bluuebunny/crossref_metadata_2025 huggingface dataset:
Daratumumab Maintenance after Autologous Hematopoietic Stem Cell Transplantation for Newly Diagnosed Multiple Myeloma
Authors: Ma Yi , Xiao Xiubin , Yuan Shunzong , Chen Xilin , Zhao Shihua , Cheng Nana , Lu Yun , Chen Junli , Wang Yueqi , Huang Wenrong
Published: December 2023

Background: Maintenance therapy after autologous hematopoietic stem cell transplantation (ASCT) is a standard of care in newly diagnosed multiple myeloma (NDMM). Lenalidomide maintenance after transplantation is the most frequently recommended regimen. Daratumumab has been increasingly used as a maintenance drug in NDMM. However, there is no enough clinical information about daratumumab as maintenance strategy after ASCT. This study is aim to evaluate the efficacy and safety of daratumumab as maintenance after ASCT for NDMM.

Patients and Methods: Clinical data, hematological and renal response, and safety of 15 post-transplant patients with NDMM who had received daratumumab maintenance between 1 May 2022 and 30 June 2023 were retrospectively analyzed. Daratumumab-naive patients received weekly dosing (16mg/kg) for the first 8 weeks.

Results: Fifteen patients (11 males and 4 females) with median age of 58 years (41～72 years) , were included. Thirteen patients were daratumumab native, 6 patients had renal impairment, and 2 patients with high-risk cytogenetics. The median infusion times of daratumumab were 12 (6～17) times. Efficacy was evaluable in all 15 patients，and daratumumab maintenance therapy increase the rate of stringent complete response (from 6 [40%] to 9 [60%] ) by 20%，improved the renal response rate and median eGFR in patients with renal injury. During maintenance therapy with daratumumab, the most common hematological grade 3 adverse events(AE) were lymphopenia (4 [26.67%] of 15 patients), non-hematologic adverse events were infusion-related reactions (7 [46.67%] of 15 patients) and grade 3 pneumonia (5 [33.33%] of 15 patients). These 5 pneumonia patients were daratumumab naive ( 5 [38.46%] of 13 patients) , with median 8 (6～10)infusions. Among them, 3 patients chest CT showed interstitial infiltrates, and combination of methylprednisolone was effective. With a median follow-up of 12 months, one patient had died (not related to daratumumab treatment), 1-year OS rate was 93.33%.

Conclusions: Daratumumab was safe and effective as a maintenance agent for post-ASCT patients with NDMM, and adverse events were controllable. The most common AE is grade 3 pneumonia, maybe a less dose-intense maintenance regimen for the first 8 weeks can reduce the incidence of pneumonia.
The Significance of a “MGUS” Tumor Board
Authors: Mazzoni Sandra , Khouri Jack , Williams Louis S. , Faiman Beth M. , Mathur Saveta , Scott Cynthia
Published: December 2023

Our large academic medical center consists of both a main campus and several regional sites that all offer hematology/oncology services where monoclonal gammopathy cases are referred for consultation. We receive well over 100 requests annually to see cases with abnormal monoclonal proteins. The challenge was how to triage these requests, with the goal of improving patient access to main campus myeloma experts while improving patient care and timely management. We decided to schedule cases initially referred for monoclonal gammopathy of undetermined significance (MGUS) with our advanced practice providers (APPs) both in the region and at main campus.

Due to the concern for varying experience managing MGUS, differences in diagnostic practice and limited guidance we developed a novel concept to have a tumor board to review MGUS cases that were initially seen by our APPs. The “MGUS” tumor board is ran by three of our main campus myeloma focused hematologists and our myeloma dedicated APRN-PhD. We meet twice a month to review cases.

Over the past 2 years we reviewed 147 cases providing recommendations for further work up. Of these, 41 cases (28%) were found to have a clinically significant monoclonal protein necessitating referral to a physician at main campus. The breakdown of these cases are detailed below in T able 1. Sixteen patients (11%) were started on treatment, the other 25 (17%) remained on observation.

Another aspect of the tumor board is to provide education regarding clinically significant scenarios that involve monoclonal proteins. This tumor board offers CME credit for both APPs and physicians. By developing a team of knowledgeable APPs we have shifted 72% of monoclonal gammopathy referrals to remain under the care of APPs which improved access to main campus myeloma physician experts. This allowed the myeloma team at main campus to see more second opinions, open more clinical trials and expand our cell-therapy options for patients. Our referral base has grown and we have hired more APPs in the region and two additional myeloma physicians at main campus since starting the “MGUS” tumor board.

Overall, we were successful in transitioning “MGUS” referrals to our APPs, but there were higher than expected percent of patients with clinically significant monoclonal proteins. Our data shows that it is essential for there to be some standardization for reviewing and working up “MGUS” referrals. This is especially important at large centers with multiple regional locations. We recommend a tumor board style review as it allows for discussion and education of “MGUS” referrals which prove to be far more complex in many cases.
Compared to Venetoclax Plus Hypomethylating Agents, High Dose Cytarabine Based Intensive Chemotherapy Induction for Newly Diagnosed Acute Myeloid Leukemia (AML) Leads to a Faster Remission Resulting in Better Survival
Authors: Solh Melhem M. , Solomon Scott R. , Morris Lawrence , Holland H Kent , Bachier-Rodriguez Lizamarie , Zhang Xu , Jackson Katelin C , Bashey Asad
Published: December 2023

Hypomethylating agents plus venetoclax is a commonly used induction regimen for newly diagnosed AML patients who are not eligible for standard intensive chemotherapy (IC). Due to the high remission rates and the easier administration of Venetoclax plus hypomethylating agents in the outpatient settings and community practices, some patients receive such induction even if IC eligible. We retrospectively analyzed 206 newly diagnosed AML patients, age 56-80 years who received either Venetoclax plus hypomethylating agents (n=65) or high dose cytarabine based IC ( n=141) to assess the difference in remission rates, time to achieve remission and the impact of time to achieving remission on survival endpoints.

Patients who received IC ( FLAG or CLAG +/- Idarubicin) were younger ( median age 66 (56,79) vs 73(56,79) years, p<0.001) but had similar race distribution, NCCN cytogenetics risk and distribution of primary versus secondary leukemia compared to patients with hypomethylating agents plus venetoclax. Patients who received IC has similar chance of achieving CR (89% vs 71%, p=0.89) and transplant rate compared to venetoclax/hypomethylating agents. Patients who received IC had better DFS (p=0.026), a trend towards better OS (p=0.06) and similar relapse risk (p=0.69) compared to venetoclax/hypomethylating agents (figure 1). In a logistic regression analysis, patients who received IC and patients with favorable/intermediate risk AML had higher likelihood of achieving CR in the first 60 days post induction ( HR 2.34, 95% CI 1.19-4.61; P=0.01 for IC versus Venetoclax/hypometh; and HR 0.24 , 95% CI 0.12-0.49, P<0.001 for NCCN adverse versus favorable/intermediate risk AML). A cox model for survival endpoints where age was factored into three equal size groups (56-63, 64-70 and 71-79) and NCCN risk fitted as strata, was performed on survival endpoints. Achieving CR1 within 60 days of induction treatment had significant impact on overall survival (HR 0.24, 95% CI 0.13-0.46, p<0.001).

Our data shows that among patients aged 55-80 years with newly diagnosed AML, receiving high dose cytarabine based IC have similar rates but more likely to achieve CR within 60 days than venetoclax plus hypomethylating agents. Achieving CR within 60 days of induction is a predictor of better overall survival. Based on our analysis, high dose cytarabine based IC should remain the preferred choice for patients eligible for such treatment.
Integrating AI and ML in Myelodysplastic Syndrome Diagnosis: State of the Art and Future Prospects
Authors: Elshoeibi Amgad , Badr Ahmed , Elsayed Basel , Metwalli Omar , Elhadary Mohamed , Alhuraiji Ahmad , Cherif Honar , Khadadah Fatima , Yassin Mohamed A
Published: December 2023

Introduction:

Myelodysplastic syndrome (MDS) is a diverse group of hematological malignancies characterized by dysfunctional pluripotent stem cells, leading to abnormal hematopoiesis and cytopenia. Early diagnosis is crucial for improving patient outcomes. Artificial Intelligence (AI) and Machine Learning (ML) have emerged as powerful tools in healthcare for disease diagnosis and monitoring. This review aims to summarize the current state of AI application in the diagnosis of MDS, discussing various ML models, their advantages, disadvantages, and performance metrics.

Methods:

A comprehensive search strategy was developed using Medical Subject Headings (MeSH) terms and relevant keywords related to MDS and AI/ML. Pubmed, Embase, and scopus were searched, and duplicates were removed. Only original research articles studying ML algorithms in the diagnosis of MDS in human subjects were included. Studies on animals, reviews, non-original articles, and non-English papers were excluded. For each article, data on the area under the receiver operating curve, sensitivity, specificity, and accuracy for the best preforming model were collected.

Results:

The search yielded 313 articles, and after screening, 16 articles met the inclusion criteria. These articles were grouped based on the data utilized in the training of their models. Five articles utilized bone marrow smears for the training of their data. Their models aimed to detect dysplastic cells and blasts in bone marrow smears. The majority of these studies utilized convolutional neural network (CNN), a ML algorithm, for image recognition. One study utilized decision tree models. The AUC of these models ranged between 0.8 and 0.996. All the models were internally validated but were not externally validates. Three studies utilized peripheral blood smears for the training of their ML models. Two of these studies also utilized CNN for the detection of dysplasia and subsequent diagnosis of MDS. One study utilized CART to create a decision tree algorithm. The AUC for these models ranged between 0.982-0.99. Four studies utilized flowcytometry data from bone marrow samples for the training of their AI models. These models achieved AUC ranging between 0.935-0.97. Two of these models were both internally and externally validated, one of which was on a prospective cohort.

Conclusion:

The integration of AI and ML in MDS diagnosis holds promise for early detection and improved patient care. The reviewed studies showed encouraging results, indicating that AI-based approaches can complement traditional diagnostic methods and aid in making more informed clinical decisions especially when it comes to analysing blood/bone smears. However, many of these models need to be externally validated in large prospective cohorts to establish AI's role in routine clinical practice.
Congenital Factor VII Deficiency : A Retrospective Analysis of 14 Cases from One Hospital
Authors: Zhou Rongfu , Xia Mao , Yuan Yanhui , Shao Xiaoyan , Xu Yuyi , Chen Bing
Published: December 2023

Objective To explore the clinical characteristics, laboratory findings,diagnosis, treatment, and prognosis of congenital factor VII (FVII) deficiency. Methods Clinical data of 14 patients of congenial FVII deficiency diagnosed from Jan 2019 to Jun 2022 were retrospectively analyzed. Results Of the 14 patients,there were 8 males and 6 females with median age 31.5 (1.3~70) years. Family history was found in 2 cases. There were 7 cases with bleeding symptoms, of which were mucocutaneous bleeding (1 case), gingival bleeding (2 cases), menorrhagia (3 cases), hemoptysis (1 case ), post-surgery hemorrhage (1 case), and 1 case with cerebral infarction. Menorrhagia occurred in 3 cases (75% of female patients who were in fertile age). Laboratory findings were characterized by significantly prolonged prothrombin time (PT) (19.2~58.9s) , of which could be corrected by mixed normal plasma, normal partial thromboplastin time (APTT), and decreased FVII activity (FVII:C) (2.4~24.7%). 1 case and his family members received gene mutation analysis and compound Heterozygous mutation (heterozygous deletion mutation and heterozygous missense mutation) was found in the proband. 3 cases were treated with prothrombin complex concentrates (PCC), 1 case with PCC and human recombinant activated FVII (rhFVIIa). 10 patients with no or mild bleeding symptoms did not receive any replacement therapy. Conclusion Most patients with congenital FVII deficiency have mild or no bleeding symptom, but have a tendency to excessive bleeding after surgery or trauma. There is no significant correlation between FVII:C and the severity of bleeding symptoms. Prophylaxis could be applied in patients with severe bleeding symptom. Gene mutation test is significant for diagnosis, prognosis prediction of the disease.
The Correlation between Alterations in the Intestinal Microbiome and Early Outcomes of Autologous Hematopoietic Cell Transplantation in Diffuse Large B-Cell Lymphoma
Authors: Sheng Lixia , WANG Jiaping , Lin Zhouning , Lai Yanli , Ouyang Guifang
Published: December 2023

Introduction: Prior research has documented a decline in the diversity of fecal microbiota among individuals with diffuse large B-cell lymphoma (DLBCL) who undergo autologous hematopoietic cell transplantation (auto-HCT). Nevertheless, the potential influence of gut microbiota on hematopoietic reconstitution, immune reconstitution, and early complications in this specific patient cohort remains uncertain.

Methods: A total of 42 fecal samples were obtained from 21 adult individuals who underwent autologous hematopoietic cell transplantation (auto-HCT) for diffuse large B-cell lymphoma (DLBCL), both prior to and two weeks following allogeneic stem cell transplantation (ASCT). The diversity of fecal microbiota was assessed using 16S ribosomal gene sequencing. Additionally, correlations between alterations in the fecal microbiome and clinical characteristics, as well as host immune indices, were examined.

Results: The beta diversity index of intestinal microecology exhibited a significant decrease following transplantation, whereas the alpha diversity index remained unaltered. Prior to transplantation, the abundance of g_Prevotella_7, g_Sutterella, and g_Enterococcus demonstrated a positive correlation with intestinal infection caused by Carbapenem-resistant Enterobacteriaceae. Conversely, the abundance of g_Blautia prior to transplantation and g_Lachnospira following transplantation exhibited a negative correlation with the duration of post-transplant fever. The abundance of g_Enterocococcus, g_Veillonella, g_Sutterella, g_Prevotella_7, and g_Streptococcus following transplantation exhibited a positive correlation with the occurrence of diarrhea. Conversely, the abundance of g_Weissella displayed a negative correlation with the duration of neutrophil and megakaryocyte engraftment, while g_Prevotella_7 exhibited a positive correlation with the duration of neutrophil engraftment. Additionally, the abundance of g_Sutterella and g_Prevotella_7 prior to transplantation, as well as the abundance of g_Lachnoclostridium, g_Blautia, g_Lachnospira, g_Veillonella, and g_Sutterella following transplantation, were positively correlated with host immunity.

Conclusions: The intestinal microbiome experienced significant fluctuations throughout the autologous stem cell transplantation (ASCT) procedure in patients with diffuse large B-cell lymphoma (DLBCL), and these changes exhibited a strong correlation with both early prognosis and post-transplant immune reconstitution. Consequently, additional investigation is necessary to establish strategies for modulating the gut microbiota and enhancing the efficacy of ASCT in DLBCL patients.
Downregulation of Mitochondrial Complex II (MC II) in Myelodysplastic Syndromes
Authors: Sezaki Maiko , Hashimoto Michihiro , Yokota Asumi , Salomonis Nathan , Grimes H. Leighton , Huang Gang
Published: December 2023

Myelodysplastic syndromes (MDS) are a group of diseases of hematopoietic stem cells (HSCs) that occur in the aging population and following therapy for unrelated cancers. The standard of care for MDS involves hypomethylating agents (HMAs) that invariably result in resistance and disease progression. We previously determined the role of mitochondrial metabolism in MDS pathophysiology, whereby suppression of mitochondrial activity leads to the accumulation of oncometabolites that competitively inhibit α-ketoglutarate-dependent dioxygenases (2OGDDs). Consequently, hypermethylation of epigenomes and hypoxia-inducible factor 1A (HIF1A) activation ensues a pseudohypoxia state that drives MDS pathogenesis.

Our recent informatics analyses of primary MDS samples identified a patient subset with high TGF-β signaling in which mitochondrial complex II (MC II) genes were downregulated. MC II, or succinate dehydrogenase (SDH) is the smallest complex of the oxidative phosphorylation system that oxidizes succinate to fumarate and thereby donates electrons to reduce FAD to FADH 2 and the ubiquinone pool. Its downregulation results in the accumulation of Krebs cycle intermediate metabolites that can act as antagonists for 2OGDDs. To understand whether SDH dysregulation alone can contribute to MDS pathobiology, we generated a doxycycline-inducible and reversible mouse model (SDH-KD) that expresses shRNAs against the SDH subunits (SDHA, B, and D) in HSCs. These mice showed a gradual buildup of intracellular succinate in c-Kit + bone marrow (BM) cells and the peripheral blood, and a corresponding decrease of fumarate levels after induction for 2 weeks, 4 weeks, and 2 months. We found that knocking down Sdh was sufficient to induce pancytopenia and trilineage dysplasia in SDH-KD mice, while intracellular iron granules that surround the nucleus of erythroblasts, similar to ring sideroblasts commonly observed in MDS patients were also detected in BM and spleen smears. Of note, removing doxycycline and restoring Sdh expression rescued macrocytic anemia among other MDS phenotypes. Furthermore, treating SDH-KD mice with HMAs restored mitochondrial respiration and reversed HIF1A activation through the upregulation of SDH subunits. Using another MDS model, MllPTD/WT/ Vav1-Cre/Runx1 Flox/Flox, we found that HMA-treated HSCs differentiate towards CD11b +Gr1 + mature myeloid cells and that this requires Sdh expression.

In conclusion, our results indicate that pseudohypoxia- and TGF-β signaling-associated changes in metabolic regulation caused by SDH downregulation underlie MDS pathogenesis. Elucidating the role of MC II in MDS pathobiology may provide alternative targets that can be combined with HMA or luspatercept therapy for MDS.
A Nationwide Analysis of the Outcomes Among Hospitalized Patients with Heparin Induced Thrombocytopenia and Concomitant COVID-19 Infection
Authors: Syal Arshi , Arya Yajur , Casipit Bruce Adrian , Dourado Claudia , Mayo Ryan J , Heller Gordon
Published: December 2023

Background

Heparin induced thrombocytopenia (HIT) and COVID-19 infection are independently associated with thromboembolic complications. Our study aims to assess outcomes among hospitalised patients with concomitant HIT and COVID-19 infection, since data among this subgroup is lacking.

Methods

We utilized the 2018-2020 National Inpatient Sample (NIS) Database in conducting a retrospective cohort study. We identified patients who have heparin induced thrombocytopenia (HIT) and COVID-19 infection using appropriate ICD-10 CM codes. We stratified patients based on the presence or absence of COVID-19 infection. A survey multivariable logistic and linear regression analysis was used to calculate adjusted odds ratios (ORs) for the primary and secondary outcomes. A p value of <0.05 was considered statistically significant. The aim of this study was to investigate the impact of COVID-19 infection on in-hospital mortality, hospital LOS, total hospitalization charge and risk for arterial or venous thrombosis among hospitalized patients with HIT.

Results

We identified a total of 9896 hospitalized patients with heparin-induced thrombocytopenia (HIT), of which 7.68% (760/9896) had concomitant COVID-19 infection. The overall in-hospital mortality among those hospitalized patients with HIT was 17.14% (1695/9896). Among those with concomitant COVID-19 infection, the mortality rate was significantly higher at 50% (380/760, p=0.001). Further, compared to those without COVID-19 infection, those with concomitant COVID-19 infection had a significantly longer median LOS (21 days vs 11 days, p=0.001) and higher total hospitalization charge ($292,325 vs $143,972, p=0.001). Utilizing a stepwise survey multivariable logistic regression model that adjusted for patient and hospital level confounders, COVID-19 infection was found to be an independent predictor of increased in-hospital mortality (adjusted OR 5.13; 95% confidence interval [CI], 3.36-7.84; p=0.001), longer LOS (adjusted OR 6.21; 95% confidence interval [CI], 3.56-8.86; p=0.001) and higher total hospitalization charge (p=0.001). However, it was not significantly associated with increased risk for arterial (adjusted OR 0.72; 95% confidence interval [CI], 0.18-2.83; p=0.63) or venous (adjusted OR 0.91; 95% confidence interval [CI], 0.48-1.74; p=0.79) thrombosis.

Conclusion

Our analysis showed that among hospitalized patients with HIT, concomitant COVID-19 infection increased the risk for in-hospital mortality, longer LOS and higher total hospitalization charge, however, it did not significantly increase the risk for the development of arterial and venous thrombosis. Prospective studies with control of possible confounders are warranted in order to better describe these associations.
Non-Traumatic Epidural Hematoma in Sickle Cell Disease
Authors: Paros Alexandra , Harwood Daniel , Ahmed Rafat , Milillo Jennifer , Fornari Marci , Ahmed Hania
Published: December 2023

Non-traumatic epidural hematoma is an extremely rare finding in patients with sickle cell disease. Spontaneous intracranial epidural hemorrhage has been associated with coagulopathy, malignancy, and infection, but few cases secondary to sickle cell disease have been reported in the literature. The mechanism of hemorrhage in these patients is often unclear, although previous cases have suggested associations with vaso-occlusive pain crises, bone infarction, and/or vascular malformations. We report a 9 year old male with sickle cell disease type SS who presented with right shoulder pain, headaches, emesis, and no neurological deficits. CT head revealed large mixed density left frontoparietal epidural hematoma leading to complete left lateral ventricle effacement and 0.5 cm left to right midline shift. He subsequently underwent a left parietal craniotomy for evacuation of epidural hematoma from which he recovered well. The prevalence and proposed pathologic process behind this rare complication will be discussed.
GLUT1 Overexpression Enhances 19-28z CAR-T Cell Expansion and Metabolic Fitness
Authors: SHI Yuzhe , Kotchetkov Ivan S , Hanina Sophie Alexandra , Sadelain Michel
Published: December 2023

The tumor microenvironment presents many obstacles to effective CAR T cell therapy. One of these obstacles is glucose competition from tumor and myeloid cells to the detriment of CAR T cells. We hypothesize here that CAR T cells with elevated levels of glucose transporter are better fit under such conditions, which will result in improved therapeutic efficacy.

We overexpressed either GLUT1 or GLUT3 in CD19-targeted CAR T cells engineered with a clinically validated CAR (19-28z). GLUT1 is ubiquitously expressed in tissues and GLUT3 has a higher affinity for glucose. Upon in vitro antigen stimulation, 1928z-GLUT1 and 1928z-GLUT3 showed greater enrichment of CAR+ T cells and underwent more cell divisions than control 1928z-LNGFR under glucose-limiting conditions. In repetitive antigen stimulation assays, 1928z-GLUT1 expanded significantly more than 1928z-LNGFR cells in 10 mM or 5 mM glucose medium. Surprisingly, 1928z-GLUT3 CAR T cells faired markedly less well than 1928z-LNGFR controls, associated with reduced viability of 1928z-GLUT3 CAR T cells after 2 or 3 exposures to antigen. In the NALM6 acute lymphoblastic leukemia model, low-dose CAR T cell treatment with 1928z-LNGFR and 1928z-GLUT3 did not curtail rapid tumor progression, while 1928z-GLUT1 CAR T cells significantly increased survival. More abundant CAR+ T cells were found in the bone marrow and spleen of 1928z-GLUT1-treated mice. Interestingly, we observed consistent increases in CD62L+CD45RA+ Tscm population in 1928z-GLUT1 cells, including both CD8 and CD4 T cells. We flow-sorted CAR T cells from treated mice and performed SMARTer RNA-seq. GLUT1, but not GLUT3, significantly upregulated TCF7 and genes encoding glycolytic enzymes and electron transport chain, indicating that GLUT1 overexpression enhances both glycolysis and oxidative phosphorylation.

To further test GLUT1-CAR-T function in tumor lines with high levels of glucose transporter, we generated a renal cell carcinoma (RCC) patient-derived xenograft (PDX) with ectopic expression of human CD19. CAR-T cells were repetitively stimulated with the RCC PDX under titrated glucose concentrations. 1928z-GLUT1 significantly improved CAR T cell viability compared to 1928z-LNGFR in low glucose conditions. The expansion of GLUT1 CAR-T cells in 10 mM and 5 mM glucose also surpassed that of LNGFR CAR-T cells. In vivo, 1928z-GLUT1-treated mice showed significantly less tumor burden than 1928z-LNGFR-treated mice.

Overall, we developed a novel strategy for enhancing the metabolic fitness of 1928z CAR-T cells by constitutively overexpressing GLUT1. GLUT1 but not GLUT3 enhances glycolysis and oxidative phosphorylation in CAR-T cells, which sustains T-cell expansion and increases therapeutic efficacy. This strategy is under investigation with other CAR formats.
Autophagy Inhibition Prevents CAR-T Exhaustion and Terminal Differentiation Via TCF7 Accumulation
Authors: Teng Xinyi , Shao Mi , Guo Xin , Cheng Tao , Qian Pengxu , Huang He
Published: December 2023

Introduction: Chimeric antigen receptor T (CAR-T) cell therapy has achieved remarkable success in hematological malignancies. However, CAR-T cell dysfunction due to exhaustion has been identified as a major roadblock to effective CAR-T cell therapy. Autophagy mediates T cell differentiation, survival, and memory by regulating the degradation of organelles and specific proteins. A recent study reported that inhibition of autophagy significantly enhanced the killing effect of CD19 CAR-T cells. Yet, the impact of autophagy inhibitors on CAR-T cell exhaustion and maintenance is still unclear. The study aims to systematically evaluate the role and mechanism of autophagy inhibitors on CAR-T cells.

Methods and Results: To explore whether autophagy inhibitors could prevent CAR-T cell exhaustion triggered by antigen-independent CAR tonic signaling, we applied 3 autophagy inhibitors: SBI0206965, MRT67307, Compound C, respectively, to the culture medium of CD19.4-1BBz CAR-T cells. We found that all the 3 autophagy inhibitors could reduce CAR-T cell exhaustion and terminal differentiation. The results were replicable with GD2.28z CAR-T cells. Among the three inhibitors, SBI0206965 showed the most significant effect. Thus, we chose SBI0206965 for further research. CD19.4-1BBz CAR-T cells pretreated with SBI0206965 demonstrated enhanced killing capacity against Nalm-6 cells in vitro. Using the Nalm-6-bearing leukemia xenograft model, we found that compared to DMSO pretreated CD19.4-1BBz CAR-T cells, SBI0206965 pretreated CD19.4-1BBz CAR-T cells showed superior persistence and antitumor efficacy, exhibited a less exhausted and differentiated state, and further prolonged mice survival (Fig.1). To evaluate whether SBI0206965 could protect CAR-T cell exhaustion triggered by antigen stimulation, we cocultured CD19.4-1BBz CAR-T cells with Nalm-6 cells in the medium with or without SBI0206965. We demonstrated that the addition of SBI0206965 could effectively inhibit CAR-T cell exhaustion and terminal differentiation driven by antigen exposure. By performing proteomics, we identified TCF7 enriched in SBI0206965-treated cells. We validated the differences using Western blot analysis, which showed increased TCF7 levels in CAR-T cells treated with SBI0206965 for 24 hours. Thus we speculated that inhibition of autophagy might contribute to the accumulation of TCF7 and further improve CAR-T maintenance. To further verify whether TCF7 was degraded in an autophagy-dependent manner, we next determined TCF7 turnover rates in the presence of cycloheximide (CHX) in CAR-T cells with or without SBI0206965 treatment. CHX chase assay showed that SBI0206965 treatment could slower the TCF7 degradation rate(Fig.2).

Significance: Our research provides a strategy to optimize CAR-T antitumor efficacy and persistence via TCF7 accumulation by autophagy inhibition.
αGal9Ab Treatment As a Novel Therapy for Blood Cancer
Authors: Lee Miyoung , Henry Curtis James
Published: December 2023

Blood cancers are the most common cancer in children accounting for roughly 28% of all childhood cancers. Of these, acute lymphoblastic leukemia ( ALL) and acute myeloid leukemia ( AML) are the most prevalent. Advancements in treatment regiments for these diseases have led to a 5-year overall survival rate of 90% for pediatric ALL and 65~70% for pediatric AML; however, in patients with relapsed or refractory disease, the overall survival rates decline to 30~50% post-treatment.

We have recently demonstrated that antibody-mediated targeting of Galectin-9 ( GAL-9), is highly cytotoxic to human B-ALL cells in vitro and confers a significant degree of protection for disease progression in pre-clinical models of B-ALL (M. Lee et al., Nature Communications, 2022). In unpublished studies, we have subsequently found that anti-GAL-9 antibody ( αGAL-9Ab) treatment is highly cytotoxic to human T-ALL and AML cells. Notably, we have found that human pediatric cell lines are more sensitive to this treatment than those derived from adult patients. Mechanistically, we have uncovered a previously unreported link between GAL-9-mediated regulation of checkpoint kinase 1 ( CHEK1) and anti-apoptotic protein myeloid leukemia 1 ( MCL-1), with αGAL-9Ab treatment leading to a deregulation of both proteins in T-ALL cells. Notably, αGAL-9Ab treatment synergized with multiple CHEK1 inhibitorscurrently being tested in phase I or II clinical trials (GDC-0575, Prexasertib and PF477736) to enhance the in vitro killing of human T-ALL, B-ALL, and AML cells. Furthermore, using xenograft models, we found that single-agent αGAL-9Ab treatment was superior to single-agent chemotherapy and vehicle treatment at conferring protection against T-ALL progression.

In conclusion, single-agent αGAL-9Ab treatment appears to be cytotoxic against multiple acute leukemia subtypes ( ALs) and its efficacy is enhanced with CHEK1 inhibition. To validate this novel combinatorial approach for treating ALs, additional studies need to be performed to define the mechanisms of action and further pre-clinical experiments will be invaluable in determining the efficacy and safety of these therapies.
Effects of Air Pollution on Respiratory Events and Pain Crises Among Children with Sickle Cell Disease in New York City
Authors: De Aliva , Jung Kyung hwa , Davis Haley , Siddiqui Abeer , Kattan Meyer , Quinn James , Rundle Andrew G. , Lovinsky-Desir Stephanie , Green Nancy S.
Published: December 2023

Introduction: Sickle cell disease (SCD) has the highest U.S. disease burden among Black and Hispanic populations, who more commonly live in urban areas with poor ambient air quality. The effect of air pollution exposure and social environmental factors on health outcomes in SCD have not been extensively studied, especially among children, and may further contribute to health disparities.

Objectives: This study aims to examine the associations between air pollution exposure and acute respiratory and pain events and to determine whether neighborhood-level social disadvantage would modify their associations.

Methods: We collected retrospective data on unscheduled outpatient visits, emergency department (ED) visits, and hospital admissions for respiratory events (i.e., respiratory tract infections, asthma exacerbation, acute chest syndrome) and vaso-occlusive pain crises requiring hospitalization (VOCs) for children with SCD in a tertiary care center in New York City (NYC) from 2015-2018. A total of 114 children with SCD (Table 1) were included in this study and had between 1-4 annual repeated measures of annual average air pollutants (i.e., black carbon (BC), fine particulate matter (PM 2.5), and nitrogen dioxide (NO 2)) with a total of 425 visits. Using home addresses, modeled data from the NYC Community Air Survey data were used to estimate street-level, annual average exposure to air pollution. The area deprivation index (ADI), a composite index of neighborhood-level social disadvantage, was dichotomized at the upper tertile (higher vs lower). Multivariable Poisson regression in generalized estimating equation (GEE) models were used to estimate relative risks (RR), after adjusting for potential covariates.

Results: Overall, there were no significant associations between air pollution levels and acute respiratory and pain events among children with SCD. When stratified by ADI, increased exposure to PM 2.5 and NO 2, but not BC, was significantly associated with more frequent respiratory outpatient and ED visits among children residing in higher ADI neighborhoods (RR (95%CI)= 1.13 (1.00, 1.28) and 1.43 (1.01, 2.02) for PM 2.5 and NO 2, respectively, p<0.05 for all), but not among those in lower ADI neighborhoods (P-value for interaction=0.2 and 0.02 for PM 2.5 and NO 2, respectively) (Figure 1). Associations of air pollution with hospital admissions and VOCs were not observed for neither high ADI nor low ADI neighborhoods (p-value>0.05).

Conclusions: Exposure to air pollutants increased respiratory complications among children living in deprived neighborhoods.
Erythropoietin Treatment in Mice Enriches Bone Marrow Osteoclast Precursors, As Reflected By Increased Macrophage Colony-Stimulating Factor (MCSF) and Elevated Expression of Osteoclast Markers in Monocytes
Authors: Gorodov Anton , Kolomansky Albert (Alex) , Piper Michelle , Lezerovich Lior , Ben-Califa Nathalie , Gabet Yankel , Neumann Drorit
Published: December 2023

The impact of erythropoietin (EPO) on erythropoiesis has long been established. However, recent studies have indicated that EPO may also exert regulatory effects on other hematopoietic lineages, including the monocyte lineage. In that respect, we have previously shown that EPO treatment in mice induces bone loss. Here, we studied the effect of EPO on bone marrow (BM) monocytes and its potential implications for osteoclastogenesis. To address the dynamic changes in monocyte populations following EPO administration, we injected EPO (180U) once, or 3 times on alternating days for one week. Mice were sacrificed 16 h after EPO injection. Our results revealed a significant 20 percent increase in the monocyte population (TER119 -, LY6G -, CD11b +) 16 hours after EPO injection, and a twofold increase in the levels of these cells after one week of 3 injections of EPO. Soluble macrophage colony-stimulating factor (MCSF) protein levels in the bone marrow were increased 1.5-fold 16 h after EPO injection, while MCSF mRNA expression levels remained unchanged. After 1-week of EPO administration (3 injections), although the global BM expression of MCSF was reduced, we observed a two-fold increase in MCSF expression levels in T cells (CD3 +). To further elucidate the functional consequences of EPO-induced monocyte expansion, we isolated monocytes from mice treated with EPO for one week. We characterized expression levels of key osteoclastic markers in these cells. Indeed, we found that in BM monocytes, EPO treatment led to the upregulation of Lymphocyte function-associated antigen 1 (LFA-1) and Receptor Activator of Nuclear Factor κ B (RANK). The former is implicated in osteoclastogenesis via its interaction with intercellular adhesion molecule-1 (ICAM−1), while the latter is the surface receptor for RANK ligand which regulates osteoclast differentiation and activation. These data suggest that EPO not only influences BM monocyte expansion, but also contributes to the regulation of osteoclast differentiation, thus linking erythropoiesis to bone homeostasis. Furthermore, we investigated the early effects of EPO treatment on the expression of transcription factors implicated in monocyte differentiation. Remarkably, we found that EPO administration for 16 hours resulted in the downregulation of KLF2 and KLF4, transcription factors involved in monocytic lineage commitment. The rapid EPO-induced expansion of the monocytic population, coupled with the EPO-regulated expression of MSCF, osteoclastic markers, and transcription factors, suggests a role for EPO in modulating monocyte-to-osteoclast differentiation. The current study opens avenues for further investigating the crosstalk between erythropoiesis, myelopoiesis, and bone homeostasis.
MRI Reliably Captures Bone Marrow Metrics in Myelofibrosis
Authors: Robison Tanner , Levinson Annabel , Lee Winston , Pettit Kristen Marie , Malyarenko Dariya , Johnson Timothy D , Chenevert Thomas , Ross Brian , Talpaz Moshe , Luker Gary
Published: December 2023

Purpose

Biopsy remains the clinical standard for evaluating bone marrow of patients with myelofibrosis (MF). However, randomly sampling a small volume of bone marrow, typically from the ilium, fails to capture heterogeneity of disease across anatomic sites. The invasive, painful nature of the procedure limits patient acceptance of repeated biopsies to monitor disease status and response to therapy. Here, we present a non-invasive, MRI technique for evaluating bone marrow in MF.

Participants and Methods

We analyzed bone marrow of 66 study participants (45 with MF; 15 with essential thrombocytosis or polycythemia vera; and 6 healthy controls). We assessed participant bone marrow across three anatomic sites (vertebral bodies, ilium, and femoral heads), using three quantitative MRI sequences: proton density fat fraction (PDFF) for fat content; apparent diffusion coefficient (ADC) for cellular effect on water mobility; and magnetization transfer (MT) for macromolecular structure and fiber deposition. We correlated these MRI metrics with patient prognosis (dynamic international prognostic scoring system; DIPSS) and iliac biopsy data for cellularity and fibrosis (MF grade).

Results

Participants with MF had elevated MT ratio (MTR) and ADC and lower PDFF compared to healthy participants. Non-MF MPN participants demonstrated the same trend, though to a lesser extent. Individual MRI metrics correlated strongly across anatomic sites (r pearson from 0.57 to 0.89, p-value < 0.01), with notable heterogeneity both within and among different regions. DIPSS risk groups had limited correlation with MRI metrics. To understand to what extent quantitative MRI biomarkers identify fibrotic marrow, we developed a multivariate logistic regression model to stratify the 45 participants with MF by early fibrosis (MF grades 0-1) and overt fibrosis (MF grades 2-3). Of the seven total anatomy-MRI metric combinations, three had the greatest contributions toward grading marrow fibrosis: PDFF in the vertebral bodies and ilium along with ADC in the ilium. We developed the logistic regression model to predict early and overt fibrosis with these three parameters using a 70%/30% train/test split with 5-fold cross validation. Based on this model, quantitative MRI metrics are associated with the severity of fibrosis in the bone marrow (test set performance: accuracy = 84.6%, sensitivity to predict overt fibrosis = 88.9%, specificity = 75%, area under the receiver operator curve = 0.94).

Conclusion

Quantitative bone marrow MRI reliably captures relevant metrics of MF bone marrow, including cellularity and fat replacement by fibrosis, and may be useful in clinical drug trials and patient management for MF.
Notch Signaling Is Required for Extramedullary Hematopoiesis in the Adult Liver
Authors: Bandara Thilinie , Shao Lijian , Paik Na Yoon , Neethling Jacob , Rodriguez Paulina , Pajcini Kostandin V
Published: December 2023

During embryonic development, hematopoietic stem cells (HSC) in the fetal liver undergo expansion and self-renewal. Fetal HSCs migrate to the bone marrow (BM) niche towards birth and remain there during adulthood. Extramedullary hematopoiesis (EMH), or hematopoiesis outside of the BM, predominantly occurs in the fetal liver or when the adult BM niche ceases to be a site for functional hematopoiesis. The mechanism by which the adult liver becomes reactivated as a site of EMH for itinerant HSCs is unknown. Our project is investigating EMH in the adult liver by inducing acute hemolytic anemia in adult mice that induces egress and homing of hematopoietic progenitors to the liver and spleen. Our work demonstrates that long-term HSCs isolated from the portal vein in the adult liver can serially reconstitute lethally irradiated mice. Thus, the adult liver serves as a potential reservoir for long-term, functional HSCs in the presence of a damaged BM niche. In pursuit of a mechanism, we focused on Notch-signaling which is an evolutionarily conserved pathway that has been shown to play a critical role in HSC emergence and fetal liver development. When comparing WT liver donors to Notch-deficient (Notch1 +/ΔTAD) or Notch knockout (Notch1 ffVavCre +) liver donors, hematopoietic transplants reveal that Notch mutant progenitors fail to efficiently reconstitute irradiated recipients. Using an HSC-specific a-catulin-GFPmouse model, we are able to track the expression of GFP + liver-originating HSCs after hematopoietic reconstitution. We show that mice reconstituted by EMH-activated a-catulin-GFP liver HSCs are able to provide long-term reconstitution in lethally irradiated recipients and resume residence in the bone marrow niche of recipients. Additionally, we see distinct localization of GFP + HSCs localized near sinusoids throughout the EMH liver niche using whole-mount imaging. We have validated receptor-ligand binding interactions of the Notch1 receptor with ligand-expressing hematopoietic cells in the adult liver. Using intracellular flow cytometry after EMH in the adult liver, we see a 3-fold increase in mean fluorescent intensity of GATA2 in HSPCs (hematopoietic stem cell progenitors) when compared to our bone marrow and isotype control mice. We demonstrate that Notch-driven GATA2 expression is a crucial regulator of self-renewal and functional maintenance of stemness during EMH of HSCs in the adult liver. Our findings reveal a developmentally conserved Notch signaling mechanism is reactivated in the adult liver to retain the functional potential and maintenance of HSCs as they adapt to inhospitable conditions in the bone marrow niche.
A Mutation in Complement Factor H (W1183R) Enhances VWF Proteolysis By ADAMTS13 Under Shear
Authors: Cao Wenjing , Liu Yi , Zhang X. Frank , Zheng X. Long
Published: December 2023

Introduction: Complement factor H (CFH) is the primary negative regulator of complement activation in the alternative pathway in both fluidic phase and on cell surface. Loss of function of CFH such as W1183R is associated with atypical hemolytic uremic syndrome (aHUS). Homozygous mutation W1206R in the murine CFH, corresponding to W1183R in human CFH, results in acute aHUS. Interestingly, in these mice, while plasma VWF antigen levels are significantly increased, plasma VWF multimer size is markedly reduced in ADAMTS13-depedent manner. However, the molecular mechanism underlying such a dramatic effect of the mutant CFH (W1206R) on VWF multimer distribution is not known. The goal of this study is to determine how wild type CFH and mutant CFH W1183R affects VWF proteolysis under shear, and the mechanism of action.

Methods: Recombinant WT CFH, CFH W1183R, ADAMTS13, and VWF fragments were expressed in mammalian cells and purified to homogeneity. The effects of WT CFH or CFH W1183R on proteolytic cleavage of VWF73 peptide and full-length multimeric VWF were determined. Additionally, the binding and unfolding events were determined by micro scale thermophoresis (MST) and optical tweezer, respectively.

Results : WT CFH showed no effect on the cleavage of peptide VWF73 substrate, but CFH W1183R enhanced the proteolytic cleavage of VWF73 by ADAMTS13 in a dose-dependent manner ( Fig. 1). Counterintuitively, WT CFH appeared to inhibit the proteolytic cleavage of multimeric VWF by ADAMTS13 under arterial shear, but CFH W1183R exhibited an opposite effect, modestly enhancing the proteolytic cleavage of multimeric VWF by ADAMTS13 under the same conditions. The MST binding assay demonstrated that both WT CFH and CFH W1183R bind to a short A1, a long AIM-A1, and A1-A2-A3 fragments with a dissociation constant ( K D) of 6.4 x 10 -7, 3.0 x 10 -9 M, and 3.3 x10 -9 mol/L, respectively. These results suggest that CFH may bind to the AIM, A1-A2 junction, and A2 domain. To further assess the impact of CFH binding to A2, we performed optical tweezer experiments. The results showed that both WT CFH (Lc: from 27.4 nm at 12 nM to 40 nm at 1.0 mM) and CFH W1183R (Lc: from 26 nm at 12 nM to 31.8 nm at 1.0 mM) resulted in significantly extended contour length (Lc) of VWF-A2 domain in a concentration-dependent manner ( Fig. 2), suggesting a dramatic conformational change in the A2 domain upon binding by WT CFH or CFH W1183R.

Conclusions : Our results demonstrate that WT CFH at physiological concentrations may exhibit a modest inhibitory activity towards cleavage of VWF by ADAMTS13 under shear, but such an inhibitory activity disappears as the result of a point mutation in the C-terminal end of CFH (e.g., W1183R). Instead, the mutant CFH W1183R appears to enhance the VWF cleavage by ADAMTS13, likely through direct binding to the AIM, A1-A2 junction, and A2 domain of VWF, resulting in conformational changes in the central A2 domain.