bluuebunny/crossref_metadata_2025 huggingface dataset:
Prognosticating Survival Outcomes Within a Genetically Heterogeneous Population of KMT2A-Aberrant Acute Leukemia
Authors: Reimonn Cassandra , Ramanathan Ramkarthic , Miron Patricia Minehart , Higgins Anne , Hutchinson Lloyd , Xie Yiyu , Cerny Jan , Patel Shyam Ajay
Published: December 2024

KMT2A aberrations are frequently observed in both therapy-related and de novo acute leukemias and are often associated with a poor prognosis. However, significant heterogeneity exists among patients with KMT2A-aberrant acute leukemia. In this study, we attempt to further differentiate this heterogeneity by assessing the overall survival (OS) outcomes of 23 patients from the UMass Leukemia Registry with KMT2A-aberrant acute leukemia, stratified by leukemia type (lymphoblastic vs. myeloblastic vs. ambiguous lineage), concurrent mutations, and aberrancy type (translocation vs. other). For patients with KMT2A translocation, further stratification was performed to compare outcomes across various KMT2A translocation partners. Median age at diagnosis was 58 (IQR = 25.5 years), and median OS was 6.08 months (IQR = 28.7 months) for the entire group. No significant difference in OS was observed between KMT2A-aberrant acute myeloid leukemia (median OS 5.32 months), KMT2A-aberrant acute lymphoblastic leukemia (median OS 6.27 months), and KMT2A-aberrant acute leukemia of ambiguous lineage [(median OS 74.3 months), p = 0.77]. Additionally, no significant difference in OS was detected between patients with KMT2A translocation and other KMT2A aberrancy types such as frameshift mutation or amplification (p = 0.57). Among patients with KMT2A translocations, translocation partners included AF9 (n=5), AF4 (n=4), ENL (n=3), AF6 (n=3), ELL (n=2), AF10 (n=2), and SEPT2 (n=1). No significant difference in OS was detected based on KMT2A translocation partner (p = 0.91). Finally, the most common concurrent mutations observed were KRAS (n=6), NRAS (n=6), and TP53 (n=5). Patients with concurrent TP53 mutations had significantly decreased OS compared to those without (p = 0.0024), whereas no significant difference in OS was observed in patients with concurrent KRAS (p = 0.15) and NRAS (p = 0.62) mutations compared to those without. The above data supports that there are no significant differences in OS/prognosis of KMT2A-aberrant acute leukemia based on leukemia subtype, aberrancy type, translocation partner, or concurrent KRAS/NRAS mutation status, but co-mutation with TP53 results in significantly inferior OS. This is consistent with our prior reports from the UMass Leukemia Registry (Patel SA et al., Leuk Lymphoma 2021) (Patel SA et al., EJHaem 2023). Additional larger studies should be conducted to validate these findings and further develop menin inhibitors that might improve overall survival for this high-risk subset of patients.
Prospective Evaluation of the Prevalence of Pulmonary Embolism in Patients with Chronic Obstructive Pulmonary Disease Exacerbation: Challenges and Feasibility
Authors: Mai Vicky , Fergusson Dean , Le Gal Gregoire , Mulpuru Sunita
Published: December 2024

Introduction: Data on the prevalence of pulmonary embolism (PE) in patients with acute exacerbation of chronic obstructive pulmonary disease (AECOPD) in North America is lacking, with most of the prospective studies conducted in Europe and Asia. The aim of this study is to describe feasibility of prospective collection of data evaluating the prevalence of PE in patients admitted for AECOPD in Canada, and to describe challenges of this approach.

Methods: This is a secondary analysis of a prospective pilot patient cohort aiming to assess the diagnosis of AECOPD among hospitalized patients. Items of the Wells PE and revised Geneva scores were prospectively collected. All patients underwent D-dimer testing within 24 hours of admission. All patients were followed for 3 months. Feasibility outcomes were prospective data collection to evaluate the prevalence of PE, D-dimer level measurement and 3-month follow-up. The study was considered feasible if >=90% of the patients had prospective data collected to assess the prevalence of PE, >=80% of the patients had D-dimer level measured and their 3-month follow-up, respectively. Challenges were also assessed.

Results: Twenty patients were included. According to the Wells PE score, 16 (80.0%), 4 (20.0%) and 0 (0.0%) patients had low, moderate and high clinical pre-test probability (c-PTP), respectively. According to the revised Geneva score, 2 (10.0%), 17 (85.0%) and 1 (5.0%) patients had low, intermediate and high c-PTP, respectively. Based on age-adjusted D-dimer cut-off, 7 and 11 patients had negative and positive D-dimer, respectively. All patients (100.0%) had prospective data collection to assess the prevalence of PE. D-dimer level was available for 18/20 (90.0%) patients and 3-month follow-up evaluation was performed in 16/20 (80.0%) patients. Challenges included the difficulty in recruiting patients since it is a population with many comorbidities, adequate c-PTP assessment using validated clinical decision rules, absence of consistent PE diagnostic strategy use since patients were mostly investigated according to the physician's impression and adequate 3-month follow-up since 25% of the patients had some protocol deviation for the follow-up. No PE was diagnosed during the entire study period.

Conclusion: This pilot study shows the feasibility in prospectively collecting data to evaluate the prevalence of PE in patients admitted for AECOPD in Canada. Also, challenges were assessed and included recruitment of patients, adequate c-PTP assessment, consistent PE diagnostic strategy use and adequate 3-month follow-up.
Sequential Infusion of Anti-CD19 and Anti-BCMA Chimeric Antigen Receptor T-Cells: A Promising Treatment Strategy for Refractory Immune-Mediated Platelet Transfusion Refractoriness
Authors: Ma Yunju , Cui Qingya , Yang Chunxiu , Cui Wei , Li Mengyun , Li Zheng , Dai Haiping , Kang Liqing , Yu Lei , Tang Zaixiang , Fu Jianxin , Jiang Min , Wu Depei , Tang Xiaowen
Published: December 2024

Background: Immune-mediated platelet transfusion refractoriness (iPTR) represents a challenging, high-cost, and life-threatening problem among patients with hematological malignancies. Refractory iPTR (riPTR), which exhibits resistance to multiple therapeutic plasma exchange, high dose of IgG and rituximab, may result from incomplete elimination of allo-specific B cells and long-lived plasma cells. Therefore, sequential anti-CD19 and anti-B cell maturation antigen (BCMA) chimeric antigen receptor (CAR) T-cells were used to treat riPTR.

Methods: In this prospective single-arm clinical trial, patients with riPTR received a combination of murine anti-BCMA CAR T-cells and murine anti-CD19 CAR T-cells at a dose of 2×107 cells/kg. The efficacy and safety of this treatment were assessed. This study was registered at clinicaltrials.gov (identifier: NCT04846439).

Results:Twelve patients with riPTR received CAR T-cell treatment with a median follow-up of 33.5 (range, 13.5-34.5) months. Among these 12 patients, the median of the maximum mean fluorescence intensity (MFI) values of anti-human leukocyte antigen (HLA) antibodies decreased by 91.1%, from 15319 (range, 10807-20374) before CAR T-cell infusion to 1369 (range, 997-2923) at 4 months post-infusion (P=0.0001). And four months after CAR T-cell infusion, the median number of anti-HLA class I antibodies with an MFI > 5000 decreased significantly from 18 (range, 3-41) before infusion to 0 (range, 0-0) post-infusion (P=0.0005). Anti-HLA antibodies in all patients turned negative within 5 months after CAR T-cell therapy and have maintained negative for up to 30.5 months to date. After CAR T-cell treatment, the medians of the amount of platelet transfusions within 7 days and the duration of platelet count below 10×109/L were significantly lower (P = 0.0002, P = 0.001). Meanwhile, there was a significant increase in 14-hour corrected count increment (CCI) and no occurrence of 14-hour CCI below 5×109/L in all patients after the administration of CAR T-cells. RiPTR was completely corrected in all patients and never occurred during subsequent chemotherapy or transplantation. All patients who underwent transplantation achieved successful neutrophil and platelet engraftment. No patient developed grade ≥ 3 cytokine release syndrome. All patients experienced transient grade ≥ 3 cytopenia. Only 2 patients (16.7%) experienced infection. All adverse events were reversible and manageable.

Conclusions:CAR T-cell therapy could be well-tolerated and highly effective in overcoming riPTR and eradicating allo-antibodies, making it applicable to antibody-mediated diseases.
Runx1+23 Enhancer Reporter Marks Erythro-Megakaryocyte Lineages in Human ESC Differentiation Culture
Authors: Syed Alina , Nishida Chika , Kobayashi Michihiro , Wenzel Pamela L. , Davis Brian , Yoshimoto Momoko
Published: December 2024

Runx1 is an essential transcriptional factor for hemogenic endothelial cells (HECs) to transit to hematopoietic stem/progenitors in mammalian embryos. In mice, it has been shown that Runx1 +23kb enhancer region (Runx1+23) marks HECs in the embryo and HSCs in the adult bone marrow (BM) specifically.

In order to identify HECs in human embryonic stem cell (ESC) differentiation culture, we utilized Runx1+23 enhancer-reporter hESC lines; H9 Runx1+23 Ds-Red and H1 Runx1+23 venus. We differentiated these Runx1+23 ESCs into hematopoietic lineage on laminin coated plate. We examined CD34+CD43- EC and CD34+CD43+ hematopoietic progenitor (HPC) productions as well as CD235a+ RBC and CD43+CD41 megakaryocyte production on day 6, day 8, and day 10 culture. We found robust Runx1+23+ Ds-Red (denoted Runx1+23) expression on CD235a+ RBCs, CD43+CD41+ cells, and CD34+CD43+ HPCs. However, Runx1+23 expression was not detected in CD34+CD43- EC population. When we gated Run1+23 population, they were relatively large, did not express CD144 EC markers, but expressed CD43 and CD41. Cytospin preparation of sorted Runx1+23+ cells showed that Runx1+23+ cells contained megakaryocytes and erythroblast. We also sorted Runx1+23 positive or Runx1+23 negative CD34+CD43+ HPCs or CD34+CD43-ECs and plated them onto Methylcellulose culture to test colony forming ability and onto MS5 stromal cells for testing B cell potential. Runx1+23+ cells displayed a greater number of megakaryocytes colony-forming cells compared to Runx1+23 negative cells and expressed erythro-megakaryocyte genes including GATA1, GATA2, TAL1, and hemoglobin genes. On MS5 cell co-culture, only Runx1+23 negative ECs produced CD19+B-cells and CD56+ NK cells. Finally, scRNA-sequencing of day 10 differentiation culture revealed that Runx1+23+ cells were exclusively expressed in the erythro-megakaryocytes populations.

These results indicated that Runx1+23 enhancer region is exclusively active in erythro-megakaryocyte lineage specification in human ESC differentiation culture, different from mouse system, suggesting different usage of Runx1+23 enhancer region between mice and humans.
R-CVP in Nodular Lymphocyte Predominant Hodgkin Lymphoma in Children
Authors: VU Thi An , Olivier Laura , Simonin Mathieu , Boudjemaa Sabah , Jonca Bénédicte , Rigaud Charlotte , Garnier Nathalie , Mansuy Ludovic , Curtillet Catherine , Treguier Pauline , Neumann Florent , Heritier Sebastien , Dourthe Marie-Emilie , Landman-Parker Judith
Published: December 2024

Introduction : A strategy of therapeutic de-escalation has been proposed for children with localized stage nodular lymphocyte predominant Hodgkin Lymphoma (NLPHL) over the past twenty years : either local surgical treatment and watch and wait after complete resection of an isolated adenopathy or low intensity chemotherapy ±Rituximab (R). Here, we provide results on the management and efficacy of R-CVP in children with early stage NLPHL.

Methods : A retrospective multicenter national study was carried out in the centers of the French Children's Cancers Society (SFCE) to evaluate the tolerability and efficacy of 3 courses of Rituximab 375 mg/m2-Cyclophosphamide (500 mg/m2 day 1), Vinblastine 6 mg/m2 day 1 and 8, Prednisolone (40 mg/m2 day 1-8) (R-CVP) treatment in NLPHL patients under 18 years of age diagnosed between January 2015 and December 2023. NLPHL diagnosis was confirmed by hematopathologist review and the staging was evaluated by FDG-TEP/CT in all patients.

Results : Twelve children, median age at diagnosis 12 years, (range 7-16 years); boys n=10 received R-CVP between January 2019 and December 2023 in SFCE centers. No patient were treated by R-CVP before 2019 in France. Patients were classified as stage IA n=4 , stage IIA n=7 and stage IIIA n=1. Six patients were treated by R-CVP as first line treatment at initial diagnosis, 3 cases for residual adenopathy post initial surgery, 3 cases after relapse (median time to relapse 9 months) after primary treatment by local surgery and/or radiotherapy. All patients achieved complete remission. In 10/12 cases R-CVP course was performed every 21 days, only one patient received R-CVP every 15 days. Stage IIIA received 4 courses of R-CVP. With a median follow up of 23 months (8-45 months) EFS and OS is 100%. Forty-two percent presented neutropenia at d17 (nadir 0.3 G/l and 0.6 G/l first and third course, respectively). No patient needed blood transfusion support. Grade II mucositis and febrile neutropenia were reported as well as grade I/II peripheral sensitive neuropathy, constipation or digestive disorders

Conclusion: Our results show that R-CVP is an effective chemotherapy regimen in children with localized stage NLPHL. Limited hematological toxicities and peripheral neuropathy were reported during the treatment
DOCK1 Expression Enhances the Activity of the Notch Signaling Pathway and Promotes the Proliferation and Survival of Acute Myeloid Leukemia Cells
Authors: Lin Ming-En , Hsu Yueh-Chwen , Chiu Hsueh-Ling , Yao Chi-Yuan , Liu Chia-Hua , Kao Chein-Jun , Li Chang-I , Yuan Chang-Tsu , Yang Yi-Tsung , Hou Hsin-An , Chou Wen-Chien , Tien Hwei-Fang
Published: December 2024

In spite of recent advances in genetic risk categorization and treatment, the available options for patients of acute myeloid leukemia (AML) resistant to standard chemotherapy remain limited. There is an urgent need to address this problem. In a prior study, we identified DOCK1 gene expression as an independent prognostic marker by means of microarray analysis of gene expression in an AML cohort adopting the ELN 2010 risk stratification. To validate the prior results and explore deeper into the molecular mechanisms, we analyzed the clinical impacts of DOCK1 expression on a new cohort of 341 de novo non-M3 AML patients diagnosed from February 2004 to December 2016. These patients were risk-stratified using the 2022 ELN classifications, and we utilized more advanced RNA sequencing technology for our analysis. All of the patients were treated with standard chemotherapy. We demonstrate that higher DOCK1 expression still correlates with unfavorable cytogenetics and mutations, and it is an independent adverse prognostic factor.

We subsequently investigated its pathogenesis using cell lines, xenograft models, and a hematopoietic cell-specific Dock1-conditional knockout model in mice. Cell line experiments showed positive roles of DOCK1 in proliferation and viability. This observation is further corroborated by xenograft experiments, in which mice transplanted with DOCK1-knockdown cells had longer survival times than those engrafted with control cells. Although mice with a hematopoietic cell-specific knockout of Dock1 exhibited minimal phenotypic changes, the loss of Dock1 ameliorated MN1-induced AML in vivo. Transcriptome analyses of cell lines and human samples consistently revealed upregulation of NOTCH signaling pathway in cells with higher DOCK1 expression, as evidenced by higher expression levels of HES1, a direct downstream target of NOTCH pathway. Single-cell RNA sequencing of MN1-induced mouse leukemia cells within Dock1-conditional knockout background also revealed a unique cluster of cells that predominantly existed in DOCK1-wild type rather than Dock1-conditional knockout mice and exhibited the highest expression of DOCK1, MN1, and HES1 among all clusters. These cells showed upregulation of stem cell function, G2M checkpoint, E2F targets, and Myc signaling pathways, consistent with the phenotypes of DOCK1 in our study. Our findings highlight the role of DOCK1 expression in the pathogenesis of AML and suggest DOCK1 as a potential therapeutic target for this group of patients.
Serum Amyloid-a (SAA) Is an Optimal Acute Phase Protein for Cytokine Release Syndrome in CAR-T Patients Exposed to Tocilizumab
Authors: Galli Eugenio , Pansini Ilaria , Corrente Alessandro , Baroni Silvia , Chiusolo Patrizia , Sica Simona , Sorà Federica
Published: December 2024

Patients treated with CAR-T and experiencing cytokine release syndrome (CRS) are often exposed to tocilizumab (anti IL6R). After the administration, IL-6 peaks in serum determinations and looses its predictive power to associate with severity of CRS.

This study aimed to explore the role of Serum Amyloid A (SAA) as a potential biomarker for monitoring early toxicities, particularly in patients treated with CAR-T cells. The analysis included 34 consecutive patients treated with CAR-T therapy at our institution from May 2023 to June 2024. SAA levels were measured at multiple time points: before lymphodepletion, immediately before CAR-T infusion, and every other day up to day 14 post-infusion. Additional acute phase proteins (APPs), including C-reactive protein (CRP), interleukin-6 (IL6), interleukin-2 receptor (IL2R), procalcitonin, ferritin, soluble suppression of tumorigenicity 2 (SST-2), and soluble urokinase plasminogen activator receptor (SUPAR), were also assessed at these intervals.

The study found that median baseline SAA levels were elevated above the upper limit of normal and exhibited wave-shaped kinetics post-CAR-T infusion, peaking on day 4 and declining thereafter. SAA levels were significantly higher before lymphodepletion compared to day 14 levels. When comparing early responders to non-responders, non-responders showed higher median SAA levels before lymphodepletion and at the day 4 peak. SAA may therefore mark tumor-associated inflammation also in this context.

SAA levels were significantly higher in patients experiencing CRS of any grade, with a clear correlation between elevated SAA and CRS severity. However, no such correlation was found for immune effector cell-associated neurotoxicity syndrome (ICANS). Strong associations were also observed between SAA and other APPs, particularly CRP, procalcitonin, ferritin, SST2, and SUPAR, with moderate trends observed for IL6 and IL2R.

In a focused analysis on patients previously exposed to tocilizumab, SAA emerged as the best-performing APP in identifying patients at risk of clinically relevant (grade 2 or higher) CRS, with a ROC analysis yielding an AUC of 0.86, being superior to CRP, IL6. IL2R, procalcitonin, and ferritin. A cut-off of 70 mg/L for SAA was identified, offering a sensitivity of 87% and specificity of 83% in distinguishing patients with grade 2-4 CRS.

SAA is a valuable biomarker for assessing inflammation and CRS in CAR-T treated patients, particularly in those previously treated with tocilizumab, where traditional markers like IL6 may be less reliable. SAA measurement could guide clinical monitoring and intervention strategies, especially in settings where early detection of severe CRS is critical. Further research is recommended to confirm these findings and explore the broader applicability of SAA in CAR-T therapy.
Transcriptome Data Analysis of 80 B ALL Patients in China
Authors: Lin Yuehui , Liu Dan , Xia Yu , Xia Yu , Wu Tong , Zheng Qinlong
Published: December 2024

Introduction: Recently, the 5th edition of the World Health Organization Classification of Haematolymphoid Tumours (WHO-HAEM5) and the 2022 International Consensus Classification (ICC) has proposed up to 26 genetically defined B cell acute lymphoblastic leukemia（B-ALL）entities and subtypes. However, several of them are hard to recognize with traditional diagnostic tests. Sequencing methods like whole genome sequencing or transcriptome analyses of B-ALL leukemia cells have been widely used in clinical studies and demonstrated effective tools that can provide comprehensive molecular profiling including driver genetic lessions as well as gene expression patterns.

Methods: Transcriptome sequencing analyses were performed for 80 retrospective B-ALL cases admitted to our medical team between October, 2020 and July, 2024. Among them, 79 were refractory or relapse（RR） even after hematopoietic stem cell transplantation（HSCT）, while only one was newly diagnosed. Testing specimens including bone marrows, blood or tissues with at least 5% leukemia cells. Oncogenic driver events of gene fusions, chromosome large scale copy number variation, and hotspot gene mutations were analyzed with an inhouse workflow. Subtypes were identified based on the gene expression profiles and characterized with those oncogenic mutations.

Results：All patients：37 females（F） and 43 males（M）, age from 2 to 70 years old.The genetic testing results show that: 15 pts were Unclassed（15/80，18.75%），M:F 7:8 with a median age of 22；7 pts were high hyperdiploid（7/80,8.75%）,M:F 4:3 with a median age of 5；3 pts were low hypodiploid（3/80,3.75%）：M:F 2:1 with a median age of 16；4 pts were KMT2A（4/80,5%）,M:F 1:3，with a median age of 53.5;9 pts were ZNF384（9/80,11.25%）,M:F 6:3, with a median age of 31；5 pts were MEF2D（5/80,6.25%）,M/F 0:5，with a median age of 30；9 pts were Ph positive（9/80,11.25%）,M:F 2:7 with a median age of 55；11 pts were Ph like（11/80,13.75%）,M:F 10:1 with a median age of 20；2 pts were ETV6-RUNX1（2/80，2.5%），M:F 1:1 with a median age of 15；7 pts were DUX4（7/80，8.75%）：F:M 2:5 with a median age of 18;2 pts were PAX5alt（2/80，2.5%）,M:F 2.0 with a median age of 26; 2 pts were PAX5 P80R（2/80，2.5%）,M:F 2.0 with a median age of 19.5; 3 pts were TCF3-PBX1（3/80，3.75%）,M:F 3.0 with a median age of 21;1pts was HLF,M,21.

Conclusions：

Our transcriptome sequencing indicate: only about 20% of B ALL pts belong to be unclassed, while remaining 80% of pts have clear molecular subtypes. This suggests that hematologists should try to perform whole genome sequencing or transcriptome analyses as much as possible when patients first diagnosis or relapse. This can help hematologists better understand diseases, also help patients achieve accurate diagnosis and obtain the best treatment. So far there are still many unknown molecular subtyping that need to be discovered and we still need further research in future.
The Efficacy and Safety of Huaiqihuang Granules in the Treatment of Childhood Chronic Primary Immune Thrombocytopenia: A Multicentre, Randomized, Double-Blind, Placebo-Controlled Clinical Study
Authors: Han Juan , Zhou Fen , Jin Runming , Shi Xiaodong , Li Jianxin , Fang Yongjun , Yuan Xiaojun , Luo Xuequn , Li Jianqin , Sun Lirong , Liu Wenjun , Yan Mei , Liu Yufeng , Liang Changda , Jiang Hua , Wu Runhui
Published: December 2024

The efficacy and safety of Huaiqihuang Granules in the treatment of childhood chronic primary immune thrombocytopenia: a multicentre, randomized, double-blind, placebo-controlled clinical study

* Correspondence:

Fen Zhou, daisy_may@163.com

Runming Jin, jinrunm@qq.com

Abstract

Background: Chronic immune thrombocytopenia (CITP) is a hemorrhagic disease that seriously affects the quality of life of children and their families. Huaiqihuang Granules, a traditional Chinese medicine, has certain value in the treatment of ITP patients in the preliminary clinical findings. To investigate the efficacy and safety of Huaiqihuang Granules in children with CITP, we conducted a multicenter, randomized, double-blind, placebo-controlled study.

Methods: Patients with CITP were treated with Huaiqihuang Granules or with matching placebo in a randomized, double-blind, 24-week treatment period. Patients who received placebo and completed 24 weeks of treatment switched to receive Huaiqihuang Granules, and patients treated with Huaiqihuang Granules in the double-blind period continued Huaiqihuang Granules during the following open-label 24-week treatment.The patients were followed up for 96 weeks. Clinical effective (defined as those achieving a platelet count of≥30×109/L and at least twofold more than the basal platelet count, or the degree of bleeding decreased by one or more grade) was evaluated.

Results: Huaiqihuang Granules was superior to placebo in achieving a platelet response and in reducing the bleeding risk. The clinical effective rate of Huaiqihuang Granules group (71.74% in 24-week) was higher than that of the Placebo group (45.65% in 24-week, p=0.0013). The most common severe adverse events were respiratory tract infection (7.8%) and immune thrombocytopenic purpura (6.9%) .

Conclusions: In CITP children, Huaiqihuang granules can improve the clinical effectiveness especially the degree of bleeding, and shows good safety in long-term treatment.

Keywords: Chronic immune thrombocytopenia, Huaiqihuang Granules, efficacy; safety
Maximizing Safety and Efficacy in Hematopoietic Stem Cell Gene Therapy
Authors: Giommetti Aurora , Barra Arianna , Bosbach Christin Laura , Olevska Valeriya , Dzionek Julia , Soltenborn Stephanie , Hebbeker Felix , Oberbörsch Svenja , Martinez Carrera Lilian Andrea , Jelveh Nojan , Bisdorf Kevin , Matzke Simona Karolin , Khorkova Svetlana , Wöhle Christian , Tomiuk Stefan , Bosio Andreas , Papanikolaou Eleni
Published: December 2024

Hematopoietic stem cell (HSC) gene therapy has been recently successfully employed for certain rare blood disorders including hemoglobinopathies, which resulted even in marketing authorizations such as Lyfgenia™ and Casgevy™. Despite these advancements, the appearance of leukemic events in the case of Sickle Cell Disease combined with the therapy's high price render the wider access, and hence the overall applicability of this approach, doubtful. Therefore to make HSC gene therapy more widely applicable, an effort is required toward robust and less laborious manufacturing processes that ensure reproducibility and predictability to maximize the safety as well as providing the capacity to generate engineered HSCs at large scale. Automation is the key to achieve these goals and, to this end, the CliniMACS Prodigy® as an automated, closed, GMP-compliant platform for HSC transduction is an ideal candidate to streamline cell processing. To this end, the scope of this project is a) to provide alternative methodology for genetic engineering of CD34+ cells by testing several transduction enhancers to achieve a clinically translatable transduction efficiency both in the manual process (open steps) and in large scale on the CliniMACS Prodigy and b) to assess the safety of both approaches. In our first line of experiments, comparable cell recovery and viability between the small and large scale conditions were observed, however superior performance of the automated platform was repeatedly shown in terms of transduction efficiency compared to open manual steps. Additionally, significantly higher transduction efficiency was achieved in the presence of transduction enhancers, preserving cell recovery and viability, without altering the stem cell clonogenic capacity. Based on these results, we propose a combined strategy of transduction enhancers on the automated platform to achieve significantly higher transduction efficiency. Ongoing experiments investigate the mechanisms involved in better performance of lentiviral transduction on the automated platform with molecular analysis via Next Generation Sequencing (NGS) and respective bioinformatics analysis.
Treatment of Chronic ITP with Avatrombopag: A Multicentric Real-World Experience
Authors: Troiano Mario , Giordano Claudia , Rossi Elena , Scalzulli Potito Rosario , Mettivier Laura , D'Ambrosio Alessandro , Mazzoni Daniele , Vatiero Erika , Selleri Carmine , Pane Fabrizio , Frigeri Ferdinando
Published: December 2024

INTRODUCTION

Chronic immune thrombocytopenia (ITP) is a bleeding disorder characterized by a low platelet count that persists for 12 months or more. Initial treatments usually include corticosteroids or intravenous immunoglobulin (IVIG). For patients who are refractory or relapsed, or who may not adhere well to treatment, second-line therapies come into play. These include: i) Thrombopoietin receptor agonists (TPO-RAs) such as Romiplostim, Eltrombopag, or Avatrombopag, and ii) Rituximab or other immunosuppressive agents such as azathioprine, mycophenolate mofetil, or ciclosporin. Splenectomy, although an option, is generally considered a last resort for patients who do not respond to or tolerate other treatments.

AIM OF THE STUDY

To assess the efficacy and safety of Avatrombopag in patients with chronic ITP, stratified by prior exposure to other TPO-RAs.

RESULT

Fifty-seven patients with immune thrombocytopenia (ITP) treated with Avatrombopag were enrolled from five Italian hematology units. The median age at the start of treatment was 60.8 years (range 20.3 - 92.3), with a slight predominance of women (56.1%). Patients had received a mean of three prior therapies (range 0 - 10), and 43 patients (75.4%) had previously been exposed to at least one other thrombopoietin receptor agonist (TPO-RA).

At baseline, the median platelet count (evaluated in 54 patients) was 11000/mm3 (range 2000-50000). An additional 3 patients switched to Avatrombopag due to intolerance to other TPO-RAs, with no washout time, and had baseline platelet counts > 90000/mm3.

The median time to platelet counts greater than 50000/mm3 (assessed from 54 patients who started with low counts) was 27 days (range 3 - 456 ). At a median follow-up of 4.9 months (range 0.3 - 14.6), 40 patients (70.2%) were still on treatment, with a median platelet count of 96000/mm3 (range 13000 - 403000). Thrombotic events were observed in 4 patients (7%), while bleeding events occurred in 5 patients (8.8%), but only 3 of these led to treatment discontinuation. Additionally, 4 deaths were recorded: 1 due to a COVID infection, 1 from acute renal failure, 1 from a subdural hematoma, and 1 unrelated death.

CONCLUSIONS

To our knowledge, this is the first large real-world experience reported in Italy for Avatrombopag treatment in patients with immune thrombocytopenia (ITP). Our data confirmed the high efficacy of this therapy, even in heavily pretreated patients, including those with prior exposure to one or two TPO-RAs. The safety profile was acceptable, with few adverse events (including thrombotic and hemorrhagic events) leading to discontinuation. However, more data and longer follow-up are needed to evaluate the duration of response.
Dynamic Changes in Peripheral Blood Lymphocyte Subset Counts and Functions in Children with B-Cell Acute Lymphoblastic Leukemia during and after Chemotherapy
Authors: Yin Tiantian , Han Juan , Qu Ping , Jin Runming , Zhou Fen
Published: December 2024

Background: B-cell acute lymphoblastic leukemia (B-ALL) is the most common malignant tumor in children. This study aimed to investigate the lymphocyte subsets, their activities and dynamic changes during and after chemotherapy in children wiht B-ALL.

Methods: Peripheral blood samples were prospectively and cross-sectional collected from B-ALL children at different treatment time points (50 cases at initial diagnosis, 40 cases after induction, 20 cases at the beginning of maintenance, 15 cases at 61 weeks, the middle of maintenance, 20 cases at the end of treatment, 18 cases at 5-year follow-up) and healthy control children. T, B and NK lymphocyte subsets, immunophenotypes and functions were detected by flow cytometry.

Results: Compared with healthy controls, T cells in patients newly diagnosed with B-ALL were in a state of inhibition, senescence and exhaustion. The percentages of CD4+, CD4/CD8, γδ T, CD4+Tnaive as well as CD8+Tnaive were significantly decreased (P < 0.05). The percentages of CD8+, Tregs, CD4+Tcm, CD8+Tem, CD4+PD-1, CD4+TIM-3, CD8+PD-1 and CD8+TIM-3 were significantly increased (P < 0.05). The levels of IFN-γ, IL-4 and IL-2 were significantly lower. NK and NKT cells in B-ALL children were in a state of inhibition, which mainly showed that the percentage of NK and NKT was significantly decreased, and the activator receptor NKP46 and NKG2D were significantly decreased (P < 0.05). After induction therapy, the percentage of B cells decreased significantly (P=0.000), and T cells and NK cells had a partial recovery (P < 0.05). When entering to maintenance therapy, T, B, NK cell percentage and T cell function of patients were the most severely impaired, and then gradually recovered. Till to the end of treatment, the function of lymphocytewas inhibited. At 5 years follow-up, the counts and functions of B and NK cells recovered, but that of T cells did not recover completely.

Conclusions: The absolute counts and functions of T cells and NK cells, which were significantly lower in children with B-ALL, gradually recovered after chemotherapy. Detecting the counts and functions of peripheral blood lymphocytes is important for identifying new prognostic markers of ALL, guiding to prevent infection and administering effective immunotherapy.
Visualization of Bortezomib Induced Peripheral Neuropathy By Dorsal Root Ganglion MRI in Patients with Multiple Myeloma
Authors: Zhou Xiang , Schindehuette Magnus , Cebulla Nadine , Öztürk Baran , Weiner Simon , Mersi Julia , Rasche Leo , Waldschmidt Johannes , Einsele Hermann , Blum Robert , Rittner Heike , Sommer Claudia , Pham Mirko , Kortüm Martin K.
Published: December 2024

Background:

Whereas most treatments in multiple myeloma (MM) are stopped due to their inefficiency over time and the development of drug resistance, the use of the proteasome inhibitor bortezomib (BTZ) is mainly limited by its neurotoxic effects and the induction of BTZ induced neuropathy (BIPN). Of particular pathophysiological importance is the dorsal root ganglion (DRG), which plays a central role in the development of BIPN (Argyriou et al. Blood 2008). As early BIPN diagnosis and treatment adjustment is critical, we here investigated the predictive potential of DRG MRI and measured the DRG T2 signal intensity as a biomarker of inflammation in BTZ treated MM patients.

Methods: We recruited a total of 112 MM patients receiving BTZ either currently or in the past. Of these, 92 (82 %) had BIPN, which was painful in 37% (n=41) and mild and only detectable by sophisticated clinical-neurological diagnostics in 52% of patients (n = 58, severity Grade 1 with/without pain). DRG MRI was performed in a subset of 54 patients, including 45 of them with detectable BIPN (all grades), and additionally in 12 healthy controls. Using high-resolution three-dimensional T2-weighted fast spin-echo DRG MRI, we evaluated the lumbosacral DRG Th12-S2 bilaterally in all patients. Comparative analyses were performed using one-way ANOVA with Bonferroni correction. Pearson's Rho was used for the correlation analyses.

Results: Analysis revealed a significant increase in DRG T2 signal intensity in MM patients with BIPN compared to MM patients without BIPN (p= 0.005) and healthy controls (p= 0.003). Furthermore, an inverse correlation was found between DRG T2 signal intensity and intraepidermal nerve fiber density (IENFD) (p= 0.035). The mean DRG volume on L5+S1 level (in mm³) did not differ significantly between the groups, however, IENFD was significantly decreased in BIPN MM patients (n=26; 3.41 fibers/mm) compared to the healthy controls (n=12, 5.28 fibers/mm; p = 0.031), MM patients without BIPN showed no difference in IENFD (4.45 fibers/mm) compared to healthy controls.

Conclusion:

DRG T2 signal intensity is a surrogate marker of inflammation, and our observations suggest that inflammatory changes at the DRG level underlie BIPN in MM patients. Our imaging holds potential to be used as a non-invasive biomarker for early BIPN detection and to define patients at risk. This is an ongoing study, and imaging from the full cohort and from additional longitudinal assessment will be presented at the meeting.
Real World Outcomes of Ruxolitinib Treatment of Steroid Refractory/Dependent Acute Graft Versus Host Disease
Authors: Leung Becky , Butler Jason , Curley Cameron , Henden Andrea S , Hunt Stewart , Morton James , Subramoniapillai Elango , Stewart Caroline , Tey Siok-Keen , Kennedy Glen A , Scott Ashleigh P
Published: December 2024

Aim:

Steroid-refractory and/or dependent acute graft versus host disease (SR/SD-aGVHD) is a life-threatening complication following allogeneic haematopoietic stem cell transplantation (HSCT). The REACH-2 study established ruxolitinib as standard of care compared to historical salvage options, however patients with lower gastrointestinal (GI) involvement still have poor outcomes1. We aimed to review local outcomes of patients treated with ruxolitinib for SR/SD-aGVHD.

Method:

We performed a retrospective analysis of HSCT recipients treated between March 2021 and January 2024. The primary endpoint was overall survival (OS), with secondary endpoints including non-relapse mortality (NRM) and requirement for third line therapy.

Results:

We identified 37 patients with SR/SD-aGVHD, median age of 52 years at time of HSCT. All patients underwent T-replete HSCT using GCSF-stimulated PBSC. Donor source was matched sibling (27%), matched unrelated donor (49%), or haploidentical/mismatched (24%); with myeloablative conditioning in 35%. GVHD prophylaxis was cyclosporin/methotrexate (78%) or post-transplant cyclophosphamide/tacrolimus/mycophenolate (22%).

Acute GVHD occurred at median 41 days post-HSCT with grade III-IV disease in 48%, lower GI involvement in 68%, and skin-only disease in 27% at diagnosis. Following ruxolitinib commencement for SR/SD-aGVHD, the proportion of patients at D28/D56 with complete response was 62%/54% and overall response 78%/70%. Ruxolitinib failure occurred more frequently in patients with lower GI involvement compared to those without, with D28/D56 overall response of 69%/58% versus 100%/100%. Flares of aGVHD after initial response requiring third-line therapy occurred more frequent in patients with lower GI aGVHD (40% vs 0%).

At median follow-up of 159 days post ruxolitinib commencement, overall survival was 51% with mortality due to GVHD (56%), infection (22%) and relapse (6%). NRM occurred more frequently in patients with lower GI aGVHD (64% vs 8%).

Conclusion:

Our single-centre data supports ruxolitinib as second-line therapy for SR/SD-aGVHD. However, patients with lower GI involvement remain at significant risk for non-sustained response requiring third-line therapy and non-relapse mortality.

References: Zeiser R, von Bubnoff N, Butler J, Mohty M, Niederwieser D, Or R, Szer J, Wagner EM, Zuckerman T, Mahuzier B, Xu J, Wilke C, Gandhi KK, Socié G; REACH2 Trial Group. Ruxolitinib for Glucocorticoid-Refractory Acute Graft-versus-Host Disease. N Engl J Med. 2020 May 7;382(19):1800-1810. doi: 10.1056/NEJMoa1917635. Epub 2020 Apr 22. PMID: 32320566.
Multi-Cohort Meta-Analysis of Rnaseq Data to Determine Sex Bias in Pediatric Acute Myeloid Leukemia
Authors: Gardner Eve , Tokarski Ember , Jensen Ava , Harmon Lauren , Triche Timothy J
Published: December 2024

Leukemia is the most common childhood cancer, and remains a leading cause of childhood cancer mortality. The most common leukemia in children, B-cell acute lymphoblastic leukemia, has a 92% cure rate (5-year overall survival). In contrast, pediatric acute myeloid leukemia (pAML) has a 68% cure rate, which has not changed significantly despite 40 years of trials.

AML exhibits some of the most striking sex-specific disparities in childhood cancer, affecting disease risk, therapy response, and transplant outcomes. pAML cohorts are consistently about 45% female and 55% male, yielding a male excess risk of 22%. Some aggressive subtypes, such as NUP98::NSD1 fusions seen in many treatment failures, the male:female ratio rises to 2:1, and to 3:1 near puberty. Up to 50% of pAML patients show loss of X or Y, with loss of Y (LOY) prognostic for lifetime all-cause mortality in males. Even mutation-targeted therapies such as midostaurin show significant differences in efficacy by sex in Phase III trials.

We sought to illuminate these differences via meta-analysis of bulk RNA sequencing (RNAseq) data from 2,000 TARGET (Therapeutically Applicable Research to Generate Effective Treatments) initiative patients and matched controls to identify sex-specific transcriptional differences. Subsequently, we interrogated these findings by performing a meta-analysis and cross-projection of publicly available single-cell RNA sequencing (scRNAseq) data from a variety of pediatric hematopoietic malignancies from eight published GEO datasets and two Alex's Lemonade Stand Foundation (ALSF) datasets, comprising 225 myeloid or myeloid-like pediatric leukemia patients and 23 non-leukemic specimens. Using Non-negative Matrix Factorization (NMF), we projected all scRNAseq data into the same latent space as the Chan Zuckerberg Initiative (CZI) human, matched-tissue scRNAseq. These NMF projections were then utilized to inform a comprehensive meta-analysis of both bulk and scRNAseq data from myeloid-like diseases. By leveraging both bulk and single-cell transcriptomic data, we aimed to elucidate the role of sex in the molecular underpinnings of pediatric hematopoietic malignancies, potentially unveiling novel therapeutic targets and contributing to personalized medicine approaches in these diseases.

Combined with the development of new models for predisposition conditions such as RUNX1-FPDMM, our work begins to reveal the molecular underpinnings of sex bias in treatment response and immunogenicity for pAML patients, and creates a foundation to better understand how genetic lesions interact with age and sex to shape risk in this lethal malignancy.
Clinical Characteristics, Molecular Features and Transplantation Outcomes in MDS with Low Blasts and without Defining Genetic Abnormalities: An Analysis of MDS-h and MDS-LB Patients
Authors: Guo Wenjing , Jia Li , Mi Yingchang , Feng Sizhou
Published: December 2024

Objective: According to the World Health Organization (WHO) 2022 classification, myelodysplastic neoplasms/syndromes (MDS) patients without increased blasts and defining genetic abnormalities are categorized as either hypoplastic MDS (MDS-h) or MDS with low blasts (MDS-LB). This study aims to evaluate the clinical characteristics, genetic and cytogenetic profiles, and outcomes of MDS-h and MDS-LB patients undergoing allogeneic hematopoietic stem cell transplantation (allo-HSCT).

Methods: We retrospectively analyzed data from 25 MDS-h and 64 MDS-LB patients who underwent allo-HSCT between January 2010 and January 2023. A one-to-one propensity score-matched analysis was performed to minimize confounding factors. Additionally, genetic mutations and chromosomal abnormalities were compared among 25 MDS-h, 64 MDS-LB and 205 severe aplastic anemia (SAA) patients.

Results: Most patients were classified into higher-risk group by Molecular International Prognostic Scoring System (IPSS-M) (MDS-h: 80.00% vs. MDS-LB: 71.88%, P=0.605). Pre-transplantation transfusion dependency was observed in 80.0% of MDS-h and 79.7% of MDS-LB patients (P=0.541). MDS-h patients had lower platelet counts (35×109/L vs. 59×109/L, P=0.034) and a higher incidence of severe cytopenia (56.00% vs. 34.38%, P=0.104). Both MDS-h and MDS-LB patients exhibited significantly more mutations and chromosomal abnormalities compared to SAA patients (P<0.030). BCOR mutations were prevalent in MDS-h (23.81%), while U2AF1 (39.62%), ASXL1 (30.19%) and RUNX1 (20.75%) mutations were common in MDS-LB. The estimated 3-year overall survival (OS) was 76.0% (61.0%-94.7%) for MDS-h and 83.9% (75.2%-93.6%) for MDS-LB (P=0.328). Failure-free survival, transplant-related mortality and cumulative incidences of engraftment, graft-versus-host disease and infection were also similar between groups. Risk factors for worse OS included bloodstream infection within 100 days post-HSCT and FAT1 mutations in MDS-h, and RUNX1 mutations in MDS-LB. Donor type and the interval from diagnosis to HSCT had no significant impact on post-HSCT outcomes.

Conclusions: MDS-h and MDS-LB patients undergoing allo-HSCT exhibited slightly distinct clinical and genetic features, but both groups achieved favorable outcomes after transplantation. This study highlights the potential efficacy of allo-HSCT in mitigating adverse genetic mutations in MDS-h and MDS-LB patients, with no significant impact from donor type or diagnosis-to-HSCT interval on outcomes.
Disruption of the Retinoblastoma Lxcxe Region Induces Cell Death in AML By Activation of the Unfolded Protein Response Leading to NOXA Induction
Authors: Bhuniya Avishek , Lu Lily , Ploi Adi Narayana Reddy , Tcyganov Evgenii , Mendoza Maria De Grecia Cauti , Sharma Paridhima , Ortiz Adiana Ochoa , Zhigarev Dmitry , Salvino Joseph , Montaner Luis
Published: December 2024

The Retinoblastoma (Rb1) protein is expressed in high amounts in leukemia and myeloid cells. While RB1 controls cell cycle progression, it also regulates cell death through protein binding regions such as the LxCxE binding cleft. To test whether blocking the Rb1 LxCxE domain would induce specific cell death in acute myeloid leukemia (AML) cells, we developed a small molecule (AP-3-84) that can selectively bind the LxCxE domain of Rb1 blocking protein-protein interactions. Statistical analysis was performed with GraphPad Prsim.

The activity of AP-3-84 was evaluated in vitro using multiple AML cell lines (THP-1, AML-193, Kasumi-1, MOLM-14) and in vivo using luciferase-transduced THP-1 cells implanted in NGS mice. AP-3-84 induced cell death in a concentration-dependent manner in all AML cell lines tested. In vivo treatment with AP-3-84 also reduced tumor growth as measured by luciferase signal using an IVIS platform. Importantly, AP-3-84 treatment did not induce cell death in PMA-differentiated THP1 cells or non-hematopoietic cancer cell lines. PROTAC-mediated Rb1 degradation reduced AP-3-84-induced cell death in THP-1 cells, confirming Rb1 as the target for AP-3-84.

Bulk RNA sequencing showed that 30 min treatment of THP-1 cells with AP-3-84 resulted in upregulation of the NRF2-mediated oxidative stress response and unfolded protein response (UPR) and induction of ATF4 and DDIT3 (CHOP). Further evaluation of the UPR pathway via western blotting indicated increased expression of pPERK, peIF2a, ATF4, ATF3, and NOXA in a sequential time-dependent manner. Notably, the levels of BAX and BAK remained unchanged at early time points (up to 6 hours), suggesting that the UPR pathway is the predominant pathway triggered by AP-3-84. In the same experiments we observed induction of lipid peroxidation. As lipid peroxidation induce ferroptosis, treatment with ferrostatin 1 reduced the effect of AP-3-84, supporting that induction of ferroptosis is the predominant cell death pathway induced by AP-3-84. We also observed an increase in cardiolipin levels consistent with lipid peroxidation changes after AP-3-84 treatment, supporting the contribution of mitochondria to activation of cell death pathways.

We conclude that AP-3-84 triggers cell death in AML by activating the UPR via PERK-eIF2a-ATF4-NOXA pathway involving mitochondrial regulation. Our results indicate that Rb1 LxCxE domain may represent a novel therapy target in AML.