bluuebunny/crossref_metadata_2025 huggingface dataset:
Reduction of Lactate Production By Phospholipase C Inhibition in Cold Stored Platelets
Authors: Oh Thomas T , Talackine Jennifer R , Lee Ji H , Barrera Gema D , Meledeo Michael A , Xiang Lusha
Published: December 2024

Introduction: Anaerobic glycolysis and resultant lactic acid accumulation have been shown to impair platelet function during cold storage. The goal of current study is to understand the contribution of phospholipase C (PLC) activity to glucose utilization in cold stored platelets.

Methods: Healthy human platelets were stored in 35% plasma and 65% platelet additive solution (PAS) at 4oC for 15 days after PLC inhibitor (U73122 in 1% dimethyl sulfoxide (DMSO)). Mini aliquot storage bags (BCSI) were used to store platelets in PAS. An iSTAT blood analyzer (Abbott Point of Care) using CG4+ and CHEM8 analysis cartridges measured levels of glucose and lactate values on days 1 - 15 of storage. Five groups were included: control, DMSO vehicle, and three doses of U73122: 10, 20, and 50 µM.

Results: The mean basal level (BL) of lactic acid in the PAS was 0.31 ± 0.01 mM in control and DMSO vehicle groups on the storage day 1 and increased on day 15 of storage to 1.67 ± 0.14 mM in control (5.4-fold increase) and 1.83 ± 0.18 mM in vehicle (6-fold increase). However, lactic acid level only slightly increased to 0.96 ± 0.06 mM, 0.62 ± 0.07 mM, and 0.51 ± 0.05 mM in 10, 20, and 50 µM U73122 groups, respectively, after storage for 15 days. The BL glucose in PAS was 29 ± 0.92 mg/dl in control and DMSO vehicle groups, decreasing to 12.75 ± 2.44 mg/dl (control) and 12.14 ± 2.14 mg/dl (vehicle) after 15 days. Only mild decreases in glucose were observed 15 days after U73122 treatments (24.4 ± 1.47 mg/dl, 24.4 ± 1.94 mg/dl, and 25.2 ± 1.3 mg/dl in 10, 20, and 50 µM, respectively).

Conclusion: The dose-dependent decrease in glucose usage and lactate production with PLC inhibitor U73122 indicate that anaerobic glycolysis is being inhibited in these cold-stored platelets. This suggests that PLC may be a potential therapeutic target to alleviate acidosis and platelet storage lesions.

DoD disclaimer: The views expressed in this abstract are those of the author(s) and do not reflect the official policy or position of the U.S. Army Medical Department, Department of the Army, DoD, or the U.S. Government.
Inhibiting Free Fatty Acid Transport to Improve CAR-T Cell Therapy of Relapsed B-Cell Acute Lymphoblastic Leukemia
Authors: Garcia Clarissa , Lyons Kaylyn , Liu Tian , Iacobucci Ilaria , Novak Amanda , Argabright Amy , Donnelly Alexis , Grandvallet Contreras Julian , Geng Huimin , Smith Sabrina , Zhou Xin , Müschen Markus , Dick John Edgar , Haines Julie R , D'Alessandro Angelo , Mullighan Charles G. , Phang Tzu , Kohler M. Eric , Witkowski Matthew
Published: December 2024

Introduction: B-cell acute lymphoblastic leukemia (B-ALL) resistant to conventional chemotherapy is a leading cause of cancer-related mortality in children and young adults. Although CD19-directed Chimeric Antigen Receptor T-cell (CAR-T) therapy shows promise for relapsed/refractory B-ALL, its effectiveness is often compromised by leukemia-intrinsic resistance mechanisms that remain unclear. Recent evidence suggests that TP53 mutations negatively affect the long-term efficacy of CD19 CAR-T therapy in B-ALL, despite initial trials not identifying high-risk genetic features impacting early clinical responses. TP53 loss-of-function mutations, although rare at diagnosis, are enriched in relapsed B-ALL and associated with chemotherapy resistance and poor survival.

Methods and Results: To dissect the B-ALL-intrinsic role of p53 during CAR-T therapy, we found that TP53 loss-of-function mutations in a subset of human B-ALL cell lines conferred resistance to CAR-T cell killing in vitro and in vivo. To elucidate the pathways driving CAR-T resistance in TP53-mutated (mut) B-ALL cells, we performed genome-wide CRISPR/Cas9 screening on isogenic TP53-wildtype (wt) and TP53-mut CD19+ B-ALL cell lines treated with CD19-CAR-T cells ex vivo. This screening approach identified the free fatty acid transporter FATP2 (encoded by SLC27A2) as a critical factor; as its genetic ablation enhanced CAR-T efficacy against TP53-mut B-ALL lines. Analysis of RNA-seq data from 1104 pediatric B-ALL patients revealed that high SLC27A2 mRNA expression at B-ALL diagnosis correlated with poor overall survival and increased free fatty acid metabolism. To understand the mechanism of FATP2 activity in B-ALL survival, we found FATP2-expressing B-ALL lines internalize long-chain free fatty acids more efficiently than non-FATP2-expressing human B-ALL cell lines. Additionally, FATP2-mediated CAR-T resistance in TP53-mut B-ALL cells was reversible by depleting exogenous fatty acids. Stable isotope free fatty acid uptake tracing revealed a role for FATP2 in promoting succinate accumulation in TP53-mutated B-ALL cells, thus significantly altering cellular metabolism that may have contributed to CAR-T resistance. Ongoing experiments aim to identify the precise mechanisms contributing to leukemia-intrinsic, FATP2-dependent CAR-T resistance.

Conclusions: Our studies suggest that FATP2 expression drives TP53-mutant B-ALL CAR-T resistance by facilitating the uptake of exogenous free fatty acids. Collectively, we propose targeting free fatty acid transport as a specific metabolic vulnerability in B-ALL that might improve CAR-T therapy outcomes.
Evaluating the Effectiveness and Security of Antifungal Prophylaxis in Acute Lymphoblastic Leukemia Induction Chemotherapy
Authors: Zhou Yang , Zhou Hongsheng , Liu Qifa
Published: December 2024

Background and objectives: Patients suffering from hematologic malignancies face a high risk of developing invasive fungal disease (IFD). Primary antifungal prophylaxis (PAP) is a key strategy to reduce the incidence of IFD in patients with acute myeloid leukemia (AML), those receiving hematopoietic stem cell transplantation (HSCT), and those with myelodysplastic syndrome (MDS). Nonetheless, clinical data regarding PAP in ALL patients is scarce, owing to major drug interactions. This paper aims to evaluate the efficacy and safety of PAP in ALL patients through real-world research.

Methods: A retrospective cohort study was carried out at a single center, focusing on ALL patients aged over 14 who received their first induction chemotherapy at our hospital from January 2011 to January 2021. Our study excluded individuals with a background of IFD, those who declined treatment and were released from the hospital, and those whose cases were inaccessible. The identification of IFD adhered to the guidelines set by the European Organization for Research and Treatment of Cancer/Mycosis Study Group (EORTC/MSG).

Results: The study included 482 patients, 316 of whom undergoing PAP treatment. Of the 482 patients, 13 developed proven or probable IFD (2.7%), and 60 developed possible IFD. There was a notably reduced occurrence of IFD in the prevention group compared to the non-prevention group (36/316, 11.4% vs. 37/166, 22.3%, P = 0.002). The multifactorial Cox regression analysis revealed a notable decrease in IFD occurrences due to PAP (HR = 0.306, P < 0.001). The primary medications administered in PAP included caspofungin (n = 113), voriconazole (n = 99), and micafungin (n = 53). The caspofungin group exhibited no adverse drug reactions. Patients experiencing neutropenia for more than 10 days and those on corticosteroids for 7 days or more showed a reduced mortality risk in the PAP group compared to the nonprophylactic group (HR = 0.615, P = 0.021).

Conclusions: The practical research we conducted revealed that PAP independently safeguards against IFD in ALL patients undergoing induction therapy. Furthermore, caspofungin's safety record for treating PAP in ALL patients is positive.
Treatment of First Relapse of Multiple Myeloma in Australia and New Zealand (ANZ): Treatment Patterns and Outcomes: An ANZ Myeloma and Related Diseases Registry (ANZ MRDR) Analysis
Authors: Gration Betty , Wellard Cameron , Moore Elizabeth , Tan Marianne , Murphy Matthew , Lim Kenneth J C , Rowland James , Hamad Nada , Rajagopal Rajeev , Mollee Peter , Spencer Andrew , Quach Hang , McCaughan Georgia
Published: December 2024

Aim:

There remains lack of consensus regarding whether relapsed multiple myeloma should be treated at clinical progression (CP) or biochemical progression (BP), and the ensuing clinical outcome. We described practices in Australia and New Zealand (ANZ) and the outcomes in these cohorts.

Method:

We conducted a retrospective review of ANZ Myeloma and Related Diseases Registry (ANZ MRDR) patients across 5 sites, that had progressed following first line therapy. Additional information was sought regarding timing of treatment at myeloma relapse (BP or CP) and presence of end organ damage (EOD) at 3-6 months. We determined progression free survival (PFS) and overall survival (OS) in these cohorts using Kaplan Meier methods.

Results:

Among 258 patients, 52% were treated at BP and 48% at CP. There was no significant difference in age, gender, ECOG, stage (ISS-3) or high-risk cytogenetics at diagnosis between groups. There was no significant difference in time to first progression (CP=20.6 months vs BP=21.7 months) or time between progression and initiation of therapy (CP=1.7 vs. BP=2.1 months). Patients treated at CP had a higher rate of persistent opioid requirement at 6 months (CP=32% vs BP=13.9%, p=0.001) and need for a mobility aid at 6 months (CP=12.7% vs. BP=3.5%, p=0.012), but there was no difference in fracture rates between the groups at 6 months (CP=10.6% vs. BP=10.5%, p=0.99). 68% of patients with renal EOD had no renal response as per IMWG at 3 months. There was a trend toward longer PFS and improved OS from initiation of second line therapy for those treated at BP versus those treated at CP however no statistical significance recognised at this study power (PFS: CP=8.5 months vs. BP=11.2 months, p=0.203, OS: CP=23.5 months vs. BP=33.7 months, p=0.06).

Conclusion:

Initial disease characteristics, time to first progression and time from progression to initiation of second line therapy did not differ between patients treated at BP versus those treated at CP. Treatment at CP was associated with more persistent EOD measured by requirement for opioids and mobility aids and the majority of those with renal EOD failed to have a renal response. Those treated at BP showed a trend towards improved PFS and OS compared to treatment at CP. This trend may be related to persistent EOD or more aggressive disease biology. The lack of statistical significance likely reflects poor power and warrants further investigation.
Analysis of Arterial Occlusive Events (AOE) in CML Patients Using Real-World Data : A Large Cohort
Authors: Takaku Tomoiku , Kojima Yoshitsugu , Yanai Yoshiaki
Published: December 2024

Background

While the prognosis of chronic myeloid leukemia (CML) has been significantly improved by the development of tyrosine kinase inhibitors (TKIs), the importance of managing various adverse events associated with long-term TKI treatment is increasing. In particular, arterial occlusive events (AOEs) can be fatal and potentially cause irreversible sequelae in patients, requiring the choice of TKIs according to individual patient risk and proactive therapeutic interventions for prevention. In this study, we conducted an analysis using data from the DPC database, MDV (Medical Data Vision Co., Ltd., and examined AOE in the real-world setting of a large cohort.

Methods

We extracted data from MDV for 2,113 patients aged 18-100 (average age 61.3; 1,326 men and 787 women) who were diagnosed with CML and received TKI treatment from Jan 2017 to Dec 2020. We used logistic regression analysis to analyze the frequency and timing of AOE, and individual AOE risk factors such as gender, age, and history of cardiovascular event, hypertension, diabetes, and dyslipidemia, by various TKI treatment lines.

Results

Of the 2,113 MDV registrants, 1,381 were 1st-line TKI treatment patients, 550 were 2nd-line, 150 were 3rd-line, and 32 were 4th-line or later. The incidence of AOE by treatment line was increasing with the later treatment lines (8.5% for 1st-line, 12.5% for 2nd, and 14.7% for 3rd: p=0.0079 (1st vs 2nd), p=0.0164 (1st vs 3rd)). The person-year method (/1000 person-years) was 37.7 for 1st-line, 95.1 for 2nd-line, and 143.9 for 3rd-line. The median time to AOE onset was 27.0 days for 1st-line, 25.0 days for 2nd-line, and 25.5 days for 3rd-line. Furthermore, analyzed by type of AOEs, we found that the median time of stroke and peripheral arterial occlusive disease were similar regardless of the treatment line. However, incident rate of myocardial infarction was increased at later line (26.0 days for 1st-line, 29.0 days for 2nd-line, and 45.0 days for 3rd-line).

In the analysis of AOE risk factors, all factors showed significant differences in the comparison between 1st-line and 2nd-line in univariate analysis, but in multivariate analysis, age, gender, dyslipidemia, and hypertension were significant risk factors in 1st-line, while age and hypertension were significant in 2nd-line. On the other hand, hypertension, diabetes, and dyslipidemia were significant in univariate analysis, and diabetes was significant in multivariate analysis in 3rd-line group.

【Conclusion】The results of the real-world data analysis of a large cohort using the DPC database showed that the incidence of AOE increased with the estimation of treatment lines, and significant risk factors varied in each treatment line. In particular, hypertension and diabetes were significant risk factors for AOE in all treatment lines in both univariate and multivariate analyses, suggesting that the proactive therapeutic interventions are strongly recommended for these risk factors.
Gain or Amplification of 1q21 in Systemic Light Chain Amyloidosis Is Associated with Advanced Mayo Stage, Plasma Cell Disease and Worse Overall Survival
Authors: Oubari Sara , Papathanasiou Maria , Michel Lars Christian , Rassaf Tienush , Thimm Andreas , Hagenacker Tim , Ehling Daniela , Wieczorek Stefan , Naser Eyad , Duehrsen Ulrich , Reinhardt Hans Christian , Carpinteiro Alexander
Published: December 2024

Systemic light chain amyloidosis is an acquired protein misfolding disease characterized by extracellular deposition of misfolded amyloidogenic immunoglobulin light chain fibrils secreted most often from clonal plasma cells in the bone marrow. Interphase fluorescence in situ Hybridization (iFISH) is an essential method to characterize disease biology and to define the treatment approach in multiple myeloma. In this retrospective study we included 169 patients with AL amyloidosis who presented between 2015-2024 and analyzed the role of iFISH aberrations on clinical characteristics and outcome. The most commonly detected aberrations were t(11;14) in 56.2%, deletion 13q14 in 33%, +1q21 in 21%, hyperdiploidy in 20.3% and deletion 16q23 in 16.4%. Significant elevations in dFLC levels were observed in patients with +1q21 (median 406 vs. 208 mg/l, p = 0.02), deletion 16q23 (median 476 vs. 203, p = 0.005), with a trend noted in t(11;14) (median 272 vs. 140 mg/l, p = 0.055). Only +1q21 was significantly associated with increased levels of biomarkers for cardiac damage including NTproBNP (median 8845 vs. 3482 pg/ml, p = 0.002) and hsTnT (median 104 vs. 52 ng/l, p = 0.001). This resulted in an increased proportion of patients with Mayo stage IIIb (53% vs. 26%, p = 0.02). Patients positive for +1q21 also had more advanced plasma cell disease (p = 0.001). In the overall population, +1q21 showed worse overall survival after a median follow-up time of 38 months (p = 0.006). In light of current induction regimens with bortezomib and daratumumab, our study is the first that identifies +1q21 as the main chromosomal aberration that predicts poor survival.
Functional and Molecular Analyses Reveal Impaired HSPCs in Multiple Myeloma Patients Post-Induction
Authors: Baumhardt Thanh Mai , Sakk Vadim , Soller Karin , Kull Miriam , Kronke Jan , Geiger Hartmut , Mulaw Medhanie Assmelash
Published: December 2024

Multiple Myeloma (MM) is an incurable disease due to relapse, despite development of novel agents and progress in autologous stem cell transplantation (ASCT) (Rajkumar, 2022). There remains a high variability in patient ASCT response in MM. We investigated the function and the transcriptome of CD34+ hematopoietic stem and progenitor cells (HSPC) of newly diagnosed, treatment-naïve MM (NDMM) patients and that of patients that underwent induction treatment prior to ASCT (treated MM) to inform us on possibilities to further improve ASCT outcomes. HSPCs from treated MM patients, when transplanted into NBSGW recipients, presented with a significantly impaired function compared to control HSPCs from healthy aged-matched donors (CD45+ frequency in recipients 0.8% vs. 16.7%, P < 0.0001), while HSPCs from NDMM patients showed a function similar to control HSPCs. In addition, there was a significant lymphoid differentiation deficiency of HSPCs from NDMM patients (CD19+ frequency 57.7% vs. 75.5%, P 0.02). To delineate likely mechanisms, single-cell RNA-sequencing analyses of CD34+ cells were performed. Interestingly, the transcriptomic profile of HSPCs from MM patients (both before, but also especially after treatment) already present with a MM gene signature, despite the fact that HSPCs do not comprise the target malignant cell population of MM (CD138+ plasma cells). Using machine learning (ML) algorithms, we developed a model to identify gene signatures among distinct types of HSPCs from MM patients. Based on a blinded dataset, the model correctly predicted 55.2%, 94.2% and 77.5% cells to healthy control, NDMM or treated MM group, respectively, revealing that NDMM HSPCs show the highest gene signature specificity. Interestingly, 16.8% of the treated MM cells showed a NDMM-like transcriptomic profile while 5.7% were assigned to the healthy control. We next asked which specific HSPC cell types, including HSCs, (Calvanese et al., 2022; Bandyopadhyay et al., 2024) present with the ML model MM signatures. GMPs showed the highest correlation whilst HSCs showed no association and remained unaffected. Furthermore, gene set enrichment analysis of EMC92 signature (Kuiper et al., 2015) based on ML gene ranking revealed a high correlation to a high-risk MM profile in the HSPCs from treated MM patients. In summary, we show that CD34+ HSPCs from MM patients present with MM gene signatures. We identified that HSPCs acquire a high-risk MM expression profile upon MM treatment, which likely adds to the risk of relapse. This MM profile resides primarily within GMPs, while HSCs stay unaffected by transcriptional changes. Our findings imply that ASCT for MM treatment might be improved by either harvesting CD34+ cells already before treatment or transplanting specific types of CD34+ cells that do not show an MM signature.
SLC25A1 Reprograms Mitochondrial and Fatty Acid Metabolism to Promote the Progression of Acute Myeloid Leukemia
Authors: Yan Xiaojing , Chen Miao , Tao Yuan , Kang Sijing , Yue Pengjie , Ge Rui
Published: December 2024

Metabolic reprogramming is a hallmark of tumors, contributing to tumor initiation, progression and drug resistance. In acute myeloid leukemia, cells usually depend on both enhanced glycolysis and oxidative phosphorylation for their energy needs. The persistence of therapy resistance in some AML patients underscores the urgency of identifying novel metabolic targets. Here, we revealed the vital role of citrate transporter SLC25A1 in maintaining AML cell survival and regulating its drug sensitivity, establishing it as a promising metabolic target for AML therapy.

We first showed that the SLC25A1 expressions were elevated in AML patients both on mRNA and protein level, which were associated with poorer outcomes. Genetic and pharmacological inhibition of SLC25A1 significantly suppressed the proliferation and clonogenicity, promoted apoptosis of leukemic cell lines, as well as the primary AML cells from patients. The specific inhibitor of SLC25A1 had no effect on the proliferation of mononuclear cells (MNCs) from healthy bone marrow at the same concentrations. Our data indicated that SLC25A1 was a potential target for AML therapy, which was further confirmed in AML mice carrying with AE9a or MLL-AF9.

Functionally, inhibition of SLC25A1 revealed the mitochondrial damage and ROS increasing in AML cells by transmission electron microscopy (TEM), mitochondrial membrane potential and extracellular flux analysis, as well as flow cytometry and fluorescence microscopy. The restoration of cell viability by ROS scavenger N-acetyl-L-cysteine (NAC) in the context of SLC25A1 inhibition suggested that SLC25A1 deficiency inhibited cell survival through the upregulation of ROS. Given this, we found that targeting SLC25A1 could enhance the sensitivities of AML cells to two clinically available anti-leukemic drugs, BCL-2 inhibitor venetoclax and chemotherapy drug daunorubicin (DNR).

Mechanically, we found that SLC25A1 was closely correlated with ROS metabolic process, extrinsic apoptotic signaling pathway and oxidative phosphorylation, as well as nicotinate metabolism and fatty acid metabolism, analyzed by gene expression data and metabolic data. We next verified that a series of key enzymes involved in fatty acid metabolism were down-regulated following SLC25A1 KD, accompanied by metabolites in fatty acid synthesized pathway. We also revealed that a self-protection mechanism after stress by up-regulating phosphoglycerides (PGs) and triglycerides (TAGs) in SLC25A1 KD cells, and the effect could be reversed by inhibiting DGAT1, a key enzyme catalyzing the conversion of fatty acids to TAGs.

In conclusion, targeting mitochondrial metabolism and fatty acid metabolism using the SLC25A1 inhibitor might be a potential novel strategy for enhancing AML therapy and overcoming drug resistance.
Implementation of a Mass Spectroscopic Method (Mass-Fix) to Replace Traditional Electrophoresis Methods for Patients with Monoclonal Gammopathies
Authors: Murray David , Kohlhagen Mindy , Coker Jon , Willrich Maria Alice V. , Dasari Surendra , Dispenzieri Angela
Published: December 2024

Background

The detection, isotyping and quantitation of over-expressed immunoglobulin(s) (Igs) (M-protein) is key for diagnosing and monitoring patients with monoclonal gammopathies. Traditionally, our lab has used a combination of agarose gel protein electrophoresis (SPEP) and immunofixation electrophoresis (IFE) for these purposes. In 2018, a mass spectrometric method (Mass-Fix) was implemented to replace IFE for M-protein isotyping. This work documents the validation of a quantitative version (Q-Mass-Fix) to replace both SPEP and IFE.

Methods

24,645 samples submitted for M-protein testing between 7/2018 and 9/2023 with quantitative IgG, IgA and IgM measurements, Mass-Fix analysis and SPEP results were utilized for this study. A separate cohort of 100,000 M-protein negative samples were utilized to mathematically model the spectral shapes IgG, IgA and IgM polyclonal background. To detect M-proteins, software was developed that fits the polyclonal Ig portion of the spectra and M-proteins are detected as peaks above the polyclonal background. M-proteins isotypes are determined by matching similar peaks in spectra in 5 sperate immuno enrichments (IgG, IgA, IgM, kappa and lambda). M-protein quantitation was obtained by multiplying total M-protein isotype quantity by the fraction area of the M-protein in the spectra. M-protein dilutional studies were used to examine the linearity and level of detection of Q-Mass-Fix.

Results

SPEP and Q-Mass-Fix M-protein quantitation had the best agreement between the range of 10 to 40 g/L. Below 10 g/L, Q-Mass Fix had on average, lower M-protein values. Above 40 g/dL, Q-Mass-Fix values were on average, higher than SPEP. The automated isotyping assignment was 98.8 percent consistent with the isotype assigned by visual inspection. Dilutional studies revealed that Q-Mass-Fix quantitation was linear over a range of 70 to 0.04 g/L (slope 1.0, r2 0.99) with an analytical limit of quantitation of 0.1 g/L and a limit of detection of 0.04 g/L.

Conclusion

Q-Mass-Fix represents a significant improvement in comparison to our current SPEP method. Q-Mass-Fix is more capable of removing the interference of polyclonal Igs below 10 g/L and is not subjective to the dye saturation effect at high M-protein levels. The increased analytical measuring range and improved limit of detection of detection could be beneficial in tracking patient's response. Q-Mass Fix is more automated than SPEP and the workflow in our laboratory.
Impact of the Immune Landscape in Follicular Lymphoma: Insights into Histological Transformation in the Rituximab Era
Authors: Enemark Marie Hairing , Jensen Maja Lund , Andersen Maja Dam , Plesner Trine , Hamilton-Dutoit Stephen , Ludvigsen Maja
Published: December 2024

Follicular lymphoma (FL) presents significant clinical heterogeneity, with some patients experiencing transformation into aggressive disease, a key contributor to FL-related mortality. Based on gene expression profiles, this study aimed to provide insights into immunological differences associated with transformation. Gene expression analysis using the NanoString nCounter Tumor Signaling 360 Panel was performed on diagnostic lymphoma samples from 70 FL patients diagnosed in the rituximab era, either non-transforming FL (nt-FL, n=34) or subsequently-transforming FL (st-FL, n=36), with paired high-grade transformed FL (tFL, n=36) samples available. In silico immunophenotyping was performed to infer immune cell infiltration using the CIBERSORTx algorithm. The gene expression analysis revealed 164 significantly differentially expressed genes distinguishing st-FL from nt-FL, generally presenting upregulation of B cell-related genes (CD40, IRF4, RELB), immunosuppressive molecules (IL10, SOCS3), and immune checkpoint molecules (CD276, TIM3). Analysis of immune cell proportions indicated significant differences in infiltrates of M1-like macrophages (p=0.007) and neutrophils (p=0.012) in nt-FL versus st-FL samples. Transformation-free survival (TFS) was associated with high numbers of both these cellular subsets (p=0.006 and 0=0.002, respectively). This was even more evident when combined, with inferior TFS in lymphomas with high infiltrates of both cell types (p<0.001). After transformation, tFL samples showed a reduction in T follicular helper cells (p=0.008) and an increase in immunosuppressive M2-like macrophages and neutrophils (p<0.001 and p=0.028, respectively). By elucidating the distinct molecular and immune landscapes of FL at the time of diagnosis and transformation, this study underscores the importance of immune microenvironment in FL transformation and patient outcome.
Clinician's Knowledge and Awareness of Patient-Reported Outcomes in Immune Thrombocytopenia: Results of a National Survey (ITP NET, GIMEMA Working Group Anemia and Thrombocytopenia)
Authors: Oliva Esther Natalie , Efficace Fabio , Valvano Maria Rosa , Arcuri Paola , Battistini Roberta , Buzzatti Elisa , Clissa Cristina , Danesin Cristina , Salemme Alessia , Sparano Francesco , Chiurazzi Federico
Published: December 2024

Background: Patient-reported outcomes (PROs) are gaining attention in hematological diseases. Nonetheless, clinicians' knowledge of PROs is not widespread. Immune thrombocytopenia (ITP) is an autoimmune hematological disease in which fatigue significantly impacts patients' quality of life (QoL).This national Italian study focused on assessing the knowledge of PROs among hematologists experienced in ITP within the ITP NET network.

Methods: From April 4 to 20, 2023, 63 hematologists from 49 Italian centers were contacted to complete a 63-item online survey covering physician's knowledge of QoL issues and the impact of QoL on physician behaviour, decision making and patient care.

Results: Thirty-nine hematologists (72% from Northern Italy), 72% female, with a mean age of 44 (SD 11), completed the survey. Median years of practice were 15 (IQR 9-24), and median years of experience treating ITP were 6 (IQR 4-16). In the previous 12 months, each respondent saw a median of 50 (IQR 20-150) ITP patients with a median of 40 (IQR 20-77) patients undergoing treatment. Knowledge of PROs was unspecific in 36% and absent in 10%. Thirty-one percent had experience only in research, 8% used PROs in clinical practice, and 15% had knowledge without use. Discrepancies included subsequent 51% reporting frequency of PRO use in research (28% once, 23% several times) and 56% reporting general use of QoL studies in their practice. Clinicians with >17 years of medical practice had more knowledge of PRO studies than those with ≤17 years of practice (72% vs. 38%, respectively; p=0.033), as those with >4 years ITP experience (64% vs. 27%, p=0.037). Physicians with larger ITP practices had more PRO experience in clinical trials (p=0.0345) and more recognition of the value of PROs in severely ill patients (100% vs. 73%, p=0.027), If QoL results were easier to understand, 93% of physicians working in northern Italy would be more likely to use them, versus 64% of physicians working in southern Italy (p=0.042).

Conclusions: There is a general lack of knowledge and use of PROs among hematologists, highlighting the need for further education and awareness. More experienced hematologists and those with larger ITP practices have better knowledge and use of PROs, suggesting that experience plays a crucial role in understanding and implementing PROs. The study serves as a foundation for future initiatives aimed at improving awareness and implementation of PROs, ultimately striving for more comprehensive and patient-focused care in ITP.
Development and Validation of a Machine Learning-Based Early Warning System for Predicting Venous Thromboembolism Risk in Hospitalized Lymphoma Patients Undergoing Chemotherapy: A Multicentre, Retrospective Cohort Study
Authors: Jiang Tingting , Yang Zailin , Tang Xinyi , Zhang Yakun , Chen Shuang , Peng Yu , Zhang Xiaomei , Jun Li , Guo Bingling , Li Jieping , Liu Tingting , Lei Haike , Hu Zuhai , Fang Na , Wei Xia , Liu Xuefen , Chen Yong , Liu Yao
Published: December 2024

Background: Hospitalized lymphoma patients undergoing chemotherapy face a heightened risk of Venous thromboembolism (VTE) due to prolonged treatment and extended bed rest. However, predicting VTE risk early in these patients is challenging. The objective of this study was to develop a machine learning-based early warning system for VTE tailored to this patient group.

Methods: Data were collected from 1,141 patients at four academic medical centers from January 2019 to February 2024. Twelve variables were included and six machine learning algorithms were utilized to develop a VTE early warning system. Models were evaluated based on accuracy, precision, sensitivity, specificity, and area under the curve (AUC). Subsequently, the variables importance of each model was analyzed and comprehensively evaluated by permutation importance analysis. Finally, we visualized the VTE early warning system (VTE-EWS) using a nomogram and compared its performance to the classic Khorana Score (KS) system.

Results: Among the 1141 patients, 799 from Chongqing University Cancer Hospital were evaluated as a training set, while 342 from other three academic hospitals were the external validation set. In the external validation set, all six models demonstrated strong predictive performance, with accuracy ranging from 0.71 to 0.87, AUC from 0.78 to 0.84, sensitivity from 0.62 to 0.73, and specificity from 0.73 to 0.90. Subsequently, we identified the six most important variables-white blood cell (WBC), D-dimer, central venous catheter (CVC), age, chemotherapy cycles, and Eastern Cooperative Oncology Group performance status (ECOG score)-to be included in the nomogram for scoring and visually predicting the risk of VTE. Finally, compared to the KS, VTE-EWS identified more patients at VTE high risk (24/37) and demonstrated a higher probability of clinical benefit within a range of 1% to 78%.

Conclusions: We developed a visual and online VTE-EWS based on machine learning algorithms. Based on simple indicators, VTE-EWS can accurately identify, visually predict, and interpret the risk of VTE in hospitalized lymphoma patients undergoing chemotherapy.
Transient Menin Inhibition Is Sufficient to Induce Irreversible Polycomb-Mediated Silencing and Potent Anti-Proliferative Effects in MLL-Rearranged AML
Authors: Neville Daniel , Ferguson Daniel , Heikamp Emily , Bell Charles , Armstrong Scott A , Gilan Omer
Published: December 2024

MLL-Fusion oncoproteins (MLL-FP) are potent oncogenes that drive an aggressive form of acute myeloid leukaemia (AML). MLL-FP activity can be disrupted by blocking its chromatin occupancy with MLL-Menin inhibitors, or by blocking its biochemical activity through inhibition of DOT1L. Despite these inhibitors having potent anti-leukaemia effects, our understanding of how Menin and DOT1L contribute to MLL-FP function remains incomplete. Here, using functional genomics, we uncover a key role for the non-canonical Polycomb repressive complex 1.1 (PRC1.1) in the silencing of MLL-FP target genes required for the efficacy of Menin and DOT1L inhibitors across a range of in vitro and in vivo MLL-FP models. Mechanistically, Menin inhibition elicits a PRC1.1-dependent selective deposition of the repressive H2AK119ub mark at key MLL-FP target genes, whereas DOT1L inhibition leads to a more global increase in H2AK119ub. Consistent with these differences, we demonstrate that the treatment-induced increase in PRC1.1 activity is specifically associated with loss of H3K79me2 rather than eviction of the MLL-FP or reduction in transcription. In addition, temporal analyses after Menin inhibition revealed that H3K79me2 loss precedes the deposition of H2AK119ub and H3K27me3, which correlates with the gradual downregulation of transcription. Upon this switch in chromatin state, Polycomb target genes undergo stable PRC2-mediated repression, leading to proliferation arrest and the irreversible commitment of leukaemia cells towards differentiation even after early drug withdrawal. Taken together, our findings demonstrate that Polycomb activity is required for the efficacy of these differentiation therapies and highlights why DOT1L is hijacked in MLL-leukaemia with important implications for future therapeutic strategies employing Menin inhibitors.
Validation of Imatinib-Therapy Failure (IMTF) Score in Children with Chronic Myeloid Leukemia in the Chronic Phase
Authors: Zheng Fangyuan , Yuan Mengyao , Zhang Xiaoshuai , Jiang Qian
Published: December 2024

Background About 27% children with chronic myeloid leukemia (CML) still experienced imatinib failure. However, there is no appropriate prediction model to accurately predict the outcome at diagnosis.

Objective To validate the predictive performance of the imatinib-therapy failure (IMTF) score and compare the predictive discrimination of IMTF score with ELTS score in newly-diagnosed children with chronic phase CML receiving imatinib-therapy.

Methods Data of children with chronic phase CML diagnosed at < 18 years receiving initial imatinib (260 to 340 mg/m2 daily) from January 2006 to December 2023 were analyzed. Diagnosis, monitoring and treatment response assessment were according to ELN recommendations. The cumulative incidence of therapy response and imatinib-therapy failure by IMTF score and ELTS score was calculated using the Fine-Gray test considering competing events defined as stopping imatinib-therapy, a switch to a 2nd or 3rd generation TKIs, a transplant or death unrelated to CML. FFS, PFS and OS were calculated by Kaplan-Meier method with the log-rank test.

Result s A total of 324 children were included in this study. 196 (61%) were male. Median age at diagnosis was 13 years (IQR, 9-17 years). 298 (92%) patients achieved a CHR by 3 months. With a median follow-up of 77 months (IQR, 51-107 months) for survivor, the incidences of CCyR, MMR, MR4.0 and MR4.5 at 8 years were 96% (94, 98%), 84% (79, 88%), 60% (53, 67%), and 45% (38, 53%); the 8-year probabilities of FFS, PFS and survival were 80% (75, 84%), 92% (88, 95%), 97% (95, 99%), respectively. By IMTF score, 63 (19%), 62 (19%), 103 (32%), 67 (21%), and 29 (9%) children were classified as very low-, low-, intermediate-, high-, and very high-risk groups. Since there was no significant difference between the low- and intermediate-risk groups (p = 0.941), these groups were combined into a integrated low/intermediate risk group. In the very low-, low/intermediate-, high- and very high risk groups, the 8-year cumulative incidences of imatinib-therapy failure were 5% (0,11%), 18% (12, 24%), 30% (18, 41%) and 43% (24, 62%;p < 0.001). There were significant differences in imatinib-therapy failure rate among the very low-, low/intermediate- and high-risk groups (p < 0.001 - 0.036), but not between the high and very high-risk groups (p = 0.184). By ELTS score, the 8-year cumulative incidences of imatinib-therapy failure in the low- (n = 223, 69%), intermediate-(n = 81, 25%) and high risk group (n = 20, 6%) were 14% (9, 19%), 32% (22, 43%) and 36% (14, 58%) (p < 0.001), but no significant difference between the intermediate- and high risk group were observed (p = 0.590). The IMTF score exhibited a higher C-statistic values of 0.680 than the ELTS score (0.644).

Conclusion IMTF score had better predictive discrimination for imatinib-therapy failure in children with CML-CP compared to ELTS score.
Evaluation of Risk Assessment Models for Predicting Venous Thromboembolic Events in Cancer Patients Using Implanted Central Venous Access Devices
Authors: Ma'koseh Mohammad , Farfoura Heba , Abunasser Mahmoud , El-Atrash Maryam , Zayed Anas , Abdel Rahman Zaid , Hamdan-Mansour Renad , Al-Jaghbeer Mohammed J , Abdel-Razeq Hikmat
Published: December 2024

Introduction

Cancer patients are at an increased risk of venous thromboembolic events (VTEs). Implanted central venous access devices (ICVADs) are commonly used for administering chemotherapy and may further increase the risk of VTEs. Our study aims to evaluate the incidence and predictors of VTEs, utilizing the commonly used VTE risk assessment models (RAMs), in cancer patients receiving chemotherapy through ICVADs.

Methods

Medical records data of adults (≥18 years) who received chemotherapy through ICVADs from January 2022 until December 2023 were retrospectively reviewed. Patients on therapeutic anticoagulation at the time of ICVAD insertion were excluded.

Khorana, COMPASS and ONCOTEV RAMs were calculated as previously described. No routine imaging studies were performed to detect asymptomatic VTEs.

VTEs were defined as venous thrombotic events reported after insertion of ICVAD until six months post-removal. Patient characteristics and VTE frequency were compared using Fisher's exact and Mann Whitney U tests. Overall survival (OS) was calculated using the Kaplan Meier method and compared using log-rank test.

Results

A total of 264 patients with a median age of 56 (range: 22-81) years were included in the analysis. The most common cancer types were gastric (n=78, 29.5%), colorectal (n=61, 23.1%), pancreatic (n=51, 19.3%), and breast (n=45, 17%). 148 (56.1%) had metastatic disease.

According to RAMs, 72 (27.3%) had Khorana score >2, 131 (49.6%) had COMPASS score >6, and 42 (15.9%) had ONCOTEV score >1.

During a median follow-up of 11.8 (range: 0.2-29.6) months, VTEs were reported in 51 (19%) of patients, occurring at a median of 3.6 (range: 0.1-20.3) months after ICVAD insertion. 27 (53%) were line related. VTEs were more frequent in patients with history of diabetes (p=0.02), a white blood cell count > 11 X 103/μL at the time of line insertion (p=0.037), and when the time from diagnosis to line insertion was >6 months (p=0.038).

When evaluating the three RAMs for correlation with VTE, there was no significant correlation between VTEs and Khorana (VTE rates: 16% in the low-risk group and 22% in the high-risk group, p=0.075) or COMPASS (VTE rates: 17% in the low-risk group and 21% in the high-risk group, p=0.4) risk groups. However, VTEs were significantly correlated with the ONCOTEV risk (VTE rates: 17% in the low-risk group vs 31% in the high-risk group, p=0.037). The occurrence of VTEs was associated with inferior OS, with 1-year OS rates of 63% vs 75%, (p=0.028).

Conclusion

Cancer patients receiving chemotherapy through ICVAD are at high risk for VTEs. Among the assessed models, ONCOTEV RAM was the most effective to predict VTEs in this group of patients. Further studies are needed to explore the potential role of prophylactic anticoagulation in this context.
ULK1 Regulates the Proliferation and Differentiation of Erythroid Progenitors
Authors: Chen Yijin , Zheng Haiqiong , Luo Qian , Huang Mengmeng , Zhang Meng , Qian Pengxu , Huang He
Published: December 2024

Erythropoiesis depends on the proliferation and differentiation of erythroid progenitors. However, underlying mechanism of balance of these two processes in erythroid progenitors is largely unknown. Unc-51 like autophagy activating kinase 1 (ULK1) drives the maintenance and differentiation of stem cells, and ULK1 keeps low expression in erythroid progenitors and rapidly increases during differentiation. Therefore, we hypothesize that the proliferation of progenitor cells and their transition to differentiation is regulated by increasing expression of ULK1.

To explore the function of ULK1 in erythroid progenitors, ULK1 was inhibited by inhibitor SBI-0206965 (SBI) or ULK1 shRNA at day 4 (D4) during UCB-derived erythroid differentiation in vitro. Quantifying treated progenitors by colony-forming assay and flow cytometry indicated ULK1 influenced erythroid differentiation at CFU-E cell stages. The results showed that the differentiation of CFU-E cells was depressed, but their proliferation was enhanced when ULK1 was inhibited. Meanwhile, opposite results were observed when over-expressing ULK1 in erythroid progenitors. Moreover, the number of mitochondria, cell cycle, and apoptosis were not impacted by inhibiting ULK1. Further, the proteomics and RNA-sequencing data showed that the expression of AHR signaling-related genes like CYP1B1, CYP1A1, and AHRR were up-regulating by ULK1 inhibition suggesting the activation of AHR. Both AHR antagonist StemReginin 1 and AHR shRNA rescued the effects of ULK1 repressed in erythroid progenitors. We also observed that the number of AHR in the nucleus was increased, though the total expression of AHR was slightly decreased due to ULK1 inhibition, indicating that ULK1 plays a role in regulating AHR activation.

Mechanically, we demonstrated that ULK1 directly interacted with AHR in cytoplasm, and phosphorylates Ser-73 of AHR. Then phosphomimetic (Ser-73 –> Asp, AHRS73D) and loss-of-function (Ser-73 –> Ala, AHRS73A) mutants were generated respectively to study whether Ser-73 of AHR can impact differentiation erythroid progenitors. Compared to AHRWT, AHRS73D promoted the differentiation, while AHRS73A showed the opposite result. Meanwhile, we observed that more AHRS73D were located in the cytoplasm, while more AHRS73A were located in the nucleus. These results were consistent with the observations of ULK1. Together, our data indicate that during the differentiation of erythroid progenitors, increased expression of ULK1 inhibited AHR translocated to the nucleus by phosphorylating Ser-73. On the contrary, low expression of ULK1 benefits AHR translocated to the nucleus, which enhances the proliferation of erythroid progenitors.
Characterizing the Microbiome-Immune Axis in Hematopoietic Stem Cell Recipients
Authors: Becker Sabrina A. , Enssle Julius C. , Wilke Anne C. , Pfundt Sylvia , Zeltner Nico , Jankovic Ana , Wolf Sebastian , Wichelhaus Thomas A. , Kempf Volkard A. J. , Vehreschild Maria J. G. T. , Serve Hubert , Oellerich Thomas , Steffen Björn , Scheich Sebastian
Published: December 2024

Hematologic diseases such as malignant lymphomas and multiple myelomas are potentially life-threatening diseases, that can be treated with high-dose chemotherapy (HDCT) and subsequent autologous stem cell transplantation (HSCT). However, these therapies lead to severe immunosuppression and an increased risk of infection. Recent studies have demonstrated the importance of gut microbiome diversity for treatment efficacy and preclinical data suggests a direct crosstalk between the microbiome and the immune system of the host. Although it is known that the gut microbiome can impact immune cell subsets, the impact of the microbiome on the immune cell reconstitution in transplant recipients is largely unknown. Therefore, we investigated the effect of the gut microbiome on immune cell reconstitution and outcomes in patients receiving HDCT and autologous HSCT.

We collected paired fecal and blood specimens from patients at five defined time points before and up to 100 days after autologous HSCT. The microbiome diversity and composition were assessed using 16S amplicon sequencing and in a subset metagenomics sequencing. To characterize the immune cell changes, we set up a large flow cytometry panel defining NK, B and T cells including a variety of their subsets e.g. regulatory T cells.

Overall, we analyzed 815 stool samples from 222 patients and 151 matched blood samples from 90 patients. Our data show that microbiome diversity decreases rapidly after autologous stem cell transplantation, hitting a nadir around 14 days post-HSCT, and recovers thereafter up to day 100, similarly in lymphoma and multiple myeloma patients.

To model the microbiome data over time, we utilized multi-omics factor analysis accounting for temporal relationships (MEFISTO) among the samples. Here, the bacterial family of Prevotellaceae was frequently represented among the top variance-explaining latent factors. Immune cell profiling revealed that patients with a high abundance of Prevotellaceae had significantly higher CD4+ lymphocyte counts (p = 0.04) and higher activated CD4+ counts after autologous HSCT. Furthermore, high Prevotellaceae abundance was associated with a significantly increased overall survival (p = 0.02) after autologous HSCT. Notably, the main cause of death in these patients were infections (47 %).

Our findings indicate an association between specific bacterial families and certain immune cell subsets in patients receiving autologous HSCT, that translates into clinical outcome differences. This data opens the rationale for microbiome reshaping studies in stem cell recipients to prevent infectious related deaths.