Biology subjects

Heidenreich, O.

Publications and source records attributed to Heidenreich, O..

13 recordsLinked to original sources

bioRxiv2026

Spatial multi-omics reveals targetable immunosuppressive macrophage T-cell interactions in human AML bone marrow

The immunosuppressive bone marrow microenvironment is an important contributor to the limited success of immunotherapy in acute myeloid leukemia (AML), but the cellular interactions underlying AML immune evasion are incompletely understood. We therefore generated a single-cell spatial transcriptomic and proteomic atlas using 148 bone biopsies from 113 individuals comprising pediatric and adult AML at diagnosis and non-leukemic controls. We observed an expansion of regulatory T cells (Tregs) in AML, with stronger colocalization between Tregs and macrophages compared to non-leukemic bone marrow. Distinct cellular neighborhoods were enriched for myeloid progenitor-like cells together with macrophages and T cells, which correlated with higher macrophage and T cell immune checkpoint expression. Moreover, these neighborhoods were associated with specific AML subtypes, especially KMT2A-rearranged and RUNX1::RUNX1T1 AML. These spatial patterns were validated by identification of malignant cells via in situ fusion detection in RUNX1::RUNX1T1 cases. Functional experiments revealed that macrophages and AML cells not only actively recruit Tregs, but also promote naive T cell differentiation into Tregs. Spatially informed ligand-receptor analysis predicted the involvement of the Galectin-9 - CD44/TIM-3 axis in this immunosuppressive crosstalk, which was supported by in vitro inhibition of CD44 and/or TIM-3 preventing macrophage- and AML-induced Treg differentiation. Collectively, this comprehensive spatial map of the AML bone marrow identified tripartite crosstalk between AML, macrophages, and T cells mediated by the Galectin-9 - CD44/TIM-3 axis as a key component of the immunosuppressive microenvironment. Targeting Galectin-9 - CD44/TIM-3 interactions may be a promising strategy to overcome immune evasion and enhance immunotherapeutic success in AML. HighlightsO_LISpatial transcriptomic and proteomic atlas of pediatric and adult acute myeloid leukemia (AML) bone marrow C_LIO_LIIncreased colocalization of macrophages and regulatory T cells (Tregs) in AML C_LIO_LIMacrophages and AML cells induce differentiation of naive T cells to Tregs in vitro, which can be prevented by inhibition of CD44 or TIM-3 C_LI O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=166 SRC="FIGDIR/small/743431v1_ufig1.gif" ALT="Figure 1"> View larger version (47K): org.highwire.dtl.DTLVardef@14f313aorg.highwire.dtl.DTLVardef@290bd3org.highwire.dtl.DTLVardef@c749baorg.highwire.dtl.DTLVardef@1b955d0_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2026

The bone marrow microenvironment of RAS pathway mutant B-ALL is enriched for immunosuppressive regulatory T cells

Somatic mutations in the RAS pathway are highly prevalent in B-Cell Acute Lymphoblastic Leukemia (B-ALL), yet their impact on the bone marrow immune microenvironment and response to immunotherapy remains poorly defined. In this study, we integrated bulk RNA-sequencing, single-cell RNA-sequencing (scRNA-seq), and spectral flow cytometry to characterize the immune landscape of RAS-mutant B-ALL. We identified pathogenic mutations in KRAS, NRAS, PTPN11, or BRAF in 42% of the cohort, predominantly as clonal events. Despite similar T-cell frequencies by flow cytometry, bulk transcriptomes from RAS-mutant samples showed suppression of immune-response and T-cell-activation pathways, and T cells from RAS-mutant patients exhibited impaired proliferation ex vivo. Single-cell analysis revealed higher CD8 dysfunction scores and enrichment of regulatory T cells (Tregs) in RAS-mutant bone marrow. These findings were validated by spectral flow cytometry and by CIBERSORTx deconvolution of bulk data. Trajectory analysis supported a higher CD4 to Treg differentiation in the RAS-mutant niche, and CellChat mapping identified contact-dependent and checkpoint interactions (including TIGIT-NECTIN2 and CTLA-4-CD86/ICOSL) enriched in RAS-mutant samples. Functionally, blinatumomab produced limited leukemic-cell killing ex vivo overall, but addition of CTLA-4 blockade (ipilimumab) selectively restored blinatumomab efficacy in RAS-mutant samples. Together, these results indicate that RAS-pathway activation associates with a Treg-enriched, immunosuppressive bone-marrow microenvironment and point to CTLA-4-targeted strategies to enhance T-cell-engager efficacy in this subgroup.

cancer biology↗

bioRxiv2025

B-cell precursor acute lymphoblastic leukaemia with IGH::CEBP rearrangement: what have we learnt over the years?

B-cell precursor acute lymphoblastic leukaemia (BCP-ALL) is a haematologic malignancy marked by the rapid proliferation of immature B cells in the bone marrow. While BCP-ALL most commonly affects children aged 1-5 years, it remains the most prevalent subtype of ALL in adolescence and adulthood. Chromosomal translocations involving the immunoglobulin (IG) locus and partner genes are proven useful for risk stratification and guiding clinical trials for therapeutic decision. This includes translocations with CCAAT/enhancer-binding proteins (CEBP), which are particularly rare. This rarity has limited efforts to characterise their genetic and clinical profiles, making risk stratification for IGH::CEBP-rearranged BCP-ALL challenging. In this letter, we review the clinical and demographic characteristics of all reported IGH::CEBP cases prior to 2024 and introduce new cases, with preliminary analysis to encourage further investigation into this poorly understood subtype. This study delivers new insights into the molecular and cytogenetic landscape of IGH::CEBP rearrangements in BCP-ALL, and lays a foundation for further investigation into CEBP family roles in haematopoietic development and leukemogenesis, especially in the context of Down syndrome. Finally, it introduces the ongoing international collaborative effort to assemble the largest known IGH::CEBP cohort for comprehensive risk stratification and prognostic evaluation.

cancer biology↗

bioRxiv2025

Bone marrow lymphocyte dynamics during chemotherapy in pediatric acute myeloid leukemia

A better understanding of lymphocyte dynamics during current treatment regimens in pediatric AML is urgently needed to understand whether the application of bispecific T-cell-engagers (BiTEs) during periods of low tumor burden could be a viable treatment strategy. In this study, we found that induction 1, comprising mitoxantrone-etoposide-cytarabine in nearly all patients (as part of the NOPHO-DBH AML-2012 protocol), led to preserved or increased relative lymphocyte abundances alongside marked blast reduction in most cases. This was accompanied by a shift towards higher T-cell fractions, potentially creating a favorable window for BiTE therapy. The absence of a correlation between blast reduction and lymphocyte changes suggests that chemotherapy exerts differential effects on the lymphocyte compartment. Despite the heterogeneity of agents used in induction 2, more than half of patients showed a decline in lymphocyte levels. Nonetheless, the increase in T- and B-cells observed in most patients from the NOPHO-AML 2004 cohort after induction 2 suggests that lymphocyte recovery at this treatment stage is not uniformly impaired. Our transcriptomic and ex vivo functional data align with preclinical findings in adult AML and provide a basis for further investigations in in vivo models and early clinical trials. Such efforts should prioritize novel BiTE constructs targeting multiple tumor-associated (e.g., NCT05673057) or tumor-specific antigens.

cancer biology↗

bioRxiv2025

RUNX1::RUNX1T1 Depletion Eliminates Stemness and Induces Bidirectional Differentiation of Acute Myeloid Leukemia

Chromosomal rearrangements that generate novel fusion genes are a hallmark of acute myeloid leukemia (AML). Depletion experiments in cell line models have suggested that continued expression is required for maintaining their leukemic phenotype and they therefore represent ideal cancer-specific therapeutic targets. However, to which extent this result holds true for the different stages of hematopoietic development in primary cells and whether therapeutic agents can be efficiently delivered to those cells is still unclear. In this study, we demonstrate that primary AML cells harboring the chromosomal translocation t(8;21) are critically dependent on the corresponding fusion gene, RUNX1::RUNX1T1, to suppress differentiation and maintain stemness. Silencing RUNX1::RUNX1T1 expression using siRNA-loaded lipid nanoparticles induces substantial changes in chromatin accessibility, thereby redirecting the leukemia-associated transcriptional network towards a myeloid differentiation program. Single-cell analyses reveal that this transcriptional reprogramming is associated with the depletion of immature stem and progenitor-like cell populations, accompanied by an expansion of granulocytic and eosinophilic/mast cell-like populations with impaired self-renewal capacity. These findings underscore the essential role of RUNX1::RUNX1T1 in sustaining AML and highlight the therapeutic potential of targeting fusion gene expression in primary AML cells. O_FIG O_LINKSMALLFIG WIDTH=167 HEIGHT=200 SRC="FIGDIR/small/637937v1_ufig1.gif" ALT="Figure 1"> View larger version (53K): org.highwire.dtl.DTLVardef@cf4838org.highwire.dtl.DTLVardef@8e6d90org.highwire.dtl.DTLVardef@bb46c5org.highwire.dtl.DTLVardef@12357e4_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2024

Distinct Stromal Cell Populations Define the B-cell Acute Lymphoblastic Leukemia Microenvironment

The bone marrow microenvironment plays a critical role in B-cell acute lymphoblastic leukemia (B-ALL) progression, yet its cellular heterogeneity remains poorly understood. Using single-cell RNA sequencing on patient-derived of bone marrow aspirates from pediatric B-ALL patients, we identified two distinct mesenchymal stromal cell (MSC) populations: early mesenchymal progenitors and adipogenic progenitors. Spatial transcriptomic analysis further revealed the localization of these cell types and identified a third stromal population, osteogenic-lineage cells, exclusively present in the bone biopsy. Functional ex vivo assays using sorted stromal populations derived from B-ALL patient bone marrow aspirates demonstrated that both early mesenchymal and adipogenic progenitors secrete key niche-supportive factors, including CXCL12 and Osteopontin, and support leukemic cell survival and chemoresistance. Transcriptomic profiling revealed that B-ALL cells interact differently with stromal subtypes. Notably, adipogenic progenitors, but not early mesenchymal progenitors, provide support to leukemic cells through interleukin-7 and VCAM1 signaling. Stromal cells from B-ALL patients exhibited an enhanced adipogenic differentiation capacity compared to healthy controls. Moreover, co-culture experiments showed that B-ALL cells induce adipogenic differentiation in healthy MSCs through a cell contact-dependent mechanism. Adipogenic progenitors were also enriched in relapse samples, implicating them in disease progression. These findings highlight the complexity of the B-ALL microenvironment and identify different specialized stromal niches with which the leukemic cells can engage.

cancer biology↗

bioRxiv2024

Repurposing CD19-directed immunotherapies for pediatric t(8;21) acute myeloid leukemia

In contrast to patients with B cell precursor acute lymphoblastic leukemia (BCP-ALL), patients with acute myeloid leukemia (AML) have not yet benefited from recent advances in targeted immunotherapy. Repurposing immunotherapies that have been successfully used to target other hematological malignancies could, in case of a shared target antigen, represent a promising opportunity to expand the immunotherapeutic options for AML. Here, we evaluated the expression of CD19 in a large pediatric AML cohort, assessed the ex vivo AML killing efficacy of CD19-directed immunotherapies, and characterized the bone marrow immune microenvironment in pediatric AML, BCP-ALL, and non-leukemic controls. Out of 167 newly diagnosed de novo pediatric AML patients, 18 patients (11%) had CD19+ AML, with 61% carrying the translocation t(8;21)(q22;q22). Among CD19+ samples, we observed a continuum of CD19 expression levels on AML cells. In individuals exhibiting unimodal and high CD19 expression, the antigen was consistently present on nearly all CD34+CD38- and CD34+CD38+ subpopulations. In ex vivo AML-T cell co-cultures, blinatumomab demonstrated substantial AML killing, with an efficacy similar to BCP-ALL. In addition, CAR T cells could effectively eliminate CD19+ AML cells ex vivo. Furthermore, our immunogenomic assessment of the bone marrow immune microenvironment of newly diagnosed pediatric t(8;21) AML revealed that T- and NK cells had a less exhausted and senescent phenotype in comparison to pediatric BCP-ALL. Altogether, our study underscores the promise of CD19-directed immunotherapies for the treatment of pediatric CD19+ AML.

immunology↗

bioRxiv2023

The neuronal homeobox transcription factor HMX3 is a crucial vulnerability factor in MECOM-negative KMT2A::MLLT3 acute myelomonocytic leukemia

The KMT2A::MLLT3 fusion protein causes acute myeloid leukemia (AML) by activating the oncogenic transcription factor MECOM. However, MECOM expression occurs in only half of the KMT2A::MLLT3 cases. By integrating gene expression and enhancer activity data from patient cells, we identified neuronal homeobox transcription factor HMX3 as cell fate determining factor in MECOM-negative KMT2A::MLLT3 AML. HMX3 expression associated with younger age and KMT2A-rearranged leukemia in large AML cohorts (p<0.002). HMX3 was not expressed in other major genetic risk groups and healthy blood cells. Transcriptomic analyses revealed that HMX3 drives cancer-associated E2F, MYC and cell cycle gene programs. Ectopic HMX3 expression completely inhibited monocytic but not granulocytic colony formation of healthy CD34+ adult cells. Silencing of HMX3 in KMT2A::MLLT3 AML cell lines and patient cells resulted in cell cycle arrest, monocytic differentiation, and apoptosis. Thus, HMX3 is a leukemia-specific vulnerability that enhances proliferation and blocks differentiation of MECOM-negative KMT2A::MLLT3 leukemia.

cancer biology↗

bioRxiv2023

Leukemic stem cells hijack lineage inappropriate signalling pathways to promote their growth

Acute Myeloid Leukemia (AML) is caused by multiple mutations which dysregulate growth and differentiation of myeloid cells. Cells adopt different gene regulatory networks specific to individual mutations, maintaining a rapidly proliferating blast cell population with fatal consequences for the patient if not treated. The most common treatment option is still chemotherapy which targets such cells. However, patients harbour a population of quiescent leukemic stem cells (LSCs) which can emerge from quiescence to trigger relapse after therapy. The processes that allow such cells to re- grow remain unknown. Here, we examined the well characterised t(8;21) AML sub-type as a model to address this question. Using a novel t(8;21) patient-derived xenograft model, we show that t(8;21) LSCs aberrantly activate the VEGF and IL-5 signalling pathways. Both pathways operate within a regulatory circuit consisting of the driver oncoprotein RUNX1::ETO and an AP-1/GATA2 axis allowing LSCs to re-enter the cell cycle while preserving self-renewal capacity.

cancer biology↗

bioRxiv2022

Nanoparticle-mediated Targeting of the Fusion Gene RUNX1/ETO in t(8;21)-positive Acute Myeloid Leukaemia

A hallmark of acute myeloid leukaemias (AMLs) are chromosomal rearrangements that give rise to novel leukaemia-specific fusion genes. Most of these fusion genes are both initiating and driving events in AML and therefore constitute ideal therapeutic targets but are challenging to target by conventional drug development. siRNAs are frequently used for the specific suppression of fusion gene expression but require special formulations for efficient in vivo delivery. Here we describe the use of siRNA-loaded lipid nanoparticles for the specific therapeutic targeting of the leukaemic fusion gene RUNX1/ETO. Transient knockdown of RUNX1/ETO reduces its binding to its target genes and alters the binding of RUNX1 and its co-factor CBF{beta}. Transcriptomic changes in vivo were associated with substantially increased median survival of a t(8;21)-AML mouse model. Importantly, transient knockdown in vivo causes long-lasting inhibition of leukaemic proliferation and clonogenicity, induction of myeloid differentiation and a markedly impaired re-engraftment potential in vivo. These data strongly suggest that temporary inhibition of RUNX1/ETO results in long-term restriction of leukaemic self-renewal. Our results provide proof for the feasibility of targeting RUNX1/ETO in a pre-clinical setting and support the further development of siRNA-LNPs for the treatment of fusion gene-driven malignancies.

cancer biology↗

bioRxiv2021

Single cell mRNA signals reveal a distinct developmental state of KMT2A-rearranged infant B-cell acute lymphoblastic leukemia

Infant B-cell acute lymphoblastic leukemia (B-ALL) has not followed the increasing trend towards cure seen in other childhood B-ALLs. The prognosis for infants with KMT2A gene fusions is especially poor, and the origins of this aggressive leukemia remain unknown. Here, we investigated the developmental state of KMT2A-rearranged infant B-ALL within hematopoietic hierarchies of human fetal bone marrow, using bulk mRNA meta-analysis of childhood leukemia and examination of single lymphoblast transcriptomes. KMT2A-rearranged infant B-ALL was uniquely dominated by an early lymphocyte precursor (ELP) state. Direct comparison of infant lymphoblasts with ELP cells distilled the core oncogenic transcriptome of cancer cells which harboured potentially targetable hybrid myeloid-lymphoid features. Overall our quantitative molecular analyses demonstrate a distinct developmental state of KMT2A-rearranged infant B-ALL.

genomics↗

bioRxiv2021

A human iPSC derived bone marrow milieu identifies a novel target against niche-conferred leukaemia proliferation and treatment resistance

Leukaemia cells re-program their microenvironment to provide proliferation support and protection from standard chemotherapy, molecularly targeted therapies, and immunotherapy. Although much is becoming known about molecules that drive niche-dependent treatment resistance; means of targeting these in the clinics has remained a key obstacle. To address this challenge, we have developed human induced pluripotent stem cell engineered niches ex vivo to reveal insights into druggable cancer-niche dependencies. We show that mesenchymal (iMSC) and vascular niche-like (iANG) cells support ex vivo proliferation of patient-derived leukaemia cells, impact dormancy and mediate therapy resistance. iMSC protected both non-cycling and cycling blasts against dexamethasone treatment while iANG protected only dormant blasts. Leukaemia proliferation and protection from dexamethasone induced-apoptosis was dependent on direct cell-cell contact and mediated by CDH2. To explore the therapeutic potential of disrupting this cell-cell interaction, we tested the CDH2 antagonist ADH-1 (previously in phase I / II for solid tumours) in a very aggressive patient-derived xenograft leukaemia mouse model. ADH-1 showed high in vivo efficacy. ADH-1/ dexamethasone combination therapy was superior to dexamethasone alone with no ADH1 conferred additional toxicity. These findings provide a proof-of-concept starting point to develop novel, potentially safer therapeutics that target niche-mediated cancer cell dependencies in haematological malignancies. SummaryCDH2 mediated niche-dependent cancer proliferation and treatment resistance is clinically targetable via ADH-1, a low toxic agent that could be potentially repurposed for future clinical trials in acute leukaemia.

cancer biology↗

bioRxiv2019

RUNX1/RUNX1T1 controls alternative splicing in the t(8;21)-positive acute myeloid leukemia cells

The fusion oncogene RUNX1/RUNX1T1 encodes an aberrant transcription factor, which plays a key role in the initiation and maintenance of the t(8;21)-positive acute myeloid leukemia. Here we show that this oncogene is a regulator of the alternative RNA splicing for a sub-set of genes in the leukemia cells. We found two primary mechanisms underlying changes in the production of RNA isoforms: (i) RUNX1/RUNX1T1-mediated regulation of alternative transcription start sites selection in target genes, and (ii) direct or indirect control of the expression of the genes encoding splicing factors. The first mechanism leads to the expression of RNA isoforms with alternative structure of the 5-UTR regions. The second mechanism generates alternative transcripts with new junctions between internal cassettes and constitutive exons. We also show that RUNX1/RUNX1T1-mediated differential splicing affects several functional groups of genes and produces proteins with unique conserved domain structures. In summary, this study reveals a novel layer of RUNX1/RUNX1T1-dependent transcriptome organization in t(8;21)-positive acute myeloid leukemia.

cancer biology↗