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Chimeric antigen receptor T cells (CAR-T) therapy has achieved remarkable therapeutic success in treating a variety of hematopoietic malignancies. However, the high relapse rate and poor in vivo persistence, partially caused by CAR-T cell exhaustion, are still important barriers against CAR-T therapy. It remains largely elusive on the mechanisms of CAR-T exhaustion and how to attenuate exhaustion to achieve better therapeutic efficacy. In this study, we initially observed that CAR-T cells showed rapid differentiation and increased exhaustion after co-culture with tumor cells in vitro, and then performed single-cell ATAC-seq to depict the comprehensive and dynamic landscape of chromatin accessibility of CAR-T cells during tumor cell stimulation. Analyses of differential chromatin accessible regions and motif accessibility revealed that TFs were distinct in each cell type and reconstituted a coordinated regulatory network to drive CAR-T exhaustion. Furthermore, we performed scATAC-seq in patient-derived CAR-T cells and identified BATF and IRF4 as pivotal regulators in CAR-T cell exhaustion. Finally, knockdown of BATF or IRF4 enhanced the killing ability, inhibited exhaustion, and prolonged the persistence of CAR-T cells in vivo. Together, our study unraveled the epigenetic regulatory mechanisms of CAR-T exhaustion and provided new insights into CAR-T engineering to achieve better clinical treatment benefits.

Although widely applied in treating hematopoietic malignancies, transplantation of hematopoietic stem/progenitor cells (HSPCs) is impeded by HSPC shortage. Whether circulating HSPCs (cHSPCs) in steady-state blood could be used as an alternative source remains largely elusive. Here we develop a three-dimensional culture system (3DCS) including arginine, glycine, aspartate, and a series of factors. Fourteen-day culture of peripheral blood mononuclear cells (PBMNCs) in 3DCS led to 125- and 70-fold increase of the frequency and number of CD34+ cells. Further, 3DCS-expanded cHSPCs exhibited the similar reconstitution rate compared to CD34+ HSPCs in bone marrow. Mechanistically, 3DCS fabricated an immunomodulatory niche, secreting cytokines as TNF to support cHSPC survival and proliferation. Finally, 3DCS could also promote the expansion of cHSPCs in patients who failed in HSPC mobilization. Our 3DCS successfully expands rare cHSPCs, providing an alternative source for the HSPC therapy, particularly for the patients/donors who have failed in HSPC mobilization.

The dysregulation of miRNAs has been increasingly recognized as a critical mediator of cancer development and progression. Here, we show that frequent deletion of the MIR135A1 locus is associated with poor prognosis in primary breast cancer. Forced expression of miR-135a decreased breast cancer progression, while inhibition of miR-135a with a specific miRNA sponge elicited opposing effects, suggestive of a tumor suppressive role of miR-135a in breast cancer. Estrogen receptor alpha (ERα) bound the promoter of MIR135A1 for its transcriptional activation, whereas tamoxifen treatment inhibited expression of miR-135a in ERα+ breast cancer cells. miR-135a directly targeted ESR1, ESRRA, and NCOA1, forming a negative feedback loop to inhibit ERα signaling. This regulatory feedback between miR-135a and ERα demonstrated that miR-135a regulated the response to tamoxifen. The tamoxifen-mediated decrease in miR-135a expression increased the expression of miR-135a targets to reduce tamoxifen sensitivity. Consistently, miR-135a expression was downregulated in ERα+ breast cancer cells with acquired tamoxifen resistance, while forced expression of miR-135a partially resensitized these cells to tamoxifen. Tamoxifen resistance mediated by the loss of miR-135a was shown to be partially dependent on the activation of the ERK1/2 and AKT pathways by miR-135a–targeted genes. Taken together, these results indicate that deletion of the MIR135A1 locus and decreased miR-135a expression promote ERα+ breast cancer progression and tamoxifen resistance.Significance: Loss of miR-135a in breast cancer disrupts an estrogen receptor-induced negative feedback loop, perpetuating disease progression and resistance to therapy.Graphical Abstract: http://cancerres.aacrjournals.org/content/canres/78/17/4915/F1.large.jpg. Cancer Res; 78(17); 4915–28. ©2018 AACR.

Supplementary table S1 lists all the sequence of miR-135a mimic, siRNAs, shRNAs, and primers for plasmid construction, ChIP assay, and qRT-PCR used in this study. Supplementary table S2 shows the information of antibodies used in this study. Supplementary table S3 and S4 show the correlation of MIR135A1 deletion and low miR-135a-1 with the clinicopathologic features of breast cancer in the TCGA breast tumors, respectively. Supplementary table S5 lists genes including EMT markers, ERα cofactors and interacting proteins, and predicted miR-135a targets which expression levels negatively correlate with miR-135a in breast tumors. Supplementary figure S1 shows that the copy number of MIR135A1 gene and the expression levels of pri-miR-135a-1 are decreased in breast cancer. Supplementary figure S2 shows that miR-135a is a tumor suppressor gene in T47D and MDA-MB-231 cells and inhibits EMT of breast cancer cells. Supplementary figure S3 shows that miR-135a impairs the in vivo tumor growth. Supplementary figure S4 shows that the transcription of MIR135A1 positively correlates with E2-ERα axis. Supplementary figure S5 shows that miR-135a modulates the estrogen independent growth and the response to tamoxifen in ERα+ breast cancer cells. Supplementary figure S6 shows the validation of miR-135a targets in breast cancer cells. Supplementary figure S7 shows that miR-135a modulates ERα-ERK-AKT crosstalk.

PDF file - 1291K, FIGURE S1: Representative data of genomic real-time PCR screening for DNA copy numbers of aberrant microRNAs on human chromosome 3p in breast carcinoma and the correlation between expression levels of miR-128 and ARPP21; FIGURE S2: miR-128 depletion confers non-tumorigenic MCF-10A cells with a malignant phenotype in vitro; FIGURE S3: Forced expression of miR-128 alters the morphology and reduces migration and invasion of MCF-7 cells in vitro, whereas miR-128 depletion confers non-tumorigenic HBL-100 cells with a malignant phenotype in vitro; FIGURE S4: miR-128 depletion confers a malignant phenotype on non-tumorigenic MCF-10A cells in vivo and miR-128 represses the CD44+CD24 Neg/Low population in mammary carcinoma cells; FIGURE S5: Interactions between miR-128 and its binding targets; FIGURE S6: Luciferase assay for promoter activity

PDF file - 226K, TABLE S1: List of miRNAs on human chromosome 3p and the changes of their genome copies determined by genomic real-time PCR; TABLE S2: Clinical significance of the expression of miR-128 in breast cancer patients; TABLE S3: Invariable analysis of clinical outcome according to the expression of miR-128 in breast cancer patients (85 cases); TABLE S4: List of mRNAs predicted to be targeted by miR-128 by all the three programs: MiRanda, Pictar and TargetScan; TABLE S5: qRT-PCR analysis of the effect of forced expression of miR-128 in MDA-MB-231 cells or depleted expression of miR-128 in MCF-10A cells on mRNA levels of several key genes functionally involved in oncogenicity and neoplastic progression; TABLE S6: Sequences of the oligonucleotides for miRNAs, siRNAs, construct-making and ChIP assays; TABLE S7: Sequence of the oligonucleotides for real-time PCR experiments; TABLE S8: List of proteins tested by antibodies and characteristics of the corresponding antibodies used

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Screening of the entire let-7 family of microRNAs (miRNA) by in situ hybridization identified let-7g as the only member, the diminished expression of which was significantly associated with lymph node metastasis and poor survival in breast cancer patients. Abrogation of let-7g expression in otherwise nonmetastatic mammary carcinoma cells elicited rapid metastasis from the orthotopic location, through preferential targets, Grb2-associated binding protein 2 (GAB2) and fibronectin 1 (FN1), and consequent activation of p44/42 mitogen-activated protein kinase (MAPK) and specific matrix metalloproteinases. Treatment with estrogen or epidermal growth factor specifically reduced the expression of mature let-7g through activation of p44/42 MAPK and subsequently stimulated expression of GAB2 and FN1, which, in turn, promoted tumor invasion. We thus identify let-7g as a unique member of the let-7 miRNA family that can serve as a prognostic biomarker in breast cancer and also propose a paradigm used by specific signaling molecules via let-7g to cooperatively promote breast cancer invasion and metastasis. Thus, let-7 family members neither possess equivalent clinicopathologic correlation nor function in breast cancer. Cancer Res; 71(20); 6463–74. ©2011 AACR.

PDF file - 190K, Includes patient and specimen data, reagent information, miRNA and siRNA transfection, and statistical analysis. Also includes supplementary references

Acute myeloid leukemia (AML) is a deadly hematological malignancy. Cellular morphology detection of bone marrow smears based on the French–American–British (FAB) classification system remains an essential criterion in the diagnosis of hematological malignancies. However, the diagnosis and discrimination of distinct FAB subtypes of AML obtained from bone marrow smear images are tedious and time-consuming. In addition, there is considerable variation within and among pathologists, particularly in rural areas, where pathologists may not have relevant expertise. Here, we established a comprehensive database encompassing 8245 bone marrow smear images from 651 patients based on a retrospective dual-center study between 2010 and 2021 for the purpose of training and testing. Furthermore, we developed AMLnet, a deep-learning pipeline based on bone marrow smear images, that can discriminate not only between AML patients and healthy individuals but also accurately identify various AML subtypes. AMLnet achieved an AUC of 0.885 at the image level and 0.921 at the patient level in distinguishing nine AML subtypes on the test dataset. Furthermore, AMLnet outperformed junior human experts and was comparable to senior experts on the test dataset at the patient level. Finally, we provided an interactive demo website to visualize the saliency maps and the results of AMLnet for aiding pathologists’ diagnosis. Collectively, AMLnet has the potential to serve as a fast prescreening and decision support tool for cytomorphological pathologists, especially in areas where pathologists are overburdened by medical demands as well as in rural areas where medical resources are scarce.

The nucleolus is the organelle for ribosome biogenesis and sensing various types of stress. However, its role in regulating stem cell fate remains unclear. Here, we present evidence that nucleolar stress induced by interfering rRNA biogenesis can drive the 2-cell stage embryo-like (2C-like) program and induce an expanded 2C-like cell population in mouse embryonic stem (mES) cells. Mechanistically, nucleolar integrity maintains normal liquid-liquid phase separation (LLPS) of the nucleolus and the formation of peri-nucleolar heterochromatin (PNH). Upon defects in rRNA biogenesis, the natural state of nucleolus LLPS is disrupted, causing dissociation of the NCL/TRIM28 complex from PNH and changes in epigenetic state and reorganization of the 3D structure of PNH, which leads to release of Dux, a 2C program transcription factor, from PNH to activate a 2C-like program. Correspondingly, embryos with rRNA biogenesis defect are unable to develop from 2-cell (2C) to 4-cell embryos, with delayed repression of 2C/ERV genes and a transcriptome skewed toward earlier cleavage embryo signatures. Our results highlight that rRNA-mediated nucleolar integrity and 3D structure reshaping of the PNH compartment regulates the fate transition of mES cells to 2C-like cells, and that rRNA biogenesis is a critical regulator during the 2-cell to 4-cell transition of murine pre-implantation embryo development., The role of the nucleolus in cell fate is not well known. Here the authors show nucleolar integrity maintained by rRNA-mediated liquid-liquid phase separation and 3D chromatin structure of perinucleolar heterochromatin regulates the fate transition of mouse embryonic stem cells to 2 cell-stage embryo-like cells, and that rRNA biogenesis regulates the 2-cell-to-4-cell transition of development.

Chimeric Antigen-Receptor (CAR) T-cell therapies have shown dramatic efficacy in treating relapsed and refractory cancers, especially B cell malignancies. However, these innovative therapies cause adverse toxicities that limit the broad application in clinical settings. Hematologic cytopenias, one frequently reported adverse event following CAR T cell treatment, are manifested as a disorder of hematopoiesis with decreased number of mature blood cells and subdivided into anemia, thrombocytopenia, leukopenia, and neutropenia, which increase the risk of infections, fatigue, bleeding, fever, and even fatality. Herein, we initially summarized the symptoms, etiology, risk factors and management of cytopenias. Further, we elaborated the cellular and molecular mechanisms underlying the initiation and progression of cytopenias following CAR T cell therapy based on previous studies about acquired cytopenias. Overall, this review will facilitate our understanding of the etiology of cytopenias and shed lights into developing new therapies against CAR T cell-induced cytopenias.

Immune cells such as T cells, macrophages, dendritic cells, and other immunoregulatory cells undergo metabolic reprogramming in cancer and inflammation‐derived microenvironment to meet specific physiologic and functional demands. Myeloid‐derived suppressor cells (MDSCs) are a heterogeneous population of immature myeloid cells that are characterized by immunosuppressive activity, which plays a key role in host immune homeostasis. In this review, we have discussed the core metabolic pathways, including glycolysis, lipid and fatty acid biosynthesis, and amino acid metabolism in the MDSCs under various pathologic situations. Metabolic reprogramming is a determinant of the phenotype and functions of MDSCs, and is therefore a novel therapeutic possibility in various diseases., Graphical Abstract This work intends to declare what metabolic alterations happen in MDSCs to support their physiologic and functional demands under a variety of pathologic situations.

Acute graft-versus-host disease (aGVHD) is the most severe complication and the major cause of morbidity and mortality in patients who have received hematopoietic stem cell transplantation. Inflammation mediated by donor T cells and neighboring recipient cells is considered to be responsible for intestinal aGVHD. Interleukin (IL)-17A-producing γδT (γδT17) cells have been investigated as key players in cancer, immunity, and inflammatory responses because of their phenotypic plasticity, memory-like activity, and unique migratory features. However, the precise roles and underlying mechanisms of γδT17 cells in aGVHD immunopathogenesis remain elusive. Here, we found that γδT17 cells constituted the major resident γδT population in the intestinal lamina propria of aGVHD mice. Adoptive infusion of induced γδT17 cells markedly attenuated murine aGVHD and increased infiltration of Gr-1+CD11b+ myeloid-derived suppressor cells (MDSCs) into the inflammatory intestine, and did not affect graft-versus-leukemia(GVL) effect. Further experiments indicated that γδT17 cells enhanced both migration ability and immunosuppressive activity of MDSCs. γδT17 cell-mediated protection in aGVHD was blunted by depletion of IL-17A or MDSCs. Our study clarifies an immune pathway where γδT17 cells play a protective role in murine aGVHD by recruiting MDSCs to the inflammatory intestine.

Evidence on the importance of lymph node (LN) dissection during resection for small cell lung cancer (SCLC) is scarce. This study sought to investigate the clinical impact of the extent of lymphadenectomy on the survival of patients with SCLC.Patients who underwent resection for primary SCLC between 2000 and 2016 were identified from the Surveillance, Epidemiology, and End Results (SEER) cancer registry. The patients were stratified based on the number of LNs dissected (0, 1-3, 4-11, and ≥12) via an X-Tile software analysis, and lung cancer-specific survival (LCSS) and overall survival (OS) were compared between these stratified groups using Kaplan-Meier curves. A propensity score-matched analysis and a Cox regression model were used to adjust for potential confounders.A total of 1,883 patients with SCLC met our criteria and were enrolled in the study. The LCSS and OS analyses revealed that patients who underwent LN dissection during surgery had longer survival times significantly than patients who did not. Similarly, patients who underwent more extensive LN dissection (≥4 LNs) had longer survival times than those who underwent less extensive LN dissection (1-3 LNs). However, no significant increase in survival time was found for patients who underwent the dissection of ≥12 LNs compared to those who underwent the dissection of 4-11 LNs. These results were confirmed in our propensity-matched and Cox regression analyses.Our study revealed that patient survival after surgical resection for SCLC is associated with the number of dissected LNs, and the number of LNs for dissection ranges from 4 to 11 achieve the best survival outcome.

Summary Mouse embryonic stem cells (ESCs) cultured in defined medium resemble the pre-implantation epiblast in the ground state, with full developmental capacity including the germline. β-Catenin is required to maintain ground state pluripotency in mouse ESCs, but its exact role is controversial. Here, we reveal a Tcf3-independent role of β-catenin in restraining germline and somatic lineage differentiation genes. We show that β-catenin binds target genes with E2F6 and forms a complex with E2F6 and HMGA2 or E2F6 and HP1γ. Our data indicate that these complexes help β-catenin restrain and fine-tune germ cell and neural developmental potential. Overall, our data reveal a previously unappreciated role of β-catenin in preserving lineage differentiation integrity in ground state ESCs., Highlights • β-Catenin depletion irreversibly compromised lineage development of ground state ESCs • TCF3-independent role of β-catenin in determining lineage differentiation potential • E2F6, HP1γ, and HMGA2 are β-catenin interaction partners and co-bound to target genes • β-Catenin and protein partners fine-tune germline and neural development potential, In this article, Tao and colleagues show that β-catenin functions beyond derepressing TCF3 targeted pluripotency network in ESCs. β-Catenin physically interacts with protein partners, including transcription factor and chromatin modulators. Together, they preserve the integrity of lineage differentiation of ground state ESCs by restraining and fine-tuning their germline and neural development potential.

Leukaemia stem cells (LSCs) underlie cancer therapy resistance but targeting these cells remains difficult. The Wnt-β-catenin and PI3K-Akt pathways cooperate to promote tumorigenesis and resistance to therapy. In a mouse model in which both pathways are activated in stem and progenitor cells, LSCs expanded under chemotherapy-induced stress. Since Akt can activate β-catenin, inhibiting this interaction might target therapy-resistant LSCs. High-throughput screening identified doxorubicin (DXR) as an inhibitor of the Akt-β-catenin interaction at low doses. Here we repurposed DXR as a targeted inhibitor rather than a broadly cytotoxic chemotherapy. Targeted DXR reduced Akt-activated β-catenin levels in chemoresistant LSCs and reduced LSC tumorigenic activity. Mechanistically, β-catenin binds multiple immune-checkpoint gene loci, and targeted DXR treatment inhibited expression of multiple immune checkpoints specifically in LSCs, including PD-L1, TIM3 and CD24. Overall, LSCs exhibit distinct properties of immune resistance that are reduced by inhibiting Akt-activated β-catenin. These findings suggest a strategy for overcoming cancer therapy resistance and immune escape.

Adults with acute myeloid leukaemia have unsatisfactory clinical outcomes and rates of complete remission. Venetoclax combined with azacytidine or low-dose cytarabine has shown efficacy in adults aged 75 years or older (or 18-74 years with comorbidities precluding intensive chemotherapy) with acute myeloid leukaemia. We aimed to investigate the activity and safety of venetoclax plus 3+7 daunorubicin and cytarabine chemotherapy in adults with acute myeloid leukaemia.We conducted a two-stage, single-arm, phase 2 trial at three public hospitals in China. We enrolled patients aged 18-60 years with previously untreated de novo acute myeloid leukaemia and an Eastern Cooperative Oncology Group performance status of 0-2. Patients received induction treatment with intravenous daunorubicin (60 mg/mBetween Dec 25, 2020, and July 7, 2021, 36 patients were assessed for eligibility and 33 were enrolled. 15 (45%) patients were men and 18 (55%) were women, and all were Asian. The composite complete remission rate after one cycle of DAV regimen was 91% (95% CI 76-98; 30 of 33 patients) in the entire cohort. 29 (97%) of 30 patients who reached complete remission had undetectable measurable residual disease (ie,0·1%). Grade 3 or worse adverse events included neutropenia in 33 (100%) of 33 patients, thrombocytopenia in 33 (100%), anaemia in 33 (100%), febrile neutropenia in 18 (55%), pneumonia in seven (21%), and sepsis in four (12%). No treatment-related deaths occurred. With a median follow-up of 11 months (IQR 9-12), estimated 1-year overall survival was 97% (95% CI 91-100) and 1-year event-free survival was 72% (56-94).The DAV regimen represents an effective induction therapy for newly diagnosed adults with acute myeloid leukaemia, which resulted in a high rate of complete remission. These findings are an important contribution to the field, showing a safe strategy to incorporate venetoclax into the most common induction regimen used to treat newly diagnosed acute myeloid leukaemia internationally.Leading Innovative and Entrepreneur Team Introduction Program of Zhejiang, National Natural Science Foundation of China, Key Research and Development Program of Zhejiang.For the Chinese translation of the abstract see Supplementary Materials section.

Myeloid-derived suppressor cells (MDSCs), a population of myeloid lineage cells with immunosuppressive capacity, can mitigate acute graft-versus-host disease (aGVHD) after allogeneic hematopoietic stem cell transplantation (allo-HSCT). We previously found that the immunosuppressive function of polymorphonuclear population (PMN-MDSCs) was impaired in aGVHD milieu. The aim of this study was to explore the intrinsic mechanism regulating the fate and function of donor-derived PMN-MDSCs during allo-HSCT. We firstly found that mitochondrial permeability transition pore (MPTP) opened in the PMN-MDSCs in response to the intense inflammatory environment of aGVHD, which induced mitochondrial damage, oxidative stress, and apoptosis of PMN-MDSCs. Inhibiting MPTP opening by a traditional immunosuppressant, cyclosporine A (CsA), could restore the immunosuppressive function and viability of PMN-MDSCs in vitro and in vivo, which reveals a new mechanism of CsA application.

Chimeric antigen receptor (CAR) T cells are potent agents for recognizing and eliminating tumors, and have achieved remarkable success in the treatment of patients with refractory leukemia and lymphoma. However, dysfunction of T cells, including exhaustion, is an inevitable obstacle for persistent curative effects. Here, the authors initially found that calcium signaling is hyperactivated via sustained tonic signaling in CAR-T cells. Next, it is revealed that the store-operated calcium entry (SOCE) inhibitor BTP-2, but not the calcium chelator BAPTA-AM, markedly diminishes CAR-T cell exhaustion and terminal differentiation of CAR-T cells in both tonic signaling and tumor antigen exposure models. Furthermore, BTP-2 pretreated CAR-T cells show improved antitumor potency and prolonged survival in vivo. Mechanistically, transcriptome and metabolite analyses reveal that treatment with BTP-2 significantly downregulate SOCE-calcineurin-nuclear factor of activated T-cells (NFAT) and glycolysis pathways. Together, the results indicate that modulating the SOCE-calcineurin-NFAT pathway in CAR-T cells renders them resistant to exhaustion, thereby yielding CAR products with enhanced antitumor potency.

Cancer cells display phenotypic equilibrium between the stem-like and differentiated states during neoplastic homeostasis. The functional and mechanistic implications of this subpopulation plasticity remain largely unknown. Herein, it was demonstrated that the breast cancer stem cell (BCSC) secretome autonomously compressed the stem cell population. Co-implantation with BCSCs decreased the tumor-initiating capacity yet increased metastasis of accompanying cancer cells, wherein DKK1 was identified as a pivotal factor secreted by BCSCs for such functions. DKK1-promoted differentiation was indispensable for disseminated tumor cell metastatic outgrowth. In contrast, DKK1 inhibitors substantially relieved the metastatic burden by restraining metastatic cells in the dormant state. DKK1 increased the expression of SLC7A11 to protect metastasizing cancer cells from lipid peroxidation and ferroptosis. Combined treatment with ferroptosis inducer and DKK1 inhibitor exhibited synergistic effects in diminishing metastasis. Hence, this study deciphers the contribution of CSC-regulated phenotypic plasticity in metastatic colonization and provides therapeutic approaches to limit metastatic outgrowth.

Relapses of CD19-expressing leukemia in patients who achieved initial remission after CART cell treatment have been reported to correlate with poor CART cells persistence. Sustained tonic signaling or strong activation drives CART cell differentiation and exhaustion, which limit the therapeutic efficacy and persistence of CART cells. Here, we identified dasatinib as the optimal candidate to prevent or reverse both CD28/CART and 4-1BB/CART cell differentiation and exhaustion during ex vivo expansion, which profoundly enhanced the therapeutic efficacy and in vivo persistence. Moreover, strong activation-induced CART cells differentiation, exhaustion and apoptosis driven by CD3/CD28 stimulation or antigen exposure were dramatically prevented or reversed by dasatinib treatment. Mechanistically, dasatinib markedly reduced the phosphorylation of Src and Lck, and downregulated the expression of genes involved in CAR signaling pathways, which resulted in the optimization of cell differentiation, exhaustion and apoptosis-related gene expression. Our study proposes a promising pharmacological approach for optimizing CART cells manufacture, and provides an experimental basis for reinvigorating CART cells in clinical application. Supplementary Information The online version contains supplementary material available at 10.1186/s13045-021-01117-y.

Polycomb group (PcG) proteins maintain cell identity by repressing gene expression during development. Surprisingly, emerging studies have recently reported that a number of PcG proteins directly activate gene expression during cell fate determination process. However, the mechanisms by which they direct gene activation in pluripotency remain poorly understood. Here, we show that Phc1, a subunit of canonical polycomb repressive complex 1 (cPRC1), can exert its function in pluripotency maintenance via a PRC1-independent activation of Nanog. Ablation of Phc1 reduces the expression of Nanog and overexpression of Nanog partially rescues impaired pluripotency caused by Phc1 depletion. We find that Phc1 interacts with Nanog and activates Nanog transcription by stabilizing the genome-wide chromatin interactions of the Nanog locus. This adds to the already known canonical function of PRC1 in pluripotency maintenance via a PRC1-dependent repression of differentiation genes. Overall, our study reveals a function of Phc1 to activate Nanog transcription through regulating chromatin architecture and proposes a paradigm for PcG proteins to maintain pluripotency., Phc1 is a subunit of the polycomb repressive complex 1 (PRC1), which represses gene expression during development. Here the authors show that Phc1 acts independently from PRC1 to activate Nanog transcription by stabilizing genome-wide chromatin interactions of the Nanog locus, and in turn stabilize pluripotency.

Fat mass and obesity-associated protein (FTO) is the first protein found to have the activity of N6-methyladenosine (m6A) demethylation. It has been reported that FTO was involved in different physiological and pathological processes, including stem cell differentiation, sex determination, tumorigenesis, and progression. To further understand the exact role of FTO in these processes, we generated a FTO knockout human embryonic stem cell (hESC) line by CRISPR/Cas9 mediated gene editing method. This cell line maintained normal karyotype, pluripotency, and trilineage differentiation potential, which are considered as a model for function studies of the FTO protein in hESC self-renewal and differentiation.

Background Decades of efforts have attempted to differentiate the pluripotent stem cells (PSCs) into truly functional hematopoietic stem cells (HSCs), yet the problems of low differentiation efficiency in vitro and poor hematopoiesis reconstitution in vivo still exist, mainly attributing to the lack of solid, reproduced, or pursued differentiation system. Methods In this study, we established an in vitro differentiation system yielding in vivo hematopoietic reconstitution hematopoietic cells from mouse PSCs through a 3D induction system followed by coculture with OP9 stromal cells. The in vivo hematopoietic reconstitution potential of c-kit+ cells derived from the mouse PSCs was evaluated via m-NSG transplantation assay. Flow cytometry analysis, RNA-seq, and cell cycle analysis were used to detect the in vitro hematopoietic ability of endothelial protein C receptor (EPCR, CD201) cells generated in our induction system. Results The c-kit+ cells from 3D self-assembling peptide induction system followed by the OP9 coculture system possessed apparently superiority in terms of in vivo repopulating activity than that of 3D induction system followed by the 0.1% gelatin culture. We interestingly found that our 3D+OP9 system enriched a higher percentage of CD201+c-kit+cells that showed more similar HSC-like features such as transcriptome level and CFU formation ability than CD201-c-kit+cells, which have not been reported in the field of mouse PSCs hematopoietic differentiation. Moreover, CD201+ hematopoietic cells remained in a relatively slow cycling state, consistent with high expression levels of P57 and Ccng2. Further, we innovatively demonstrated that notch signaling pathway is responsible for in vitro CD201+ hematopoietic cell induction from mouse PSCs. Conclusions Altogether, our findings lay a foundation for improving the efficiency of hematopoietic differentiation and generating in vivo functional HSC-like cells from mouse PSCs for clinical application.

This study demonstrates that nuclear DEK governs quiescence and metabolic homeostasis of HSC and preserves HSC potential. Mechanistically, DEK dampens chromatin accessibility in HSC by inducing deacetylation of H3K27 and governs gene expression, including Akt1/2., The oncogene DEK is found fused with the NUP214 gene creating oncoprotein DEK-NUP214 that induces acute myeloid leukemia (AML) in patients, and secreted DEK protein functions as a hematopoietic cytokine to regulate hematopoiesis; however, the intrinsic role of nuclear DEK in hematopoietic stem cells (HSCs) remains largely unknown. Here, we show that HSCs lacking DEK display defects in long-term self-renew capacity, eventually resulting in impaired hematopoiesis. DEK deficiency reduces quiescence and accelerates mitochondrial metabolism in HSCs, in part, dependent upon activating mTOR signaling. At the molecular level, DEK recruits the corepressor NCoR1 to repress acetylation of histone 3 at lysine 27 (H3K27ac) and restricts the chromatin accessibility of HSCs, governing the expression of quiescence-associated genes (e.g., Akt1/2, Ccnb2, and p21). Inhibition of mTOR activity largely restores the maintenance and potential of Dek-cKO HSCs. These findings highlight the crucial role of nuclear DEK in preserving HSC potential, uncovering a new link between chromatin remodelers and HSC homeostasis, and have clinical implications.

Venetoclax (VEN) plus azacitidine has become the first-line therapy for elderly patients with acute myeloid leukemia (AML), and has a complete remission (CR) plus CR with incomplete recovery of hemogram rate of ≥70%. However, the 3-year survival rate of these patients is BCL2 mutation was not present in our cohort, indicating that, in contrast to chronic lymphocytic leukemia, this BCL2 mutation is dispensable for acquired VEN resistance in AML. Instead, we found that reconstructed existing mutations, especially dominant mutation conversion (e.g., expanded FLT3-ITD), rather than newly emerged mutations (e.g., TP53 mutation), mainly contributed to VEN resistance in AML. According to our results, the combination of precise mutational monitoring and advanced interventions with targeted therapy or chemotherapy are potential strategies to prevent and even overcome acquired VEN resistance in AML.

Although widely applied in treating hematopoietic malignancies, transplantation of hematopoietic stem/progenitor cells (HSPCs) is impeded by HSPC shortage. Whether circulating HSPCs (cHSPCs) in steady-state blood could be used as an alternative source remains largely elusive. Here we develop a three-dimensional culture system (3DCS) including arginine, glycine, aspartate, and a series of factors. Culture of peripheral blood mononuclear cells (PBMNCs) in 3DCS led to 125- and 70-fold increase of the frequency and number of CD34+ cells. Further, 3DCS-expanded cHSPCs exhibited 1.65-fold increase in the reconstitution rate compared to CD34+ HSPCs in bone marrow. Mechanistically, 3DCS fabricated an immunomodulatory niche, secreting cytokines as TNF-α to support cHSPC survival and proliferation. Finally, 3DCS could also promote the expansion of cHSPCs in patients who failed in HSPC mobilization. Our 3DCS successfully expands rare cHSPCs, providing an alternative source for the HSPC therapy, particularly for the patients/donors who have failed in HSPC mobilization.

Acute graft-versus-host disease (aGVHD) is the most severe complication and the major cause of morbidity and mortality in patients who have received hematopoietic stem cell transplantation. Inflammation mediated by donor T cells and neighboring recipient cells is considered to be responsible for intestinal aGVHD. Interleukin (IL)-17A-producing γδT (γδT17) cells have been investigated as key players in cancer, immunity, and inflammatory responses because of their phenotypic plasticity, memory-like activity, and unique migratory features. However, the precise roles and underlying mechanisms of γδT17 cells in aGVHD immunopathogenesis remain elusive. Here, we found that γδT17 cells constituted the major resident γδT population in the intestinal lamina propria of aGVHD mice. Adoptive infusion of induced γδT17 cells markedly attenuated murine aGVHD and increased infiltration of Gr-1

The mutations in the SARS-CoV-2 virus genome during COVID-19 dissemination are unclear. In 788 COVID-19 patients from Zhejiang province, we observed decreased rate of severe/critical cases compared with patients in Wuhan. For mechanisms exploration, we isolated one strain of SARS-CoV-2 (ZJ01) from a mild COVID-19 patient. Thirty-five specific gene mutations were identified. Phylogenetic and relative synonymous codon usage analysis suggested that ZJ01 may be a potential evolutionary branch of SARS-CoV-2. We classified 54 global virus strains based on the base (C or T) at positions 8824 and 28247 while ZJ01 has T at both sites. The prediction of the Furin cleavage site (FCS) and sequence alignment indicated that the FCS may be an important site of coronavirus evolution. ZJ01 mutations identified near the FCS (F1-2) caused changes in the structure and electrostatic distribution of the S surface protein, further affecting the binding capacity of Furin. Single-cell sequencing and ACE2-Furin co-expression results confirmed that the Furin expression was especially higher in glands, liver, kidneys, and colon. The evolutionary pattern of SARS-CoV-2 towards FCS formation may result in its clinical symptom becoming closer to HKU-1 and OC43 caused mild flu-like symptoms, further showing its potential in differentiating into mild COVID-19 subtypes.

Hematopoietic stem cell (HSC) aging, which is accompanied by reduced self-renewal ability, impaired homing, myeloid-biased differentiation, and other defects in hematopoietic reconstitution function, is a hot topic in stem cell research. Although the number of HSCs increases with age in both mice and humans, the increase cannot compensate for the defects of aged HSCs. Many studies have been performed from various perspectives to illustrate the potential mechanisms of HSC aging; however, the detailed molecular mechanisms remain unclear, blocking further exploration of aged HSC rejuvenation. To determine how aged HSC defects occur, we provide an overview of differences in the hallmarks, signaling pathways, and epigenetics of young and aged HSCs as well as of the bone marrow niche wherein HSCs reside. Notably, we summarize the very recent studies which dissect HSC aging at the single-cell level. Furthermore, we review the promising strategies for rejuvenating aged HSC functions. Considering that the incidence of many hematological malignancies is strongly associated with age, our HSC aging review delineates the association between functional changes and molecular mechanisms and may have significant clinical relevance.

The outbreak of COVID-19 become enormous threat to human beings, showing unclear virus mutation during dissemination. We found, in our 788 confirmed COVID-19 patients, the decreased rate of severe/critical type, increased liver/kidney damage and prolonged period of nuclear acid positivity, when compared with Wuhan. To investigate underlining mechanisms, we isolated one strain of SARS-CoV-2 (ZJ01) in mild COVID-19 patient and found the existence of 35 specific gene mutation by gene alignment. Further phylogenetic analysis and RSCU heat map results suggested that ZJ01 may be a potential evolutionary branch of SARS-CoV-2. We classified 54 strains of viruses worldwide (C/T type) based on the base (C or T) at positions 8824 and 28247. ZJ01 has both T at those sites, becoming the only TT type currently identified in the world. The prediction of Furin cleavage site (FCS) and the sequence alignment of virus family indicated that FCS may be an important site of coronavirus evolution. ZJ01 had mutations near FCS (F1-2), which caused changes in the structure and electrostatic distribution of S protein surface, further affecting the binding capacity of Furin. Single cell sequencing and ACE2-Furin co-expression results confirmed that Furin level was higher in the whole body, especially in glands, liver, kidney and colon while FCS may help SARS-CoV-2 infect these organs. The evolutionary pattern of SARS-CoV-2 towards FCS formation may result in its clinical symptom becoming closer to HKU-1 and OC43 (the source of FCS sequence-PRRA) caused influenza, further showing potential in differentiating into mild COVID-19 subtypes.

Transplantation of hematopoietic stem cells (HSCs) shows promising prospect for broad-spectrum hematological disorder therapy, owing to their potential to generate the entire hematopoietic lineages. However, the limited number of HSCs in either bone marrow or umbilical cord blood limits their widespread use. Hence, there is great interest in developing methods for ex vivo expansion and thus self-renewal of HSCs. Strategies based on nanomaterials or nanotechnologies seem hopeful to expand HSC numbers more efficiently and tune their properties ex vivo. In this chapter, we focus on recent advances in biomaterials-based tissue engineering studies of HSCs, such as hematopoietic niche mimic, functional regulation, and other potential applications. Moreover, we proposed the future scientific issues to be solved in advancing the preclinical translations.

The dysregulation of miRNAs has been increasingly recognized as a critical mediator of cancer development and progression. Here, we show that frequent deletion of the MIR135A1 locus is associated with poor prognosis in primary breast cancer. Forced expression of miR-135a decreased breast cancer progression, while inhibition of miR-135a with a specific miRNA sponge elicited opposing effects, suggestive of a tumor suppressive role of miR-135a in breast cancer. Estrogen receptor alpha (ERα) bound the promoter of MIR135A1 for its transcriptional activation, whereas tamoxifen treatment inhibited expression of miR-135a in ERα+ breast cancer cells. miR-135a directly targeted ESR1, ESRRA, and NCOA1, forming a negative feedback loop to inhibit ERα signaling. This regulatory feedback between miR-135a and ERα demonstrated that miR-135a regulated the response to tamoxifen. The tamoxifen-mediated decrease in miR-135a expression increased the expression of miR-135a targets to reduce tamoxifen sensitivity. Consistently, miR-135a expression was downregulated in ERα+ breast cancer cells with acquired tamoxifen resistance, while forced expression of miR-135a partially resensitized these cells to tamoxifen. Tamoxifen resistance mediated by the loss of miR-135a was shown to be partially dependent on the activation of the ERK1/2 and AKT pathways by miR-135a–targeted genes. Taken together, these results indicate that deletion of the MIR135A1 locus and decreased miR-135a expression promote ERα+ breast cancer progression and tamoxifen resistance. Significance: Loss of miR-135a in breast cancer disrupts an estrogen receptor-induced negative feedback loop, perpetuating disease progression and resistance to therapy. Graphical Abstract: http://cancerres.aacrjournals.org/content/canres/78/17/4915/F1.large.jpg. Cancer Res; 78(17); 4915–28. ©2018 AACR.

Transplantation of hematopoietic stem cells (HSCs) from human umbilical cord blood (hUCB) holds great promise for treating a broad spectrum of hematological disorders including cancer. However, the limited number of HSCs in a single hUCB unit restricts its widespread use. Although extensive efforts have led to multiple methods for ex vivo expansion of human HSCs by targeting single molecules or pathways, it remains unknown whether it is possible to simultaneously manipulate the large number of targets essential for stem cell self-renewal. Recent studies indicate that N6-methyladenosine (m6A) modulates the expression of a group of mRNAs critical for stem cell-fate determination by influencing their stability. Among several m6A readers, YTHDF2 is recognized as promoting targeted mRNA decay. However, the physiological functions of YTHDF2 in adult stem cells are unknown. Here we show that following the conditional knockout (KO) of mouse Ythdf2 the numbers of functional HSC were increased without skewing lineage differentiation or leading to hematopoietic malignancies. Furthermore, knockdown (KD) of human YTHDF2 led to more than a 10-fold increase in the ex vivo expansion of hUCB HSCs, a fivefold increase in colony-forming units (CFUs), and more than an eightfold increase in functional hUCB HSCs in the secondary serial of a limiting dilution transplantation assay. Mapping of m6A in RNAs from mouse hematopoietic stem and progenitor cells (HSPCs) as well as from hUCB HSCs revealed its enrichment in mRNAs encoding transcription factors critical for stem cell self-renewal. These m6A-marked mRNAs were recognized by Ythdf2 and underwent decay. In Ythdf2 KO HSPCs and YTHDF2 KD hUCB HSCs, these mRNAs were stabilized, facilitating HSC expansion. Knocking down one of YTHDF2′s key targets, Tal1 mRNA, partially rescued the phenotype. Our study provides the first demonstration of the function of YTHDF2 in adult stem cell maintenance and identifies its important role in regulating HSC ex vivo expansion by regulating the stability of multiple mRNAs critical for HSC self-renewal, thus identifying potential for future clinical applications.

SUMMARY Tumor-initiating stem cells (TSCs) are critical for drug resistance and immune escape. However, the mutual regulations between TSC and tumor microenvironment (TME) remain unclear. Using DNA-label retaining, single-cell RNA sequencing (scRNA-seq), and other approaches, we investigated intestinal adenoma in response to chemoradiotherapy (CRT), thus identifying therapy-resistant TSCs (TrTSCs). We find bidirectional crosstalk between TSCs and TME using CellPhoneDB analysis. An intriguing finding is that TSCs shape TME into a landscape that favors TSCs for immunosuppression and propagation. Using adenoma-organoid co-cultures, niche-cell depletion, and lineaging tracing, we characterize a functional role of cyclooxygenase-2 (Cox-2)-dependent signaling, predominantly occurring between tumor-associated monocytes and macrophages (TAMMs) and TrTSCs. We show that TAMMs promote TrTSC proliferation through prostaglandin E2 (PGE2)-PTGER4(EP4) signaling, which enhances β-catenin activity via AKT phosphorylation. Thus, our study shows that the bidirectional crosstalk between TrTSC and TME results in a pro-tumorigenic and immunosuppressive contexture., Graphical Abstract, In brief He et al. examined the nature of therapy-resistant tumor-initiating stem cells (TSCs) in murine intestinal adenoma using single-cell RNA sequencing (scRNA-seq) analysis and revealed a bidirectional crosstalk between TSC and its microenvironment, TME, in which TSC shapes TME, especially tumor-associated monocyte and macrophage (TAMM), into an immunosuppressive barrier and pro-tumorigenic niche.

Advanced and recurrent endometrial carcinoma (EC) exhibits a poor response to chemotherapy and low survival rates. It has been previously reported that human prolactin (hPRL) is upregulated in endometrial cancer and is associated with worse survival outcomes. We provide evidence for the functional role of hPRL in EC progression. We generated a model for the study of autocrine hPRL-mediated cell functional effects through the forced expression of hPRL in human EC cells. Autocrine hPRL expression stimulated cell proliferation, anchorage-independent growth, migration, and invasion of EC cells and promoted tumor growth, local invasion, and metastatic colonization in xenograft models. In addition, forced expression of hPRL decreased sensitivity of EC cells to chemotherapeutic drugs (i.e., doxorubicin and paclitaxel), both in vitro and in vivo. Consistently, small interfering RNA-mediated depletion of hPRL significantly reduced oncogenicity and enhanced the chemosensitivity of EC cells. As CD24 is hPRL-regulated and has been implicated in drug resistance in EC, we further showed that CD24 is a critical mediator of hPRL-stimulated reduced sensitivity to doxorubicin and paclitaxel in EC cells. Therefore, inhibition of hPRL signaling is a potential therapeutic strategy for the treatment of late-stage EC, which can be used in combination with chemotherapy to improve the chemotherapeutic response.

Human ELTD1 (Epidermal growth factor, latrophilin and seven-transmembrane domain-containing 1), an orphan G-protein-coupled receptor (GPCR) belonging to the adhesion GPCR family, has been reported as a novel regulator of angiogenesis and a potential anti-cancer therapeutic target. However, little is known about the function of ELTD1, especially its undiscovered ligands. In this experiment, an ELTD1 homozygous knockout human embryonic stem cell line, FAHZUe001-A, was generated by the iCRISPR/Cas9 system to achieve a deeper understanding of ELTD1. The FAHZUe001-A was confirmed with normal karyotype, typical undifferentiated morphology, pluripotency and trilineage differentiation potential in vitro.

// Xiangjun Kong 1, 2, * , Wenyong Wu 3, * , Yan Yuan 1, 2 , Vijay Pandey 4 , Zhengsheng Wu 5 , Xuefei Lu 1, 2 , Weijie Zhang 1, 2 , Yijun Chen 4 , Mingming Wu 1, 2 , Min Zhang 1, 2 , Gaopeng Li 1, 2 , Sheng Tan 1, 2 , Pengxu Qian 1, 2 , Jo K. Perry 6 , Peter E. Lobie 4, 7 , Tao Zhu 1, 2 1 The CAS Key Laboratory of Innate Immunity and Chronic Disease, School of Life Sciences and Medical Center, University of Science and Technology of China, Hefei, Anhui, China 2 Hefei National Laboratory for Physical Sciences at Microscale, Hefei, Anhui, China 3 Department of General Surgery, First Affiliated Hospital of Anhui Medical University, Hefei, Anhui, China 4 Cancer Science Institute of Singapore and Department of Pharmacology, National University of Singapore, Singapore 5 Department of Pathology, Anhui Medical University, Hefei, Anhui, China 6 Liggins Institute, University of Auckland, Auckland, New Zealand 7 National University Cancer Institute of Singapore, National University Health System, Singapore * These authors have contributed equally to this work Correspondence to: Tao Zhu, e-mail: zhut@ustc.edu.cn Peter E. Lobie, e-mail: csipel@nus.edu.sg Keywords: growth hormone, prolactin, hepatocellular carcinoma, oncogenicity, survival Received: May 19, 2015 Accepted: March 24, 2016 Published: April 18, 2016 ABSTRACT The death rates of hepatocellular carcinoma (HCC) are extremely high due to the paucity of therapeutic options. Animal models and anecdotal clinical evidence indicate a potential role of hGH and hPRL in HCC. However, the prognostic relevance and the functional role of tumor expression of these hormones in human HCC are not defined. Herein, we analyzed the mRNA and protein expression of hGH and hPRL in histopathological samples of non-neoplastic liver and HCC by in situ hybridization, PCR and immunohistochemistry techniques. Increased mRNA and protein expression of both hormones was observed in HCC compared with non-neoplastic liver tissues. hGH expression was significantly associated with tumor size and tumor grade. No significant association was observed between the expression of hPRL and any histopathological features. Amplification of both hGH and hPRL genes in HCC was observed when compared to non-neoplastic tissue. Expression of both hGH and hPRL was associated with worse relapse-free and overall survival in HCC patients. In vitro and in vivo functional assays performed with HCC cell lines demonstrated that autocrine expression of hGH or hPRL in HCC cells increased STAT3 activation, oncogenicity and tumor growth while functional antagonism with hGH-G120R significantly reduced these parameters. Hence, tumor expression of hGH/hPRL is associated with a worse survival outcome for patients with HCC and hGH/hPRL function as autocrine/paracrine promoters of HCC progression.

MicroRNAs (miRNAs) typically bind to unstructured miRNA-binding sites in target RNAs, leading to a mutual repression of expression. Here, we report that miR-1254 interacts with structured elements in cell cycle and apoptosis regulator 1 (CCAR1) 5' untranslated region (UTR) and this interaction enhances the stability of both molecules. miR-1254 can also act as a repressor when binding to unstructured sites in its targets. Interestingly, structured miR-1254-targeting sites act as both a functional RNA motif-sensing unit, and an independent RNA functional unit that enhances miR-1254 expression. Artificially designed miRNA enhancers, termed "miRancers", can stabilize and enhance the activity of miRNAs of interest. We further demonstrate that CCAR1 5' UTR as a natural miRancer of endogenous miR-1254 re-sensitizes tamoxifen-resistant breast cancer cells to tamoxifen. Thus, our study presents a novel model of miRNA function, wherein highly structured miRancer-like motif-containing RNA fragments or miRancer molecules specifically interact with miRNAs, leading to reciprocal stabilization.

The mammalian imprinted Dlk1-Gtl2 locus produces multiple non-coding RNAs (ncRNAs) from the maternally inherited allele, including the largest miRNA cluster in the mammalian genome. This locus has characterized functions in some types of stem cell, but its role in hematopoietic stem cells (HSCs) is unknown. Here, we show that the Dlk1-Gtl2 locus plays a critical role in preserving long-term repopulating HSCs (LT-HSCs). Through transcriptome profiling in 17 hematopoietic cell types, we found that ncRNAs expressed from the Dlk1-Gtl2 locus are predominantly enriched in fetal liver HSCs and the adult LT-HSC population and sustain long-term HSC functionality. Mechanistically, the miRNA mega-cluster within the Dlk1-Gtl2 locus suppresses the entire PI3K-mTOR pathway. This regulation in turn inhibits mitochondrial biogenesis and metabolic activity and protects LT-HSCs from excessive reactive oxygen species (ROS) production. Our data therefore show that the imprinted Dlk1-Gtl2 locus preserves LT-HSC function by restricting mitochondrial metabolism.

Transplantation of hematopoietic stem cells (HSCs) from human umbilical cord blood (hUCB) holds great promise for treating a broad spectrum of hematological disorders including cancer. However, the limited number of HSCs in a single hUCB unit restricts its widespread use. Although extensive efforts have led to multiple methods for ex vivo expansion of human HSCs by targeting single molecules or pathways, it remains unknown whether it is possible to simultaneously manipulate the large number of targets essential for stem cell self-renewal. Recent studies indicate that N(6)-methyladenosine (m(6)A) modulates the expression of a group of mRNAs critical for stem cell-fate determination by influencing their stability. Among several m(6)A readers, YTHDF2 is recognized as promoting targeted mRNA decay. However, the physiological functions of YTHDF2 in adult stem cells are unknown. Here we show that following the conditional knockout (KO) of mouse Ythdf2 the numbers of functional HSC were increased without skewing lineage differentiation or leading to hematopoietic malignancies. Furthermore, knockdown (KD) of human YTHDF2 led to more than a 10-fold increase in the ex vivo expansion of hUCB HSCs, a fivefold increase in colony-forming units (CFUs), and more than an eightfold increase in functional hUCB HSCs in the secondary serial of a limiting dilution transplantation assay. Mapping of m(6)A in RNAs from mouse hematopoietic stem and progenitor cells (HSPCs) as well as from hUCB HSCs revealed its enrichment in mRNAs encoding transcription factors critical for stem cell self-renewal. These m(6)A-marked mRNAs were recognized by Ythdf2 and underwent decay. In Ythdf2 KO HSPCs and YTHDF2 KD hUCB HSCs, these mRNAs were stabilized, facilitating HSC expansion. Knocking down one of YTHDF2′s key targets, Tal1 mRNA, partially rescued the phenotype. Our study provides the first demonstration of the function of YTHDF2 in adult stem cell maintenance and identifies its important role in regulating HSC ex vivo expansion by regulating the stability of multiple mRNAs critical for HSC self-renewal, thus identifying potential for future clinical applications.