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Language：English   Publisher：Cancer Science  

In this study, we investigated the measurable residual leukemic stem cell (MR-LSC) population after allogeneic stem cell transplantation (allo-SCT) for high-risk acute myeloid leukemia (AML), utilizing T-cell immunoglobulin mucin-3 (TIM-3) expression as a functional marker of AML leukemic stem cells (LSCs). Analysis of the CD34⁺CD38⁻ fraction of bone marrow cells immediately after achievement of engraftment revealed the presence of both TIM-3⁺LSCs and TIM-3⁻ donor hematopoietic stem cells (HSCs) at varying ratios. Genetic analysis confirmed that TIM-3⁺ cells harbored patient-specific mutations identical to those found in AML clones, whereas TIM-3⁻ cells did not, indicating that TIM-3⁺CD34⁺CD38⁻ cells represent residual AML LSCs. In 92 allo-SCT occasions involving 83 AML patients, we enumerated the frequencies of TIM-3⁺LSCs immediately after achieving hematologic complete remission with complete donor cell chimerism. Notably, only 22.2% of patients who achieved a TIM-3⁺MR-LSC^low status (<60%) experienced relapse, with a median event-free survival (EFS) of 1581 days (median follow-up duration was 2177 days among event-free survivors). Conversely, 87.5% of patients with TIM-3⁺MR-LSC^int/high (≥60%) relapsed, with a median EFS of 140.5 days. Furthermore, MR-LSC status emerged as a significant independent risk factor for relapse (hazard ratio, 8.56; p < 0.0001), surpassing the impact of patient disease status prior to allo-SCT, including failure to achieve complete remission (hazard ratio, 1.98; p = 0.048). These findings suggest that evaluating TIM-3⁺ MR-LSCs immediately after engraftment, which reflects the competitive reconstitution of residual TIM-3⁺ LSCs and donor HSCs, may be valuable for predicting outcomes in AML patients undergoing allo-SCT.

DOI： 10.1111/cas.16431

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DOI： 10.1182/blood-2024-199102

Web of Science

Language：English   Publisher：Blood Advances  

Immunomodulatory drugs (IMiDs) are key drugs for treating multiple myeloma and myelodysplastic syndrome with chromosome 5q deletion. IMiDs exert their pleiotropic effects through the interaction between cell-specific substrates and cereblon, a substrate receptor of the E3 ubiquitin ligase complex. Thus, identification of cell-specific substrates is important for understanding the effects of IMiDs. IMiDs increase the risk of thromboembolism, which sometimes results in fatal clinical outcomes. In this study, we sought to clarify the molecular mechanisms underlying IMiDs-induced thrombosis. We investigated cereblon substrates in human megakaryocytes using liquid chromatography–mass spectrometry and found that thrombospondin-1 (THBS-1), which is an inhibitor of a disintegrin-like and metalloproteinase with thrombospondin type 1 motifs 13, functions as an endogenous substrate in human megakaryocytes. IMiDs inhibited the proteasomal degradation of THBS-1 by impairing the recruitment of cereblon to THBS-1, leading to aberrant accumulation of THBS-1. We observed a significant increase in THBS-1 in peripheral blood mononuclear cells as well as larger von Willebrand factor multimers in the plasma of patients with myeloma, who were treated with IMiDs. These results collectively suggest that THBS-1 represents an endogenous substrate of cereblon. This pairing is disrupted by IMiDs, and the aberrant accumulation of THBS-1 plays an important role in the pathogenesis of IMiDs-induced thromboembolism.

DOI： 10.1182/bloodadvances.2023010080

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Language：English   Publisher：John Wiley & Sons Australia, Ltd  

急性骨髄性白血病(AML)に特異的なミトコンドリア機能の制御に関与する分子メカニズムを検討した。CD34陽性AML細胞と健常な造血幹/前駆細胞を比較した代謝物スクリーニングを通じて、AMLにおけるリゾホスファチジン酸(LPA)合成活性の亢進を同定した。LPAは、LPA合成経路の律速酵素であるグリセロール-3-リン酸アシルトランスフェラーゼ(GPAT)によってグリセロール-3-リン酸から合成される。GPATの4つのアイソザイムのうち、グリセロール-3-リン酸アシルトランスフェラーゼ、ミトコンドリア(GPAM)がAML細胞で高発現していた。GPAMサイレンシングまたはFSG67(GPAM阻害剤)でLPA合成を阻害すると、ミトコンドリア分裂が誘導され、その結果、酸化的リン酸化が抑制され、活性酸素種が増加することで、AMLの増殖が著しく阻害された。さらに、FSG67によるGPAM阻害は、in vivoでの正常な造血に影響を与えることなく、AMLに対して有効であることが示された。以上より、GPAMを介したグリセロール-3-リン酸からのLPA合成経路は、AMLにおけるミトコンドリア動態を特異的に制御する重要な代謝機構であり、GPAMは有望な治療標的であることが示唆された。

Language：English   Publisher：Cancer Science  

Metabolic alterations, especially in the mitochondria, play important roles in several kinds of cancers, including acute myeloid leukemia (AML). However, AML-specific molecular mechanisms that regulate mitochondrial dynamics remain elusive. Through the metabolite screening comparing CD34+ AML cells and healthy hematopoietic stem/progenitor cells, we identified enhanced lysophosphatidic acid (LPA) synthesis activity in AML. LPA is synthesized from glycerol-3-phosphate by glycerol-3-phosphate acyltransferases (GPATs), rate-limiting enzymes of the LPA synthesis pathway. Among the four isozymes of GPATs, glycerol-3-phosphate acyltransferases, mitochondrial (GPAM) was highly expressed in AML cells, and the inhibition of LPA synthesis by silencing GPAM or FSG67 (a GPAM-inhibitor) significantly impaired AML propagation through the induction of mitochondrial fission, resulting in the suppression of oxidative phosphorylation and the elevation of reactive oxygen species. Notably, inhibition of this metabolic synthesis pathway by FSG67 administration did not affect normal human hematopoiesis in vivo. Therefore, the GPAM-mediated LPA synthesis pathway from G3P represents a critical metabolic mechanism that specifically regulates mitochondrial dynamics in human AML, and GPAM is a promising potential therapeutic target.

DOI： 10.1111/cas.15835

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Language：English   Publisher：Blood Advances  

Cancer-specific metabolic activities play a crucial role in the pathogenesis of human malignancies. To investigate human acute leukemia–specific metabolic properties, we comprehensively measured the cellular metabolites within the CD34+ fraction of normal hematopoietic stem progenitor cells (HSPCs), primary human acute myelogenous leukemia (AML), and acute lymphoblastic leukemia (ALL) cells. Here, we show that human leukemia cells are addicted to the branched-chain amino acid (BCAA) metabolism to maintain their stemness, irrespective of myeloid or lymphoid types. Human primary acute leukemias had BCAA transporters for BCAA uptake, cellular BCAA, α-ketoglutarate (α-KG), and cytoplasmic BCAA transaminase-1 (BCAT1) at significantly higher levels than control HSPCs. Isotope-tracing experiments showed that in primary leukemia cells, BCAT1 actively catabolizes BCAA using α-KG into branched-chain α-ketoacids, whose metabolic processes provide leukemia cells with critical substrates for the trichloroacetic acid cycle and the synthesis of nonessential amino acids, both of which reproduce α-KG to maintain its cellular level. In xenogeneic transplantation experiments, deprivation of BCAA from daily diet strongly inhibited expansion, engraftment and self-renewal of human acute leukemia cells. Inhibition of BCAA catabolism in primary AML or ALL cells specifically inactivates the function of the polycomb repressive complex 2, an epigenetic regulator for stem cell signatures, by inhibiting the transcription of PRC components, such as zeste homolog 2 and embryonic ectoderm development. Accordingly, BCAA catabolism plays an important role in the maintenance of stemness in primary human AML and ALL, and molecules related to the BCAA metabolism pathway should be critical targets for acute leukemia treatment.

DOI： 10.1182/bloodadvances.2022008242

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Language：English   Publisher：Blood Advances  

The activation of β-catenin plays critical roles in normal stem cell function, and, when aberrantly activated, the maintenance and enhancement of cancer stemness in many solid cancers. Aberrant β-catenin activation is also observed in acute myeloid leukemia (AML), and crucially contributes to self-renewal and propagation of leukemic stem cells (LSCs) regardless of mutations in contrast with such solid tumors. In this study, we showed that the AML-specific autocrine loop comprised of T-cell immunoglobulin mucin-3 (TIM-3) and its ligand, galectin-9 (Gal-9), drives the canonical Wnt pathway to stimulate self-renewal and propagation of LSCs, independent of Wnt ligands. Gal-9 ligation activates the cytoplasmic Src homology 2 domain of TIM-3 to recruit hematopoietic cell kinase (HCK), a Src family kinase highly expressed in LSCs but not in HSCs, and HCK phosphorylates p120-catenin to promote formation of the LDL receptor–related protein 6 (LRP6) signalosome, hijacking the canonical Wnt pathway. This TIM-3/HCK/p120-catenin axis is principally active in immature LSCs compared with TIM-3–expressed differentiated AML blasts and exhausted T cells. These data suggest that human AML LSCs constitutively activates β-catenin via autocrine TIM-3/HCK/p120-catenin signaling, and that molecules related to this signaling axis should be critical targets for selective eradication of LSCs without impairing normal HSCs.

DOI： 10.1182/bloodadvances.2022008405

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Language：English   Publishing type：Research paper (scientific journal)  

The activation of β-catenin plays critical roles in normal stem cell function, and, when aberrantly activated, the maintenance and enhancement of cancer stemness in many solid cancers. Aberrant β-catenin activation is also observed in acute myeloid leukemia (AML), and crucially contributes to self-renewal and propagation of leukemic stem cells (LSCs) regardless of mutations in contrast with such solid tumors. In this study, we showed that the AML-specific autocrine loop comprised of T-cell immunoglobulin mucin-3 (TIM-3) and its ligand, galectin-9 (Gal-9), drives the canonical Wnt pathway to stimulate self-renewal and propagation of LSCs, independent of Wnt ligands. Gal-9 ligation activates the cytoplasmic Src homology 2 domain of TIM-3 to recruit hematopoietic cell kinase (HCK), a Src family kinase highly expressed in LSCs but not in HSCs, and HCK phosphorylates p120-catenin to promote formation of the LDL receptor-related protein 6 (LRP6) signalosome, hijacking the canonical Wnt pathway. This TIM-3/HCK/p120-catenin axis is principally active in immature LSCs compared with TIM-3-expressed differentiated AML blasts and exhausted T cells. These data suggest that human AML LSCs constitutively activates β-catenin via autocrine TIM-3/HCK/p120-catenin signaling, and that molecules related to this signaling axis should be critical targets for selective eradication of LSCs without impairing normal HSCs.

DOI： 10.1182/bloodadvances.2022008405.

Language：Japanese   Publisher：Japanese Society for Transplantation and Cellular Therapy  

<p> The relapse of acute myeloid leukemia is a critical problem in clinics. As the prognosis of patients who relapse after allogeneic stem cell transplantation (allo-SCT) is poor, the evaluation of minimal/measurable residual disease (MRD) is crucial for determining the risk of relapse. Multiparameter flow cytometry (MFC) is an established method for quantifying MRD by European Leukemia Net. One of our primary concerns about MFC-MRD is its genetic backbone. In addition to MFC, detecting genetic abnormalities with next-generation sequencing (NGS) is a useful method for quantifying MRD. MFC-MRD and NGS-MRD are complementary approaches for predicting relapse, but conflicting results are observed in some cases. To determine the relevance of MFC-MRD and NGS-MRD, we performed single-cell targeted DNA and surface protein analysis using the Tapestri platform. Then, we established a heterogeneity characterized by the expression pattern of the surface antigen where populations with similar genetic background could produce different MRD statuses between MFC-MRD and NGS-MRD. Therefore, to track the residual disease responsible for relapse, we focused on T-cell immunoglobulin mucin-3 (TIM-3), a leukemic stem cell (LSC)-specific functional molecule. Furthermore, we demonstrated that the evaluation of TIM-3⁺ LSCs could be useful in predicting relapses after allo-SCT.</p>

DOI： 10.7889/tct-23-005

CiNii Research

Language：Japanese   Publisher：The Japanese Society of Hematology  

<p>Acute myeloid leukemia (AML) is one of the most common hematologic malignancies derived from self-renewing and highly propagating leukemic stem cells (LSCs). We have previously identified T-cell immunoglobulin mucin-3 (TIM-3) as an AML LSC-specific surface molecule by comparing the gene expression profiles of LSCs and hematopoietic stem cells (HSCs). TIM-3 expression clearly discriminates LSCs from HSCs within the CD34⁺CD38^- stem cell fraction. Furthermore, AML cells secrete galectin-9 (Gal-9, a TIM-3 ligand) in an autocrine manner, resulting in constitutive TIM-3 signaling, which maintains LSC self-renewal capacity through <i>β</i>-catenin accumulation. In this study, we investigated the LSC-specific mechanisms of TIM-3 signaling. We found that TIM-3 signaling drove the canonical Wnt pathway, which was independent of Wnt ligands, to maintain cancer stemness in LSCs. Gal-9 ligation activated the cytoplasmic Src homology 2 (SH2) binding domain of TIM-3 to recruit hematopoietic cell kinase (HCK), a Src family kinase that is highly expressed in LSCs. HCK phosphorylated p120-catenin to promote the formation of the LDL receptor-related protein 6 (LRP6) signalosome, hijacking the canonical Wnt pathway. This TIM-3/HCK/p120-catenin axis was employed principally in immature LSCs compared to TIM-3-expressing exhausted T-cells.</p>

DOI： 10.11406/rinketsu.64.547

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CiNii Research

Language：English   Publisher：American Journal of Ophthalmology Case Reports  

Purpose: This article presents a case of panuveitis that occurred after unrelated allogeneic hematopoietic stem cell transplantation (allo-HSCT) in a patient with lymphoma-type human T-cell leukemia virus type-1 (HTLV-1)-associated adult T-cell leukemia (ATL). Observations: A 45-year-old man developed unilateral panuveitis 18 months after undergoing allo-HSCT. He underwent vitrectomy, and depositions of grey-white granules localized on the retinal artery were observed in the eye. Cytological examination of the vitreous aspirates showed that the atypical lymphoid cells stained positive for CD3 and CD8, but negative for CD4, B-cell markers, and cytomegalovirus antigen. Interphase fluorescence in situ hybridization using X‐ and Y‐chromosome probes revealed complete donor chimerism in CD8+ T cells in the vitreous aspirates. Conclusions and importance: Donor-derived CD8+ T lymphocytes can induce panuveitis like HTLV-1-assiciated uveitis after allo-HSCT in patients with ATL. Pathological diagnosis of vitreous infiltration by vitrectomy is helpful in patients with ATL. Donor-derived CD8+ T lymphocytes-induced panuveitis is recurrent but susceptible to regional corticosteroid treatment.

DOI： 10.1016/j.ajoc.2022.101673

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