Akkermansia muciniphila uses human milk oligosaccharides to thrive in the early life conditions in vitro

Akkermansia muciniphila is a well-studied anaerobic bacterium specialized in mucus degradation and associated with human health. Because of the structural resemblance of mucus glycans and free human milk oligosaccharides (HMOs), we studied the ability of A. muciniphila to utilize human milk oligosaccharides. We found that A. muciniphila was able to grow on human milk and degrade HMOs. Analyses of the proteome of A. muciniphila indicated that key-glycan degrading enzymes were expressed when the bacterium was grown on human milk. Our results display the functionality of the key-glycan degrading enzymes (alpha -l-fucosidases, beta -galactosidases, exo-alpha -sialidases and beta -acetylhexosaminidases) to degrade the HMO-structures 2 ' -FL, LNT, lactose, and LNT2. The hydrolysation of the host-derived glycan structures allows A. muciniphila to promote syntrophy with other beneficial bacteria, contributing in that way to a microbial ecological network in the gut. Thus, the capacity of A. muciniphila to utilize human milk will enable its survival in the early life intestine and colonization of the mucosal layer in early life, warranting later life mucosal and metabolic health.Peer reviewe

Epigenetic modifier gene mutations in chronic myeloid leukemia (CML) at diagnosis are associated with risk of relapse upon treatment discontinuation

Non peer reviewe

IFN-α with dasatinib broadens the immune repertoire in patients with chronic-phase chronic myeloid leukemia

In chronic myeloid leukemia (CML), combination therapies with tyrosine kinase inhibitors (TKIs) aim to improve the achievement of deep molecular remission that would allow therapy discontinuation. IFN-alpha is one promising candidate, as it has long-lasting effects on both malignant and immune cells. In connection with a multicenter clinical trial combining dasatinib with IFN-alpha in 40 patients with chronic-phase CML (NordCML007, NCT01725204), we performed immune monitoring with single-cell RNA and T cell receptor (TCR) sequencing (n = 4, 12 samples), bulk TCR beta sequencing (n = 13, 26 samples), flow cytometry (n = 40, 106 samples), cytokine analyses (n = 17, 80 samples), and ex vivo functional studies (n = 39, 80 samples). Dasatinib drove the immune repertoire toward terminally differentiated NK and CD8+ T cells with dampened functional capabilities. Patients with dasatinib-associated pleural effusions had increased numbers of CD8(+) recently activated effector memory T (Temra) cells. In vitro, dasatinib prevented CD3-induced cell death by blocking TCR signaling. The addition of IFN-alpha reversed the terminally differentiated phenotypes and increased the number of costimulatory intercellular interactions and the number of unique putative epitope-specific TCR clusters. In vitro IFN-alpha had costimulatory effects on TCR signaling. Our work supports the combination of IFN-alpha with TKI therapy, as IFN-alpha broadens the immune repertoire and restores immunological function.Peer reviewe

Integrated drug profiling and CRISPR screening identify essential pathways for CAR T-cell cytotoxicity

Funding Information: The authors thank Laura Turunen, Jani Saarela, Katja Suomi, and Maria Nurmi of the High-Throughput Biomedicine Unit and the personnel of the Sequencing Laboratory at the Institute of Molecular Medicine Finland. The authors also thank Caroline Heckman, Sirpa Lepp?, and Olli Lohi for collaboration by providing the cell lines used in the study. The Biomedicum Helsinki Flow Cytometry Core Unit and the Biomedicum Virus Core are acknowledged for their services. The primary B-ALL samples were provided by the Finnish Hematology Registry and Clinical Biobank (FHRB). We thank all the patients for their generous participation. The FHRB biobank is supported by the Finnish Association of Hematology, Finnish Red Cross Blood Service, Institute for Molecular Medicine Finland, and participating hospitals in Finland. This study was supported by the Cancer Foundation Finland, the Sigrid Juselius Foundation, the Relander Foundation, state funding for university-level health research in Finland, and HiLife fellow funds from the University of Helsinki. O.D. was supported by grants from the Biomedicum Helsinki Foundation and the Finnish Medical Society. J. Koski was supported by grants from the Finnish Pediatric Cancer Foundation, the V?re Pediatric Research Foundation, the Cancer Foundation Finland, the Orion Research Foundation, and state funding for university-level health research in Finland. Funding Information: This study was supported by the Cancer Foundation Finland, the Sigrid Juselius Foundation, the Relander Foundation, state funding for university-level health research in Finland, and HiLife fellow funds from the University of Helsinki. O.D. was supported by grants from the Biomedicum Helsinki Foundation and the Finnish Medical Society. J. Koski was supported by grants from the Finnish Pediatric Cancer Foundation, the Väre Pediatric Research Foundation, the Cancer Foundation Finland, the Orion Research Foundation, and state funding for university-level health research in Finland. Funding Information: The authors thank Laura Turunen, Jani Saarela, Katja Suomi, and Maria Nurmi of the High-Throughput Biomedicine Unit and the personnel of the Sequencing Laboratory at the Institute of Molecular Medicine Finland. The authors also thank Caroline Heckman, Sirpa Leppä, and Olli Lohi for collaboration by providing the cell lines used in the study. The Biomedicum Helsinki Flow Cytometry Core Unit and the Biomedicum Virus Core are acknowledged for their services. The primary B-ALL samples were provided by the Finnish Hematology Registry and Clinical Biobank (FHRB). We thank all the patients for their generous participation. The FHRB biobank is supported by the Finnish Association of Hematology, Finnish Red Cross Blood Service, Institute for Molecular Medicine Finland, and participating hospitals in Finland. Publisher Copyright: © 2020 American Society of HematologyChimeric antigen receptor (CAR) T-cell therapy has proven effective in relapsed and refractory B-cell malignancies, but resistance and relapses still occur. Better understanding of mechanisms influencing CAR T-cell cytotoxicity and the potential for modulation using small-molecule drugs could improve current immunotherapies. Here, we systematically investigated druggable mechanisms of CAR T-cell cytotoxicity using >500 small-molecule drugs and genome-scale CRISPR-Cas9 loss-of-function screens. We identified several tyrosine kinase inhibitors that inhibit CAR T-cell cytotoxicity by impairing T-cell signaling transcriptional activity. In contrast, the apoptotic modulator drugs SMAC mimetics sensitized B-cell acute lymphoblastic leukemia and diffuse large B-cell lymphoma cells to anti-CD19 CAR T cells. CRISPR screens identified death receptor signaling through FADD and TNFRSF10B (TRAIL-R2) as a key mediator of CAR T-cell cytotoxicity and elucidated the RIPK1-dependent mechanism of sensitization by SMAC mimetics.Death receptor expression varied across genetic subtypes of B-cell malignancies, suggesting a link between mechanisms of CAR T-cell cytotoxicity and cancer genetics. These results implicate death receptor signaling as an important mediator of cancer cell sensitivity to CAR T-cell cytotoxicity, with potential for pharmacological targeting to enhance cancer immunotherapy. The screening data provide a resource of immunomodulatory properties of cancer drugs and genetic mechanisms influencing CAR T-cell cytotoxicity. Key Points: • Survey of immunomodulatory effects of >500 drugs identifies SMAC mimetics as sensitizers to CAR T-cell cytotoxicity. • Genome-scale CRISPR screen reveals essentiality of death receptor signaling for CAR T-cell cytotoxicity.Peer reviewe

Single-cell analysis of immune recognition in chronic myeloid leukemia patients following tyrosine kinase inhibitor discontinuation

Immunological control of residual leukemia cells is thought to occur in patients with chronic myeloid leukemia (CML) that maintain treatment-free remission (TFR) following tyrosine kinase inhibitor (TKI) discontinuation. To study this, we analyzed 55 single-cell RNA and T cell receptor (TCR) sequenced samples (scRNA+TCRαβ-seq) from patients with CML (n = 13, N = 25), other cancers (n = 28), and healthy (n = 7). The high number and active phenotype of natural killer (NK) cells in CML separated them from healthy and other cancers. Most NK cells in CML belonged to the active CD56dim cluster with high expression of GZMA/B, PRF1, CCL3/4, and IFNG, with interactions with leukemic cells via inhibitory LGALS9–TIM3 and PVR–TIGIT interactions. Accordingly, upregulation of LGALS9 was observed in CML target cells and TIM3 in NK cells when co-cultured together. Additionally, we created a classifier to identify TCRs targeting leukemia-associated antigen PR1 and quantified anti-PR1 T cells in 90 CML and 786 healthy TCRβ-sequenced samples. Anti-PR1 T cells were more prevalent in CML, enriched in bone marrow samples, and enriched in the mature, cytotoxic CD8 + TEMRA cluster, especially in a patient maintaining TFR. Our results highlight the role of NK cells and anti-PR1 T cells in anti-leukemic immune responses in CML.Peer reviewe

Erythroid/megakaryocytic differentiation confers BCL-XL dependency and venetoclax resistance in acute myeloid leukemia

Funding Information: This study was supported by the Cancer Foundation Finland, the Helsinki Institute of Life Science (HiLIFE) Fellow grants, the Academy of Finland (grant 1320185 ), the Sigrid Jusélius Foundation , the Finnish Cancer Institute , the Finnish Medical Foundation , University of Helsinki , and the Finnish special governmental subsidy for health sciences, research, and training . The FIMM High Throughput Biomedicine Unit is financially supported by the University of Helsinki (HiLIFE) and Biocenter Finland. O.D. was supported by the K. Albin Johansson Foundation , the Emil Aaltonen Foundation , and the Juhani Aho foundation . Publisher Copyright: © 2023 The American Society of HematologyMyeloid neoplasms with erythroid or megakaryocytic differentiation include pure erythroid leukemia, myelodysplastic syndrome with erythroid features, and acute megakaryoblastic leukemia (FAB M7) and are characterized by poor prognosis and limited treatment options. Here, we investigate the drug sensitivity landscape of these rare malignancies. We show that acute myeloid leukemia (AML) cells with erythroid or megakaryocytic differentiation depend on the antiapoptotic protein B-cell lymphoma (BCL)-XL, rather than BCL-2, using combined ex vivo drug sensitivity testing, genetic perturbation, and transcriptomic profiling. High-throughput screening of >500 compounds identified the BCL-XL–selective inhibitor A-1331852 and navitoclax as highly effective against erythroid/megakaryoblastic leukemia cell lines. In contrast, these AML subtypes were resistant to the BCL-2 inhibitor venetoclax, which is used clinically in the treatment of AML. Consistently, genome-scale CRISPR-Cas9 and RNAi screening data demonstrated the striking essentiality of BCL-XL-encoding BCL2L1 but not BCL2 or MCL1, for the survival of erythroid/megakaryoblastic leukemia cell lines. Single-cell and bulk transcriptomics of patient samples with erythroid and megakaryoblastic leukemias identified high BCL2L1 expression compared with other subtypes of AML and other hematological malignancies, where BCL2 and MCL1 were more prominent. BCL-XL inhibition effectively killed blasts in samples from patients with AML with erythroid or megakaryocytic differentiation ex vivo and reduced tumor burden in a mouse erythroleukemia xenograft model. Combining the BCL-XL inhibitor with the JAK inhibitor ruxolitinib showed synergistic and durable responses in cell lines. Our results suggest targeting BCL-XL as a potential therapy option in erythroid/megakaryoblastic leukemias and highlight an AML subgroup with potentially reduced sensitivity to venetoclax-based treatments.Peer reviewe

Erythroid/megakaryocytic differentiation confers BCL-XL dependency and venetoclax resistance in acute myeloid leukemia

Myeloid neoplasms with erythroid or megakaryocytic differentiation include pure erythroid leukemia, myelodysplastic syndrome with erythroid features, and acute megakaryoblastic leukemia (FAB M7) and are characterized by poor prognosis and limited treatment options. Here, we investigate the drug sensitivity landscape of these rare malignancies. We show that acute myeloid leukemia (AML) cells with erythroid or megakaryocytic differentiation depend on the antiapoptotic protein B-cell lymphoma (BCL)-XL, rather than BCL-2, using combined ex vivo drug sensitivity testing, genetic perturbation, and transcriptomic profiling. High-throughput screening of >500 com-pounds identified the BCL-XL-selective inhibitor A-1331852 and navitoclax as highly effective against erythroid/megakaryoblastic leukemia cell lines. In contrast, these AML subtypes were resistant to the BCL-2 inhibitor venetoclax, which is used clinically in the treatment of AML. Consistently, genome-scale CRISPR-Cas9 and RNAi screening data demonstrated the striking essentiality of BCL-XL-encoding BCL2L1 but not BCL2 or MCL1, for the survival of erythroid/megakaryoblastic leukemia cell lines. Single-cell and bulk transcriptomics of patient samples with erythroid and megakaryoblastic leukemias iden-tified high BCL2L1 expression compared with other subtypes of AML and other hematological malignancies, where BCL2 and MCL1 were more prominent. BCL-XL inhibition effectively killed blasts in samples from patients with AML with erythroid or megakaryocytic differentiation ex vivo and reduced tumor burden in a mouse erythroleukemia xenograft model. Combining the BCL-XL inhibitor with the JAK inhibitor ruxolitinib showed synergistic and durable responses in cell lines. Our results suggest targeting BCL-XL as a potential therapy option in erythroid/mega-karyoblastic leukemias and highlight an AML subgroup with potentially reduced sensitivity to venetoclax-based treatments.Peer reviewe

Immunogenomic Landscape of Hematological Malignancies

Understanding factors that shape the immune landscape across hematological malignancies is essential for immunotherapy development. We integrated over 8,000 transcriptomes and 2,000 samples with multilevel genomics of hematological cancers to investigate how immunological features are linked to cancer subtypes, genetic and epigenetic alterations, and patient survival, and validated key findings experimentally. Infiltration of cytotoxic lymphocytes was associated with TP53 and myelodysplasia-related changes in acute myeloid leukemia, and activated B cell-like phenotype and interferon-γ response in lymphoma. CIITA methylation regulating antigen presentation, cancer type-specific immune checkpoints, such as VISTA in myeloid malignancies, and variation in cancer antigen expression further contributed to immune heterogeneity and predicted survival. Our study provides a resource linking immunology with cancer subtypes and genomics in hematological malignancies.Peer reviewe