Age-related clonal hematopoiesis and myeloid malignancies arise from hematopoietic stem cells and progenitors with genetic abnormalities. Advances in next-generation sequencing technology have led to the identification of a wide variety of genetic alterations involved in disease onset. However, it remains unclear how diverse genetic alterations, lacking disease specificity, lead to the development of myeloid malignancies and the progression of clonal hematopoiesis. Mitochondrial abnormalities and their roles in various pathological conditions such as aging, inflammation, neurological diseases, cardiac diseases, and cancer have recently been revealed, and have garnered attention as new therapeutic targets. This review focuses on regulation of mitochondrial dynamics and outlines the role of mitochondria in myeloid malignancies and clonal hematopoiesis.

Hematopoietic stem cell (HSC)-targeted gene therapy is curative for various genetic blood diseases, and its efficacy has been demonstrated in recent clinical trials. HSCs have self-renewal and hematopoietic multipotency; therefore, repairing pathological mutations or defects in HSCs allows for a lifelong cure with a single treatment. Autologous HSC gene therapy has been developed by lentiviral gene addition or gene editing, and is an option for most patients because it does not require a compatible donor. Current HSC gene therapy is based on ex vivo methods, in which patient HSCs are harvested, genetically modified ex vivo, and autologously transplanted into patients. However, the complexity of this process and the high cost of treatment are hindering the spread of gene therapy. Therefore, in vivo HSC gene therapy is being developed to deliver gene therapy tools directly into bone marrow HSCs by administration without ex vivo culture.

Methods in which patient-derived T cells are genetically modified in vitro and administered to patients have been demonstrated effective in the area of cancer immunotherapy. However, these methods have some unresolved issues such as cost, time, and unstable quality. Several groups have developed strategies to overcome these barriers by regenerating T cells from iPSCs. We have been developing a method in which specific TCR genes are introduced into iPSCs and T cells are regenerated from these iPSCs (TCR-iPSC method). We are now using starting iPSCs from the iPSC stock lines provided by CiRA-F, as the iPSC stock cells are less likely to be rejected. A study aimed at application to solid tumors demonstrated the therapeutic effect of regenerated T cells in a patient tissue xenograft model of WT1 antigen-positive renal cell carcinoma. This article will also discuss strategies by other groups to regenerate various types of T cells from iPSCs.

Prediction of intestinal drug absorption and drug-induced intestinal toxicity is critical for the development of orally-administered drugs. However, it is difficult to accurately predict these events because of large species differences and a lack of appropriate in vitro assay. Then, we proposed the use of human crypt-derived intestinal cells for the prediction of intestinal absorption and the risk of intestinal toxicity. 3D human intestinal spheroids were established from fresh surgical specimens of proximal jejunum and terminal ileum using the conditioned media containing Wnt3a, R-spondin 3, and noggin. To generate 2D monolayer, spheroids were enzymatically dissociated into single cells and plated onto Matrigel-precoated culture plates/inserts. We have confirmed the activities of typical drug-metabolizing enzymes and uptake/efflux transporters in human jejunal spheroid-derived differentiated cells. Intestinal availability (Fg) estimated from the apical-to-basal permeation clearance across the jejunal monolayer showed a good correlation with in vivo human Fg values for five CYP3A substrate drugs. As for the prediction of intestinal toxicity, we found that the degree of ATP decreases in intestinal spheroids incubated with different EGFR-TKIs varied greatly depending on the drugs and the rank order of the extent of ATP decrease corresponded with that of frequency of clinically-observed diarrhea. We also constructed enterochromaffin (EC) cell-rich spheroids and quantified serotonin release from EC cells upon exposure to drugs for the prediction of drug-induced nausea and vomiting. As a result, we found that the serotonin release was related to the high/low risk of nausea and vomiting of each ALK/ROS1 kinase inhibitors.

Drug discovery research takes many years and tons of effort, and the success rate is extremely low. In order to overcome this situation, pharmaceutical companies struggle to improve the probability of success drug research and development with multiple approaches. Recently, it is important to predict the clinical effects of the candidates as early as possible in drug discovery stage, to stratify patients with diseases, and to provide appropriate readouts for evaluation of pharmacological efficacy for increasing the success rate. In this environment, the importance of non-clinical research that actively utilizes human-derived samples including patient-derived samples is increasing. In this article, author describes the use of human-derived samples in non-clinical research, especially focusing on the utilization of induced pluripotent stem cells. Human-derived samples are valuable experimental materials that have, at least in part, human-specific characteristics that experimental animals do not possess. In particular, patient-derived samples are thought to have the genetic predisposition and at least some disease characteristics that cause the disease, and are useful from the perspective of elucidating the pathogenesis, disease modeling, and predictability of clinical effects. This is also valuable for drug discovery research in diseases that are difficult to reproduce in experimental animals such as mice. Whereas, human-derived samples have some limitations, and we need ethical procedures and consideration when researchers use them. Author will provide an overview of the use of human-derived samples in non-clinical research based on the perspectives as described above and introduce our research group cases, and future research prospects using them.

Myelodysplastic syndrome (MDS) is a refractory cancer that arises from hematopoietic stem cells and predominantly affects elderly adults. In addition to driver gene mutations, which are also found in clonal hematopoiesis in healthy elderly people, systemic inflammation caused by infection or collagen disease has long been known as an extracellular factor in the pathogenesis of MDS. Wild-type HSCs have an "innate immune memory" that functions in response to infection and inflammatory stress, and my colleagues and I used an infection stress model to demonstrate that the innate immune response by the TLR-TRIF-PLK-ELF1 pathway is similarly critical in impairment of hematopoiesis and dysregulation of chromatin in MDS stem cells. This revealed that not only are MDS stem cells expanded by the TRAF6-NF-kB pathway, the innate immune response is also involved in generating MDS stem cells. In this review, I will present research findings related to "innate immune memory," one of the pathogenic mechanisms of blood cancer, and discuss future directions for basic pathological research and potential therapeutic development.

Chimeric antigen receptor-transduced autologous T (CAR-T) cell therapy targeting CD19 has revolutionized the treatment of CD19-positive hematological tumors, including acute lymphoblastic leukemia and large B-cell lymphoma. However, despite the high response rate, many problems such as exceedingly high cost, complex logistics, insufficient speed, and manufacturing failures have become apparent. One solution for these problems is to use an allogeneic cell as an effector cell for genetic modification with CAR. Allogeneic, or "off-the-shelf", CAR-expressing immune-effector cells include 1) genome-edited, T-cell receptor (TCR) gene-deleted CAR-T cells generated using healthy adult donor T cells, 2) induced pluripotent stem cell-derived CAR-T cells, and 3) CAR NK cells. NK cells are notorious for their poor ex-vivo expansion and low susceptibility to genetic modification. In this article, I will review the current state and future prospects of allogeneic CAR cell therapies, with special reference to CAR NK cells.

A 62-year-old woman with adult T-cell leukemia/lymphoma (ATL) received umbilical cord blood transplantation (CBT) in first complete remission. However, relapse of ATL was detected on day 74 post-transplantation, as evidenced by the rapid growth of lymphoma cells in peripheral blood and an increase in soluble interleukin-2 receptor (sIL2R) levels. Discontinuation of immunosuppressant therapy alone did not improve ATL findings, but treatment with lenalidomide caused lymphoma cells to disappear from the peripheral blood and sIL2R levels to return to normal. Pancytopenia was observed as a lenalidomide-associated adverse effect, but lymphocyte counts were not reduced. The patient was judged to be in complete remission based on results of Southern blot analysis and human T-cell leukemia virus 1 (HTLV-1)-infected cell analysis using flow cytometry (HAS-Flow). Flow cytometric analysis of peripheral blood and FISH analysis of X and Y chromosomes revealed that the therapeutic effect of lenalidomide was associated with an increase in the number of donor-derived peripheral natural killer cells. ATL relapse was not observed at 13 months into lenalidomide treatment. Our results suggest that lenalidomide is an effective option for the treatment of post-transplant relapsed ATL.

Allogeneic hematopoietic stem cell transplantation (allo-SCT) is a curative treatment option for multiple myeloma (MM), but few patients are eligible due to its high risk of treatment-related toxicity and relapse. Here, we report the feasibility and efficacy of allo-SCT after myeloablative conditioning with 8 Gy of total body irradiation (TBI) for reducing relapse of MM. We retrospectively analyzed data from 30 consecutive patients who received allo-SCT for MM after 8 Gy of TBI at Japanese Red Cross Medical Center between 2012 and 2021. Median age at allo-SCT was 47 (range 31-61) years. Stem-cell sources were peripheral blood from an HLA-matched related donor (MRD, n=5), bone marrow from an HLA-matched unrelated donor (MUD, n=5), bone marrow from an HLA-mismatched unrelated donor (MMUD, n=13), and cord blood (n=7). All patients received conditioning with 8 Gy of TBI combined with Flu/Mel (n=28) or others (n=2). Five-year PFS and 5-year OS were 36.7% and 46.2%, respectively. Sixteen patients died during the observation period (12 of primary disease and 4 of treatment-related toxicity). Patients with VGPR or better before allo-SCT had significantly better PFS (p=0.009) and OS (p=0.01) than others. Patients who received MMUD cells tended to have better PFS than those with other cell sources. Our report showed that allo-SCT for MM after 8 Gy of TBI is feasible, and the better PFS of MMUD suggests graft-versus-myeloma effects.

Aplastic anemia is a syndrome characterized by reduced hematopoietic stem cells, bone marrow hypoplasia, and pancytopenia, and is often considered a T-cell-mediated autoimmune disease. It is predominantly treated with hematopoietic stem cell transplantation and immunosuppressive therapy with anti-human thymocyte immunoglobulin (ATG) and cyclosporine. Only rabbit ATG was previously available in Japan, but equine ATG was recently approved for use in 2023. Thrombopoietin receptor agonists available in Japan are oral eltrombopag and injectable romiplostim. In hematopoietic stem cell transplantation for aplastic anemia, a conditioning regimen of reduced-dose cyclophosphamide and fludarabine has been used to reduce cardiotoxicity. Human leukocyte antigen haploidentical stem cell transplants have also been developed, and their use in patients without a donor is increasingly reported. Future advancements in novel drugs and transplantation therapies could revolutionize the management of aplastic anemia.

Chronic graft-versus-host disease (cGVHD) negatively impacts long-term survival and quality of life (QOL) after allogeneic hematopoietic cell transplantation. Corticosteroids are the first-line treatment for cGVHD, but approximately 30% to 70% of patients develop steroid-refractory cGVHD (SR-cGVHD), which has an extremely poor prognosis. The pathophysiology of cGVHD is more complicated than acute GVHD, but recent advances using murine models in conjunction with human studies indicate three major phases: 1) acute inflammation, 2) chronic inflammation with loss of immune tolerance, and 3) disrupted target organ homeostasis and fibrosis. Strategies that help prevent cGVHD include optimal donor selection and choice of conditioning regimen as well as pharmacologic and graft manipulation strategies. The key cellular mediators of SR-cGVHD are T cells, B cells, antigen-presenting cells, and fibroblasts. T cells and B cells are now targetable with the inhibitors ibrutinib and ruxolitinib, respectively. Recently, promising results have been obtained by modulating pathologic T cell responses with Rock2 inhibitors and targeting fibrosis with CSF-1R inhibitors. To optimize the use of these medications, a better understanding of the biological and target organ-specific mechanisms of cGVHD is needed. Here we review recent advances in cGVHD pathogenesis and discuss how best to implement recently approved biology-driven treatments for cGVHD.

There is growing recognition of post-transplant cyclophosphamide (PTCy) as the new standard prophylaxis for graft-versus-host disease (GVHD) in HLA-matched peripheral blood stem cell transplants with reduced intensity conditioning, based on recent results of randomized phase III trials of PTCy. Allogeneic hematopoietic cell transplantation (HCT) with PTCy is thought to have GVHD-dependent and -independent graft-versus-tumor (GVT) effects. Its GVHD-dependent effects may be attenuated by PTCy-induced alloreactive T cell dysfunction and preferential recovery of regulatory T cells after HCT, but its GVT effects do not appear to be significantly impaired in patients in remission or with indolent disease. As patients not in remission are often also candidates for transplantation in Japan, it will be necessary to use PTCy as a platform to establish a strategy that could also be effective in patients not in remission and to revise the donor selection algorithm.

Health surveys to assess adverse events after peripheral blood stem cell harvest (PBSCH) have conventionally been conducted by phone, but phone calls are suboptimal for conducting frequent surveys. We developed a web-based application (donor app) that enables donors to inform healthcare professionals (HCPs) of their health status as an electronic patient-reported outcome (ePRO). In this prospective observational study, we compared the usefulness of this donor app to phone calls for conducting health surveys. App users reported ePRO daily, and patients called by HCPs reported their health status at least once a week when called. The observation period was from the first administration of granulocyte colony-stimulating factor to the first follow-up visit after PBSCH, excluding the hospitalization period. Each group consisted of eight donors with a median age of 32 years (range: 19-58). Nine (56.3%) were female. There were eight related donors in the phone call group and four in the donor app group. During the observation period, HCPs obtained health status reports more frequently from app users than from phone call recipients (mean proportion of days with reports made during the observation period, 27.0% vs 53.5%; p<0.05). Average time spent by the HCPs for one follow-up and total follow-ups were both significantly shorter when the donor app was used. There were no differences in donor burden or satisfaction with donation. Our study suggests that use of a donor app could provide more detailed health survey data without increasing the burden on donors and HCPs.

Crohn's disease (CD) is a chronic and relapsing inflammatory bowel disease affecting the entire gastrointestinal tract. The prevalence of CD among Japanese people is increasing. One of the most frequent complications of CD is perianal fistulas. People living with CD may experience complex perianal fistulas, which can cause intense pain, bleeding, swelling, infection, and anal discharge. Despite medical and surgical advancements, complex perianal fistulas in CD remain challenging for clinicians to treat. CD patients living with perianal fistulas reported a negative impact on many aspects of their quality of life. Darvadstrocel is a cell therapy product containing a suspension of allogeneic expanded adipose-derived mesenchymal stem cells. It has been approved in Europe and Japan for the treatment of complex perianal fistulas that have shown an inadequate response to at least one conventional or biologic therapy in adult patients with non-active/mildly active luminal CD. By exhibiting immunomodulatory and local anti-inflammatory effects at the site of inflammation, it offers a new treatment option for complex perianal fistulas in CD patients. In this manuscript, the characteristic of darvadstrocel, the summary of results from the pivotal phase 3 studies in Europe and Japan, and the development strategy in Japan were introduced.

VEXAS syndrome is a new disease entity characterized by the presence of cytoplasmic vacuoles in blood cells, X-linked autoinflammatory symptoms, and somatic variants in UBA1, which encodes an E1 ubiquitin-activating enzyme. Around 30-50% of VEXAS syndrome patients have concurrent MDS. We and others have recently analyzed clinical and genetic features of MDS associated with VEXAS syndrome and found that most of these cases are categorized in the low-risk subgroup with low bone marrow blast percentages. MDS associated with VEXAS syndrome tended to involve a smaller number of genes and lower-risk genetic alterations than classical MDS. In addition, anemia in MDS associated with VEXAS syndrome with active inflammation before treatment tended to respond well to steroids. In this review, we will present our recent findings together with others, focusing on the new disease entity and pathophysiology of VEXAS syndrome and clinical/genetic features of associated MDS.

The primary cilium, an antenna-like structure of cell membrane, detects various signals and regulates cellular functions such as proliferation and differentiation. The impairment of primary cilium is associated with the etiologies of diseases including cancer, obesity, and congenital anomalies. In this review, novel functions of trichoplein, a suppressor of ciliogenesis, on the regulation of adipose progenitors and fibro-adipogenic progenitors are focused. Trichoplein-knockout mice show resistance to high-fat diet-induced obesity and accelerated regeneration after skeletal muscle injury. The primary cilia of adipose progenitors from trichoplein-knockout mice are elongated, leading to the inhibitions of the accumulation of lipid raft to the base of primary cilia and the phosphorylation of AKT. The primary cilia of fibro-adipogenic progenitors from trichoplein-knockout mice are also elongated, causing the increased expression of IL-13 through IL-33 receptor signaling. These mechanisms are involved in the resistance to diet-induced obesity and improved regeneration. These findings suggest that targeting the primary cilia of specific cells may be a novel therapeutic approach through modulating cellular functions.

The epigenome can adequately regulate the on/off states of genes in response to external environmental factors and stress. In recent years, it has been observed that the epigenome, which is modulated through DNA methylation, histone modifications, and chromatin remodeling, changes with age. Alterations in the epigenome lead to the loss of cell-specific epigenome/identity, which in turn triggers a decline in tissue function. In mammals, postnatal epigenomic variations are not only caused by metabolic diseases, such as diabetes or DNA damage, but also by social stress and infectious diseases. Unlike Genome-Wide Association Studies (GWAS), dynamically changing epigenomes, along with their cellular roles, need to be established as objective biomarkers in conjunction with various biological signals, such as walking speed, brain waves, and clinical data. The biological age/aging clock, determined by methylated DNA, has attracted attention, and calorie restriction not only slows the progression of aging, but also seems to suppress it. However, as indicated by gene expression analysis in aging mice, aging is not a linear model, but is represented by nonlinear dynamic changes. Consequently, the development of experimental models and analytical methods that enhance temporal resolution through time-series analysis, tailored to spatial resolution, such as cell distribution and organ specificity, is progressing. Moreover, in recent years, in addition to anti-aging efforts targeting epigenomic variations, global attention has increasingly focused on research and development aimed at rejuvenating treatments, thus leading to the birth of many biotech companies. Aging Hallmarks such as inflammation, stem cells, metabolism, genomic instability, and autophagy, interact closely with the epigenome. Various postnatal and reversible epigenomic controls of aging, including Yamanaka factors (OKSM and OSK), are now entering a new phase. In the future, the development of aging control using diverse modalities, such as mRNA, artificial peptides, and genome editing, is expected, along with an improved molecular understanding of aging and identification of useful biomarkers.

The transcription factor GATA-1 is essential for erythroid differentiation. Recently, FAM210B, which encodes a mitochondrial inner membrane protein, has been identified as a novel target of GATA-1. To clarify the role of FAM210B, we depleted endogenous FAM210B in human iPS-derived erythroid progenitor (HiDEP-1) cells, and found that erythroid differentiation was more pronounced in the FAM210B depleted cells. Comprehensive metabolite analysis revealed a decline in mitochondrial function accompanied by increased lactate production, indicative of anaerobic glycolysis. Mass spectrometry revealed that FAM210B could interact with multiple subunits of mitochondrial ATP synthases, such as subunit alpha (ATP5A) and beta (ATP5B). Our results suggested that FAM210B contributes prominently to erythroid differentiation by regulating mitochondrial energy metabolism. This review will discuss the potential association between mitochondrial metabolism and erythropoiesis.

A 41-year-old woman with right shoulder pain was found to have multiple tumors with osteolysis and M-proteinemia. Abnormal plasma cells (CD38+, CD138+, Igλ≫κ) were detected in 1.4% of bone marrow nucleated cells, and G-banding analysis revealed a 46,XX,t (8;14), (q24;q32) karyotype in 4 of 20 cells analyzed. A biopsy specimen from an extramedullary lesion had a packed proliferation of aberrant plasmacytoid cells with positive IgH::MYC fusion signals on fluorescence in situ hybridization. The patient was diagnosed with symptomatic multiple myeloma and treated with the BLd regimen, which significantly reduced M protein levels. Extramedullary lesions were initially reduced, but increased again after four cycles. The lesions disappeared with subsequent EPOCH chemotherapy and radiation, and complete remission was confirmed. The patient was then treated with high-dose chemotherapy with autologous peripheral blood stem cell transplantation. Complete remission was maintained for over one year with lenalidomide maintenance therapy. A solitary IgH::MYC chromosomal translocation is extremely rare in multiple myeloma and may be associated with high tumor proliferative capacity, multiple extramedullary lesions, and poor prognosis. Combined therapeutic modalities with novel and conventional chemotherapy and radiation might be a promising treatment strategy for patients with this type of multiple myeloma.

Recent advances in cancer therapy have significantly improved the survival rate of patients with cancer. In contrast, anti-cancer drug-induced adverse effects, especially cardiotoxicity, have come to affect patients' prognosis and quality of life. Therefore, there is a growing need to understand the anti-cancer drug-induced cardiotoxicity. Human induced pluripotent stem (iPS) cell-derived cardiomyocytes (hiPSC-CMs) have been used to assess drug-induced cardiotoxicity by improving the predictability of clinical cardiotoxicity and the principles of the 3Rs (replacement, reduction and refinement). To predict the anti-cancer drug-induced cardiotoxicity, we developed a novel method to assess drug-induced proarrhythmia risk using hiPSC-CMs by participating in the international validation. In addition, we established the chronic contractility toxicity assessment by image-based motion analysis. The compound BMS-986094, which was withdrawn from clinical trials, inhibited contractility velocity and relaxation velocity in hiPSC-CMs. Currently, we are trying to investigate the predictability of the contractility assay by comparing the hiPSC-CM data with adverse events reports from real-world database. In this review, we would like to introduce the novel imaging-based contractility method using hiPSC-CMs and future perspectives in anti-cancer drug-induced cardiotoxicity.