Myelodysplastic syndrome (MDS) describes a family of blood disorders driven by the clonal expansion of mutated blood cells that can evolve into secondary acute myeloid leukemia (sAML). Two new ...studies use single-cell and deep sequencing to elucidate the progression of MDS to AML, revealing discrete clonal architectures and the driving role of signaling mutations.
The implementation of small-molecule and immunotherapies in acute myeloid leukemia (AML) has been challenging due to genetic and epigenetic variability amongst patients. There are many potential ...mechanisms by which immune cells could influence small-molecule or immunotherapy responses, yet, this area remains understudied.
Here we performed cell type enrichment analysis from over 560 AML patient bone marrow and peripheral blood samples from the Beat AML dataset to describe the functional immune landscape of AML.
We identify multiple cell types that significantly correlate with AML clinical and genetic features, and we also observe significant correlations of immune cell proportions with
small-molecule and immunotherapy responses. Additionally, we generated a signature of terminally exhausted T cells (T
) and identified AML with high monocytic proportions as strongly correlating with increased proportions of these immunosuppressive T cells.
Our work, which is accessible through a new "Cell Type" module in our visualization platform (Vizome; http://vizome.org/), can be leveraged to investigate potential contributions of different immune cells on many facets of the biology of AML.
Many acute myeloid leukemia (AML) patients exhibit hallmarks of immune exhaustion, such as increased myeloid-derived suppressor cells, suppressive regulatory T cells and dysfunctional T cells. ...Similarly, we have identified the same immune-related features, including exhausted CD8
T cells (TEx) in a mouse model of AML. Here we show that inhibitors that target bromodomain and extra-terminal domain (BET) proteins affect tumor-intrinsic factors but also rescue T cell exhaustion and ICB resistance. Ex vivo treatment of cells from AML mice and AML patients with BET inhibitors (BETi) reversed CD8
T cell exhaustion by restoring proliferative capacity and expansion of the more functional precursor-exhausted T cells. This reversal was enhanced by combined BETi and anti-PD1 treatment. BETi synergized with anti-PD1 in vivo, resulting in the reduction of circulating leukemia cells, enrichment of CD8
T cells in the bone marrow, and increase in expression of Tcf7, Slamf6, and Cxcr5 in CD8
T cells. Finally, we profiled the epigenomes of in vivo JQ1-treated AML-derived CD8
T cells by single-cell ATAC-seq and found that JQ1 increases Tcf7 accessibility specifically in Tex cells, suggesting that BETi likely acts mechanistically by relieving repression of progenitor programs in Tex CD8
T cells and maintaining a pool of anti-PD1 responsive CD8
T cells.
Summary
Drug resistance in chronic myeloid leukaemia (CML) may occur via mutations in the causative BCR::ABL1 fusion or BCR::ABL1‐independent mechanisms. We analysed 48 patients with ...BCR::ABL1‐independent resistance for the presence of secondary fusion genes by RNA sequencing. We identified 10 of the most frequently detected secondary fusions in 21 patients. Validation studies, cell line models, gene expression analysis and drug screening revealed differences with respect to proliferation rate, differentiation and drug sensitivity. Notably, expression of RUNX1::MECOM led to resistance to ABL1 tyrosine kinase inhibitors in vitro. These results suggest secondary fusions contribute to BCR::ABL1‐independent resistance and may be amenable to combined therapies.
Transits of exoplanets observed in the near-UV have been used to study the scattering properties of their atmospheres and possible star–planet interactions. We observed the primary transits of 15 ...exoplanets (CoRoT-1b, GJ436b, HAT-P-1b, HAT-P-13b, HAT-P-16b, HAT-P-22b, TrES-2b, TrES-4b, WASP-1b, WASP-12b, WASP-33b, WASP-36b, WASP-44b, WASP-48b, and WASP-77Ab) in the near-UV and several optical photometric bands to update their planetary parameters, ephemerides, search for a wavelength dependence in their transit depths to constrain their atmospheres, and determine if asymmetries are visible in their light curves. Here, we present the first ground-based near-UV light curves for 12 of the targets (CoRoT-1b, GJ436b, HAT-P-1b, HAT-P-13b, HAT-P-22b, TrES-2b, TrES-4b, WASP-1b, WASP-33b, WASP-36b, WASP-48b, and WASP-77Ab). We find that none of the near-UV transits exhibit any non-spherical asymmetries, this result is consistent with recent theoretical predictions by Ben-Jaffel et al. and Turner et al. The multiwavelength photometry indicates a constant transit depth from near-UV to optical wavelengths in 10 targets (suggestive of clouds), and a varying transit depth with wavelength in 5 targets (hinting at Rayleigh or aerosol scattering in their atmospheres). We also present the first detection of a smaller near-UV transit depth than that measured in the optical in WASP-1b and a possible opacity source that can cause such radius variations is currently unknown. WASP-36b also exhibits a smaller near-UV transit depth at 2.6σ. Further observations are encouraged to confirm the transit depth variations seen in this study.
To understand mechanisms of response to BET inhibitors (BETi), we mined the Beat AML functional genomic dataset and performed genome-wide CRISPR screens on BETi- sensitive and BETi- resistant AML ...cells. Both strategies revealed regulators of monocytic differentiation, SPI1, JUNB, FOS, and aryl-hydrocarbon receptor signaling (AHR/ARNT), as determinants of BETi response. AHR activation synergized with BETi while inhibition antagonized BETi-mediated cytotoxicity. Consistent with BETi sensitivity dependence on monocytic differentiation,
sensitivity to BETi in primary AML patient samples correlated with higher expression of monocytic markers CSF1R, LILRs, and VCAN. In addition, HL-60 cell line differentiation enhanced its sensitivity to BETi. Further, screens to rescue BETi sensitivity identified BCL2 and CDK6 as druggable vulnerabilities. Finally, monocytic AML patient samples refractory to venetoclax
were significantly more sensitive to combined BETi + venetoclax. Together, our work highlights mechanisms that could predict BETi response and identifies combination strategies to overcome resistance.
Acute myeloid leukemia (AML) remains difficult to treat due to mutational heterogeneity and the development of resistance to therapy. Targeted agents, such as MEK inhibitors, may be incorporated into ...treatment; however, the impact of MEK inhibitors on the immune microenvironment in AML is not well understood. A greater understanding of the implications of MEK inhibition on immune responses may lead to a greater understanding of immune evasion and more rational combinations with immunotherapies. This study describes the impact of trametinib on both T cells and AML blast cells by using an immunosuppressive mouse model of AML and primary patient samples. We also used a large AML database of functional drug screens to understand characteristics of trametinib-sensitive samples. In the mouse model, trametinib increased T-cell viability and restored T-cell proliferation. Importantly, we report greater proliferation in the CD8+CD44+ effector subpopulation and impaired activation of CD8+CD62L+ naive cells. Transcriptome analysis revealed that trametinib-sensitive samples have an inflammatory gene expression profile, and we also observed increased programmed cell death ligand 1 (PD-L1) expression on trametinib-sensitive samples. Finally, we found that trametinib consistently reduced PD-L1 and PD-L2 expression in a dose-dependent manner on the myeloid population. Altogether, our data present greater insight into the impact of trametinib on the immune microenvironment and characteristics of trametinib-sensitive patient samples.
•MEK inhibition rescues T cells from activation-induced cell death in an AML model.•MEK inhibitor sensitivity is associated with inflammation pathways and PD-L1 expression.
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Abstract
Hematopoietic stem cells (HSC) frequently acquire gene mutations with age. An expansion of blood cells derived from a single HSC is termed clonal hematopoiesis (CH), a condition that often ...precedes acute myeloid leukemia (AML). CH is common in elderly individuals: by age 60, approximately 10% of individuals have a clonally expanded blood cell population with a leukemia-associated mutation. The most common mutations in CH (50% of all cases) inactivate the DNA methyltransferase DNMT3A, which regulates gene expression via methylation of CpG rich regions. These mutations provide a selective advantage over healthy HSCs during aging and inflammation. Our goal is to discover mechanisms that allow for clonal expansion of DNMT3A-driven CH. A better understanding of these mechanisms will inspire new therapeutic strategies to limit clonal expansion and increase survival. To evaluate the transcriptional consequences of DNMT3A mutations in myeloid cells, we analyzed our previously published single-cell RNA-sequencing data of DNMT3A-mutated AML patient samples. We found a significant enrichment of antigen presentation signatures in DNMT3A mutated cells along the myeloid differentiation trajectory. CD8+ T cells from these same patients had significantly increased signatures of T cell activation and T cell exhaustion. T cell exhaustion occurs after an activated T cell has been continuously exposed to antigens. Exhausted T cells lack most effector functions and the ability to properly surveil (i.e. recognize mutant peptides presented on the surface of mutated cells and kill the target upon recognition) and restrain mutant cells from expanding. We hypothesize that DNMT3A-driven CH leads to escape of immune surveillance via chronic antigen stimulation and eventual promotion of T cell exhaustion. This would allow for further expansion of the mutated cells and the transformation of CH into AML. To functionally test this hypothesis, we evaluated whether a mouse model of Dnmt3a-CH recapitulated our human data. Indeed, scRNA-seq data of mouse Dnmt3a-CH cells showed significantly increased antigen presentation signatures across the myeloid differentiation trajectory. Additionally, CD8+ T cells had increased signatures of activation. We next functionally tested whether Dnmt3a-mutated CH cells had enhanced antigen presentation by pulsing bone marrow cells derived from Dnmt3a-CH mice with DQ-Ovalbumin, a self-quenched exogenous protein that becomes fluorescent when endocytosed and digested in the phagolysosome. Dnmt3a-mutated HSCs and LSKs had significantly increased DQ-OVA uptake and processing. In conclusion, we found that DNMT3A-CH cells have increased antigen presentation and processing which corresponds with enhanced immunogenicity. These findings support a model in which T cells constrain the expansion of DNMT3A-mutated HSCs, until T cell exhaustion occurs, leading to further expansion of the mutated clone and an increased chance of transformation into AML.
Citation Format: Kyle A Romine, Daniel Ssozi, Tina Mujica, Yoke Lee, Jennifer Trowbridge, Peter van Galen. Mechanisms of adaptive immune evasion by pre-leukemic DNMT3A-mutated blood cell clones abstract. In: Proceedings of the AACR Special Conference: Acute Myeloid Leukemia and Myelodysplastic Syndrome; 2023 Jan 23-25; Austin, TX. Philadelphia (PA): AACR; Blood Cancer Discov 2023;4(3_Suppl):Abstract nr A28.
Abstract
Acute myeloid leukemia (AML) is a hematological cancer with a very poor prognosis. FLT3-ITD mutations that cause constitutive FLT3 signaling are commonly seen in patients. Our lab and others ...have shown that AML patient samples exhibit hallmarks of immune exhaustion such as T cell dysfunction, increased MDSCs, and increased Tregs that associate with worse survival. In solid tumor models DCs have been shown to be important for anti-tumor T cell activity and we aim to further describe the role of DCs in the immune response to leukemia. FLT3 signaling is critical for DC development but the effects of FLT3-ITD on DCs and how it contributes to leukemogenesis remains unclear. In a novel FLT3-ITD driven AML mouse model we found that DCs from these mice have elevated pFLT3, indicating constitutive FLT3 signaling. Moreover, significantly expanded DC progenitors and DCs in the bone marrow and spleen are observed. Analysis of serum cytokines identified increased levels of IL-27, IL-10, and IL-17a in leukemia mice compared to healthy controls, suggesting that the expansion of DCs in these leukemia mice may be driving pathogenic expansion of T helper subsets. Furthermore, we measured higher levels of circulating Th2, Th17, and Treg phenotypes in the blood compared to healthy controls. Thus, we hypothesize that FLT3-ITD DCs are a significant contributor to T cell skewing in AML resulting in poor anti-tumor responses. We are using scRNA-seq methods to identify transcriptional changes in FLT3-ITD DCs that lead to the phenotypes we identified. Understanding how FLT3-ITD DCs contribute to AML immune suppression is critical to interpreting leukemia etiology and the development of targeted therapies such as immune checkpoint blockade or small molecule inhibitors.
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