against neoantigen-expressing tumor cells. Together, these results demonstrate the applicability of this versatile precision genome engineering platform technology to yield enhanced next generation neoTCR-T cell therapies to expand the potential for clinical benefit in treating persons with solid cancers.

Treatment with TET inhibitor suppressed the clonal evolution of TET2 mutant cells in murine models and TET2-mutated human leukemia xenografts. These results suggest that TET inhibitors may constitute a new class of targeted agents in TET2 mutant neoplasia.