Background: High-dose chemotherapy combined with autologous stem cell transplantation (ASCT) remains pivotal in the treatment of multiple myeloma (MM) and lymphoma. The safety and efficacy of ASCT, as well as the quantity of stem cells harvested during apheresis, are heavily reliant on the efficacy of peripheral blood stem cell (PBSC) mobilization. In China, chemotherapy coupled with G-CSF mobilization is the predominant strategy. However, the influence of febrile neutropenia (FN) during chemomobilization on CD34+ PBSC collection and the risk of post-transplant infection in MM and lymphoma patients undergoing ASCT is still a matter of debate. Methods: This multi-center, real-world, retrospective study was conducted at Tongji Hospital in Wuhan, Shanxi Bethune Hospital, and Shanxi Province Cancer Hospital from October 1st 2010 to January 31st 2024. We reviewed the records of 148 MM and 130 lymphoma patients, all aged 18 years or older, who underwent ASCT within this period. All participants received chemotherapy followed by G-CSF stimulation for stem cell mobilization. For MM patients, the mobilization regimens primarily involved the cyclophosphamide (CTX) protocol and the etoposide protocol. For lymphoma patients, the regimens mainly included the DHAP protocol, the CTX protocol, and the etoposide protocol. The study received ethics approval from the Ethics Committees of Tongji Hospital, Shanxi Bethune Hospital, and Shanxi Province Cancer Hospital, adhering to the ethical guidelines of the Declaration of Helsinki. The impact of FN during chemomobilization on stem cell collection and post-transplant infection was analyzed using GraphPad Prism 9, with significance set at P < 0.05. Results: Patients were categorized into FN and non-FN groups. MM patients in the FN group required more than 5 days of G-CSF stimulation (P < 0.001) and more than 2 days of apheresis (P = 0.007) to achieve the target CD34+ cell count of 2 × 106/kg. The median CD34+ counts were significantly lower in the FN group (3.75 × 106/kg versus 6.22 × 106/kg, P < 0.001). Additionally, these patients exhibited lower good-mobilization rates (P < 0.001) and prolonged times to neutrophil (P = 0.006) and platelet engraftment (P = 0.03). In lymphoma patients, those in the FN group required more than 2 days of apheresis (P = 0.001) and experienced longer times to platelet engraftment (P = 0.02). FN was associated with an increased risk of pre-engraftment infection (P = 0.03) in lymphoma patients. Univariate analysis revealed that more than 5 days of G-CSF stimulation (P = 0.004), more than 2 days of apheresis (P = 0.01) and FN during mobilization (P < 0.001) were significant risk factors impacting good mobilization in MM patients. FN during mobilization was identified as a risk factor for reduced good mobilization in MM patients (univariate P < 0.001; multivariate OR = 3.06, 95% CI = 1.22-8.03, P = 0.02) and for pre-engraftment infection in lymphoma patients (OR = 0.29, 95% CI = 0.10-0.86, P = 0.03). Further scrutiny into the long-term survival implications revealed that FN during chemomobilization might impact the long-term survival of MM patients (P = 0.002). The 5-year overall survival in FN and non-FN subgroups of MM patients was 61.01% and 84.36%, respectively. Conclusions: Febrile neutropenia during chemomobilization has a detrimental effect on stem cell mobilization in multiple myeloma patients and heightens the risk of pre-engraftment infections in lymphoma patients undergoing ASCT. These findings underscore the necessity for improved strategies to manage FN in these patient populations to enhance ASCT outcomes.

The role of the compromised immune microenvironment, including immune checkpoints, in myelodysplastic syndromes (MDS) has been identified as critical This study aimed to investigate the expression patterns of immune checkpoints, particularly soluble PD-1/PD-L1 (sPD-1/sPD-L1) as well as PD-1 on effector T cell subsets, and assess their prognostic value and potential regulatory roles in MDS. 161 MDS patients were enrolled, including 129 patients were primarily diagnosed with de novo MDS, together with 59 MDS patients who underwent hypomethylating agents (HMAs) therapy. Plasma sPD-L1 level was elevated in newly diagnosed MDS patients, which was also found to be associated with MDS disease progression that further increase in higher IPSS-R score group. Patients with increased sPD-L1 expression at diagnosis exhibited notably poorer overall survival, and multivariate Cox analysis indicated that elevated sPD-L1 was an independent risk factor. Furthermore, the levels of multiple cytokines and membrane-bound PD-1 on T cells were found to correlate with sPD-1/sPD-L1 levels in plasma. Importantly, we also found sPD-L1 levels significantly increased in MDS patients who showed progression of disease following HMAs therapy. In conclusion, we found elevated plasma sPD-L1 levels in MDS patients are associated with disease progression and poorer overall survival. This study showed that sPD-L1 is a potential biomarker for prognosis and a target for immunotherapy in MDS.

Background: Plasmacytoid dendritic cells (pDCs), being a primary source of natural type I interferon (IFN-I), have pivotal roles in orchestrating the immune response. Typically, only less than 1% pDCs can be detected in the total bone marrow or peripheral blood nuclear cells. However, the uncontrollable proliferation of transformed pDCs is a remarkable characteristic of blastic plasmacytoid dendritic cell neoplasm (BPDCN). Recently, an abnormal amplification of pDCs in myeloid neoplasms has also been reported in MDS/AML patients (pDC-AML/MDS). This abnormal increase in pDCs could potentially promote disease progression. In this study, we utilized single-cell RNA-sequencing (scRNA-seq) to examine the characteristics of pDCs in pDC-AML. Method: Our study employed a mixed retrospective and prospective design. 85 patients diagnosed with PDC-AML/MDS/BPDCN, including 1 pDC-MDS, 17 pDC-AML, 50 non-pDC-AML (AML patients without pDCs expansion), 17 BPDCN from Tongji Hospital of Huazhong University of Science and Technology in Wuhan, China, between July 2014 and June 2023. scRNA-seq using 10x Genomics platform were performed on bone marrow (BM) samples from 8 pDC-AML patients and 1 BPDCN patient. Furthermore, we expanded our dataset by integrating publicly available scRNA-seq data of BM samples from 5 BPDCN patients, 5 non-pDC-AML patients, and 5 healthy donors (HD). Result: The median pDCs proportion of pDC-AML/MDS was 8.0% (range 2.5-52.0%). As of June 2023, The median overall survival (OS) of patients with pDC-MDS/AML was 10.5 months shorter than non-pDC-AML (21.5 months; P＜0.05). In addition, the prognosis of pDC-AML/MDS patients can only be significantly improved after receiving allogenic hematopoietic stem cell transplantation, showing a similar OS to that of non-pDC-AML patients. To further clarified the role of pDCs in these patients, we performed scRNA-seq and acquired 126,693 cells across all samples (8 pDC-AML, 6 BPDCN, 5 non-pDC-AML and 5 HD), and identified 13 clusters based on their transcriptional profile in the overall UMAP (Figure 1 A). Firstly, our single-cell sequencing data revealed significant differences in the proportions of bone marrow pDCs across pDC-AML, HDs, non-pDC-AML, and BPDCN patients. Notably, pDC proportions were significantly elevated in pDC-AML and BPDCN patients. Further differential expression analysis revealed striking differences in the transcriptomes of pDCs across these four groups. When comparing pDC-AML to HDs, we identified 801 upregulated and 150 downregulated genes. Upon comparing pDC-AML to BPDCN, we found 1110 upregulated and 765 downregulated genes, and when comparing pDC-AML to non-pDC-AML, we observed 1674 upregulated and 945 downregulated genes. To explore the functional implications of these differentially expressed genes (DEGs), these findings were explored further through Gene Set Enrichment Analysis (GSEA), and indicated a heightened response to interferon signaling within the pDC cells of pDC-AML patients. Concurrently, we also discovered an increased expression of gene sets associated with tumorigenic pathways (such as KRAS, PTEN, p53, RAPA, etc.) and those related to embryonic stem cell states in pDCs of pDC-AML. Drawing on our findings and existing research, we hypothesize that interferon signaling might play a pivotal role in the pathogenesis of pDC-AML. This could be achieved through the enhancement of cell proliferation and self-renewal capabilities, thereby facilitating disease progression. This hypothesis is further supported by our pseudotime analysis. After conducting a more detailed subgroup classification of pDC-AML patients, we found a developmental trajectory linking AML_HSC and pDCs specific to pDC-AML patients. Notably, such a correlation was not observed in the non-pDC-AML patient group. Conclusions: Our study revealed heterogeneity and underscores the crucial role of pDCs in pDC-AML, as we found that these patients had worse OS. We observed upregulation of interferon signaling and tumorigenic pathways in these pDCs. Moreover, pseudotime analysis suggested a potential developmental relationship between pDCs and AML_HSCs. These findings provide invaluable insights for further investigations into pDCs biological properties in pDC-AML pathology and therapy design.

Background: Myelodysplastic syndromes (MDS) are heterogeneous hematological stem cell malignancies with a complex pathophysiology involving immune, genetic, and epigenetic changes induced by aging, oxidative stress, and genotoxicity. Dysregulated immune responses and aberrant inflammatory signaling have been identified as critical drivers of MDS development. Recent research has highlighted the significant association of the IL-33/ST2 signaling axis with tumorigenicity and tumor immunity in various cancers. Soluble ST2 (sST2), a transmembrane-free receptor induced by IL-33, negatively regulates the IL-33/ST2 signaling pathway, and elevated levels of sST2 have been reported to correlate with poor prognosis in malignant tumors. However, the specific level of soluble ST2 and the precise role of the IL-33/ST2 signaling axis in the context of MDS remain uncertain and require further investigation. Method: This prospective study enrolled 89 newly diagnosed, untreated outpatient individuals with MDS to assess their clinical prognostic risk using the International Prognostic Scoring System-Revised (IPSS-R) and the International Prognostic Scoring System-Molecular (IPSS-M). A control group of 45 healthy donors (HD) was also included. The levels of various cytokines (sST2, IL-33, IL-2Rα, IL-6, IL-15) were measured using ELISA. Result: Our study revealed that sST2 levels in MDS patients were significantly higher compared to HD (22.18 vs. 12.43 ng/ml, p = 0.0313), with no significant difference between low and high-risk groups. The Receiver Operating Characteristic (ROC) curve analysis showed an Area Under the Curve (AUC) of 0.6140 for sST2 (p = 0.0315, 95%CI 0.5199-0.7081). Using the maximum Youden index, the optimal cut-off value for sST2 (>14.71 ng/ml) was determined with a sensitivity and specificity of 51.69% and 73.33%, respectively. Progressive events include conversion to leukemia(n=8), transfusion-dependent occurrence(n=1) and death (n=29). Patients with higher sST2 levels in both high and low-risk groups demonstrated poorer Event-Free Survival (EFS) (p = 0.0193) and Overall Survival (OS) (p = 0.0130) (Figure 1). Further, multivariate analysis identified cytokine sST2(HR = 0.337, 95%CI 0.163-0.696, P=0.003), blast percentage(HR = 0.472, 95%CI 0.237-0.940, P = 0.033), and transfusion dependence(HR = 0.367, 95%CI 0.179-0.753, P = 0.006) as independent risk factors, indicating sST2's potential in predicting prognosis and adverse event risk in MDS patients. Furthermore, sST2 exhibited a significant correlation with other inflammatory cytokines (IL-2Rα, r = 0.4152, p = 0.0005; IL-6, r = 0.4897, p = 0.0013; IL-15, r = 0.3535, p = 0.0253). This suggests that sST2 plays an active role in immune system activation and chronic inflammation. Multiple attempts to detect IL-33 expression in the serum of MDS patients were unsuccessful, indicating the need for a more sensitive and specific assay. In our study, no statistically significant difference was observed in the level of sST2 between bone marrow and peripheral blood (P=0.2197), as well as before treatment and after chemotherapy (P=0.3965). These findings indicate that sST2 is an easily detectable and relatively stable indicator. Conclusions: sST2 emerges as a significant, independent negative prognostic factor at the time of MDS diagnosis, offering valuable predictive insights into disease progression and a poor prognosis.

Background: Chimeric antigen receptor (CAR) T-cell therapy carries the risk of inducing severe and life-threatening toxicities such as cytokine release syndrome (CRS), neurotoxicity, and infection. Although CRS and infections have similar symptoms, their treatment strategies differ, and early diagnosis is very important. For CRS and infections, the fastest detection time currently takes more than 24 h, so a quick and simple method to identify a fever after CAR T-cell infusion is urgently needed. Methods: We enrolled 27 patients with recurrent fever treated with different types of CAR T-cells, including cluster of differentiation (CD) 7, CD19, CD22, and CD19-CD22 bicistronic CAR T-cells, and evaluated the infection events occurring in these patients. We detailed the morphology of CAR T-cells in peripheral blood smears (PBS) and reported the infection events, CAR transgene copy number, and inflammatory indicators within the first month after treatment. Results: Similar morphological characteristics were observed in the PBS of different CAR T-cells, namely, enlarged cell bodies, deep outside and shallow inside basophilic blue cytoplasm, and natural killer (NK) cell-like purplish red granules. There were ten infections in nine of the twenty-seven patients (33%). The percentage of atypical lymphocytes in PBS was significantly associated with CAR transgene copy number and absolute lymphocyte count in all patients. The atypical lymphocyte percentage was significantly higher in the non-infection group. Conclusions: In conclusion, the unique morphology of CAR T-cells in PBS can be used to evaluate CAR T-cell kinetics and provide reliable evidence for the rapid early identification of fever after CAR T-cell infusion. Clinical Trial Registrations: ChiCTR-OPN-16008526; ChiCTR-OPN-16009847; ChiCTR2000038641; NCT05618041; NCT05388695.

Plasmacytoid dendritic cells (pDCs) are a specific dendritic cell type stemming from the myeloid lineage. Clinically and pathologically, neoplasms associated with pDCs are classified as blastic plasmacytoid dendritic cell neoplasm (BPDCN), mature plasmacytoid dendritic myeloid neoplasm (MPDMN) and pDC expansion in myeloid neoplasms (MNs). BPDCN was considered a rare and aggressive neoplasm in the 2016 World Health Organization (WHO) classification. MPDMN, known as mature pDC-derived neoplasm, is closely related to MNs and was first recognized in the latest 2022 WHO classification, proposing a new concept that acute myeloid leukemia cases could show clonally expanded pDCs (pDC-AML). With the advances in detection techniques, an increasing number of pDC expansion in MNs have been reported, but whether the pathogenesis is similar to that of MPDMN remains unclear. This review focuses on patient characteristics, diagnosis and treatment of pDC expansion in MNs to gain further insight into this novel and unique provisional subtype.

Background: Myelodysplastic syndromes (MDS) consist of a spectrum of myeloid malignancies typified by regular genetic abnormalities, clonal hematopoiesis, dysfunctional myelopoiesis, inefficient blood cell production, peripheral-blood cytopenia, and an elevated risk for progressing to acute myeloid leukemia (AML). Mounting evidence suggests that the dysregulation of immune checkpoints (ICs) plays critical roles in immune evasion of MDS, which thus led to a series of trials using HMAs in combination with immune checkpoints inhibitors (ICIs). Of these, programmed cell death-1 (PD-1)/programmed cell death ligand-1 (PD-L1) pathway has been studied most extensively. Recently, forms of cell-free soluble PD-1/PD-L1 (sPD-1/sPD-L1) detection in plasm of patients with solid tumors has open a new paradigm for PD-1/PD-L1 investigations. However, within the context of MDS, the potential dysregulation and prognostic impact of sPD-1/sPD-L1 remains uncertain. This study was aimed to assess roles of sPD-1/sPD-L1 in newly diagnosed MDS patients. Method: Between July 2020 and March 2023, 130 MDS/sAML patients matched the inclusion criteria and 55 healthy individuals with no familial or personal history of autoimmune or cancer disorders were included. The criteria for patient inclusion included: (1) a diagnosis of MDS confirmed pathologically according to 2016/2022 World Health Organization (WHO) classification, or sAML patients with a verified prior MDS history; (2) absence of other conditions like autoimmune diseases or different types of tumors; (3) availability of complete clinical information and survival data. MDS patients were categorized by IPSS-R as: lower-risk (very low risk/ low risk; higher-risk (intermediate risk/high risk/very high risk. Follow-up was conducted until June 16th, 2023. The median follow-up was 14.32 months (range 3.2-34.63 months). Profile of sICs (sPD-L1, sPD-1, sCTLA-4, sGITR, sTIM-3, sOX40, sLAG-3, s4-1BB, sICOS) were tested by ELISA, cytokines (IFN-γ, IFN-α2, IL-2, IL-2α, IL-6, IL-7, IL-10, IL-15, IL-17, TNF-α) were detected using Luminex analysis, and the expression levels of PD-1 on T cells (CD3 +, CD4 +, CD8 +, DPT, DNT）were measured using flow cytometry (FCM). Result: In the plasma of 98 newly diagnosed MDS patients, sPD-L1 levels (median 66.80 pg/ml, range 25.00-219.5 pg/ml) were significantly elevated compared to 55 healthy blood donors (median 49.22 pg/ml, range 12.5-218.1 pg/ml). sPD-L1 level was higher in higher-risk IPSS-R groups ( P=0.0380). In addition, higher sPD-L1 levels also showed a correlation with lower hemoglobin concentration, implying a potential link between higher sPD-L1 levels and a more advanced MDS disease state. The AUC for sPD-1 was found to be 0.6333 [95% CI 0.5331-0.7334, P=0.0100] and for sPD-L1, it was 0.6785 [95% CI 0.5836-0.7733, P=0.0002]. The optimal cut-off values were established at 187.5 pg/mL for sPD-1 and 55.08 pg/mL for sPD-L1. Based on these cut-off values, patients with increased sPD-L1 experienced shorter OS ( P=0.0305). By using multivariate Cox model, high expression of sPD-L1 ( P=0.019, HR = 4.172, 95% CI: 1.265-13.755) and high IPSS-R scores ( P=0.030, HR = 2.898, 95% CI: 1.111-7.565) were found to be independent risk factors for newly diagnosed MDS patients. In addition, compared to healthy controls, the PD-1 + T cell subpopulations in MDS patients were significantly reduced, sPD-1 levels were positively correlated with proportion of CD4 + PD-1 + T cells, absolute count of CD4 + PD-1 + T cells and PD-1 + DPT cells, suggesting that CD4 + T cells and DPT cells may be potential sources of sPD-1 in plasma. Subsequently, we analysed the potential roles of cytokines and HMAs in sPD-1/sPD-L1 regulation. It was found that sPD-1 levels had a positive correlation with sPD-L1, IL-2Rα, IP-10, MIG, TNFα, GROα, and TRAIL. sPD-L1 was positively correlated with IL-2Rα, IP-10, MCP-1, MIG, MIP1α, and SCF (Figure 1 A). Notably. We found a reduced sPD-L1 levels in MDS patients who achieved remission after HMA treatment compared to treatment-naïve patients. ( P=0.0302) (Figure 1 B). Conclusions: In conclusion, we found sPD-L1 is an independent risk factors for overall survival in newly diagnosed MDS patients. The elevation of plasma sPD-L1 levels is associated with disease progression in MDS, and may be a potential target for MDS immunotherapy.