bims-tremyl 2022-12-18 papers

bims-tremyl

Biomed News

on Therapy resistance biology in myeloid leukemia

Issue of 2022–12–18
37 papers selected by
Paolo Gallipoli, Barts Cancer Institute, Queen Mary University of London

Erythroid/megakaryocytic differentiation confers BCL-XL dependency and venetoclax resistance in acute myeloid leukemia.
Overlapping features of therapy-related and de novoNPM1-mutated AML.
Applying molecular measurable residual disease testing in acute myeloid leukaemia.
A key role of the WEE1-CDK1 axis in mediating TKI-therapy resistance in FLT3-ITD positive acute myeloid leukemia patients.
Impact of FLT3-ITD Mutation Status and Its Ratio in a Cohort of 2901 Patients Undergoing Upfront Intensive Chemotherapy: A PETHEMA Registry Study.
Targeting Menin and CD47 to Address Unmet Needs in Acute Myeloid Leukemia.
Outcomes in Patients With FLT3-Mutated R/R AML Who Underwent Transplantation in the Phase 3 ADMIRAL Trial of Gilteritinib Versus Salvage Chemotherapy.
IL3-zetakine combined with a CD33 costimulatory receptor as a Dual CAR approach for safer and selective targeting of AML.
Heterogeneous nuclear ribonucleoprotein K is overexpressed in acute myeloid leukemia and causes myeloproliferation in mice via altered Runx1 splicing.
The transcriptomic landscape of elderly acute myeloid leukemia identifies B7H3 and BANP as a favorable signature in high-risk patients.
Dual inhibition of CHK1/FLT3 enhances cytotoxicity and overcomes adaptive and acquired resistance in FLT3-ITD acute myeloid leukemia.
CCRL2 affects the sensitivity of myelodysplastic syndrome and secondary acute myeloid leukemia cells to azacitidine.
A randomized phase II/III study of 'novel therapeutics' versus azacitidine in newly diagnosed patients with acute myeloid leukemia (AML), high-risk myelodysplastic syndrome (MDS), or chronic myelomonocytic leukemia (CMML), age 60 or older: a report of the comparison of azacitidine and nivolumab to azacitidine: SWOG S1612.
An unexpected partnership targeting FLT3wt AML.
Development of Gilteritinib-Based Chimeric Small Molecules that Potently Induce Degradation of FLT3-ITD Protein.
Ex vivo venetoclax sensitivity testing predicts treatment response in acute myeloid leukemia.
NUC-7738 regulates β-catenin signalling resulting in reduced proliferation and self-renewal of AML cells.
A pro B cell population forms the apex of the leukemic hierarchy in Hoxa9/Meis1-dependent AML.
Splicing factor deficits render hematopoietic stem and progenitor cells sensitive to STAT3 inhibition.
Infectious complications after intensive chemotherapy with CLAG-M versus 7+3 for AML and other high-grade myeloid neoplasms.
Viewing AML through a New Lens: Technological Advances in the Study of Epigenetic Regulation.
A genetic disorder reveals a hematopoietic stem cell regulatory network co-opted in leukemia.
Asxl1 deletion disrupts MYC and RNA polymerase II function in granulocyte progenitors.
Deciphering the immuno-pathological role of FLT, and evaluation of a novel dual inhibitor of topoisomerases and mutant-FLT3 for treating leukemia.
Heterogeneity of Patient-Derived Acute Myeloid Leukemia Cells Subjected to SYK In Vitro Inhibition.
Indirect treatment comparison of oral versus injectable azacitidine as maintenance therapy for acute myeloid leukemia.
Myeloid cells promote interferon signaling-associated deterioration of the hematopoietic system.
Adaptation of Serious Illness Care Program to be Delivered via Telehealth for Older Patients with Hematologic Malignancy.
Establishment of isogenic induced pluripotent stem cells with or without pathogenic mutation for understanding the pathogenesis of myeloproliferative neoplasms.
Asciminib monotherapy for newly diagnosed chronic myeloid leukemia in chronic phase: the ASC4FIRST phase III trial.
Integrated single-cell transcriptome analysis of CD34 + enriched leukemic stem cells revealed intra- and inter-patient transcriptional heterogeneity in pediatric acute myeloid leukemia.
In vivo characterization of glutamine metabolism identifies therapeutic targets in clear cell renal cell carcinoma.
DNA damage to bone marrow stromal cells by anti-leukemia drugs induces chemo-resistance in acute myeloid leukemia via paracrine FGF10-FGFR2 signaling.
Human genetic diversity alters off-target outcomes of therapeutic gene editing.
Venetoclax treatment in cancer patients has limited impact on circulating T and NK cells.
Raman microspectroscopy reveals unsaturation heterogeneity at the lipid droplet level and validates an in vitro model of bone marrow adipocyte subtypes.
A phase 2, prospective, randomized, open-label study of defibrotide added to standard-of-care prophylaxis for the prevention of acute graft-versus-host disease after allogeneic hematopoietic cell transplantation.

Blood. 2022 Dec 12. pii: blood.2021011094. [Epub ahead of print]

Erythroid/megakaryocytic differentiation confers BCL-XL dependency and venetoclax resistance in acute myeloid leukemia.

Heikki Kuusanmäki, Olli Dufva, Markus Vähä-Koskela, Aino-Maija Leppä, Jani Huuhtanen, Ida Maria Vänttinen, Petra Johanna Nygren, Jay Klievink, Jonas Otto Vilhelm Bouhlal, Petri Pölönen, Qi Zhang, Shady Adnan Awad, Cristina Mancebo-Pérez, Joseph Saad, Juho J Miettinen, Komal Kumar Javarappa, Sofia Aakko, Tanja Ruokoranta, Samuli Eldfors, Merja Heinäniemi, Kim Theilgaard-Mönch, Ulla Wartiovaara-Kautto, Mikko A I Keränen, Kimmo Porkka, Marina Konopleva, Krister Wennerberg, Mika Kontro, Caroline A Heckman, Satu Mustjoki.

Myeloid neoplasms with erythroid or megakaryocytic differentiation include pure erythroid leukemia (PEL), myelodysplastic syndrome (MDS) with erythroid features, and acute megakaryoblastic leukemia (FAB M7) and are characterized by poor prognosis and limited treatment options. Here, we investigate the drug sensitivity landscape of these rare malignancies. We show that acute myeloid leukemia (AML) cells with erythroid or megakaryocytic differentiation depend on the anti-apoptotic protein BCL-XL, rather than BCL-2, using combined ex vivo drug sensitivity testing, genetic perturbation, and transcriptomic profiling. High-throughput screening of > 500 compounds identified the BCL-XL-selective inhibitor A-1331852 and navitoclax as highly effective against erythroid/megakaryoblastic leukemia cell lines. In contrast, these AML subtypes were resistant to the BCL-2 inhibitor venetoclax used clinically in the treatment of AML. Consistently, genome-scale CRISPR-Cas9 and RNAi screening data demonstrated striking essentiality of BCL2L1 encoding BCL-XL, but not BCL2 or MCL1, for the survival of erythroid/megakaryoblastic leukemia cell lines. Single-cell and bulk transcriptomics of patient samples with erythroid and megakaryoblastic leukemias identified high BCL2L1 expression compared to other subtypes of AML and other hematological malignancies, where BCL2 and MCL1 were more prominent. BCL-XL inhibition effectively killed blasts in AML patient samples with erythroid or megakaryocytic differentiation ex vivo and reduced tumor burden in a mouse erythroleukemia xenograft model. Combining BCL-XL inhibitor with the JAK inhibitor ruxolitinib showed synergistic and durable responses in cell lines. Our results suggest targeting BCL-XL as a potential therapy option in erythroid/megakaryoblastic leukemias and highlight an AML subgroup with potentially reduced sensitivity to venetoclax-based treatments.

DOI: https://doi.org/10.1182/blood.2021011094
Blood. 2022 Dec 12. pii: blood.2022018108. [Epub ahead of print]

Overlapping features of therapy-related and de novoNPM1-mutated AML.

Jad Othman, Manja Meggendorfer, Enrico Tiacci, Christian Thiede, Richard F Schlenk, Richard Dillon, Sebastian Stasik, Alessandra Venanzi, Sarah Bertoli, Eric Delabesse, Pierre-Yves Dumas, Arnaud Pigneux, Audrey Bidet, Amanda Frances Gilkes, Ian Thomas, Maria Teresa Voso, Alessandro Rambaldi, Lorenzo Brunetti, Vincenzo Maria Perriello, Vibeke Andresen, Bjorn T Gjertsen, Maria Paola Martelli, Christian Récher, Hubert Serve, Carsten Müller-Tidow, Claudia D Baldus, Torsten Haferlach, Nigel H Russell, Brunangelo Falini.

NPM1-mutated AML represents a WHO leukemia entity with unique pathological and clinical features. Little is known about the characteristics of "therapy-related" NPM1-mutated AML. We compared the genetics, transcriptional profile and clinical outcome of therapy-related NPM1-mutated AML (t-NPM1 AML), de-novo NPM1-mutated AML (dn-NPM1 AML) and therapy-related AML with wild-type NPM1 (t-AML). A normal karyotype was more frequent in t-NPM1 AML (n=78/96 cases, 88%) and dn-NPM1 (n=1986/2394,88%) than in t-AML (n=103/390,28%; p <0.001). DNMT3A and TET2 were mutated in 43% and 40% of t-NPM1 AML (n=107), similar to dn-NPM1 (n=88, 48% and 30%; p-values >0.1), but more frequently than t-AML (n=162; 14% and 10%; p-values <0.001). TP53 and PPM1D, typically mutated in t-AML, were consistently wild-type in t-NPM1 AML (97% and 96%). t-NPM1 and dn-NPM1 AML were transcriptionally similar, displaying upregulation of HOX genes and down-regulation of CD133 and CD34. With a median follow-up of 62 months, 3-year overall survival (OS) for t-NPM1 AML (n=96), dn-NPM1 AML (n=2394) and t-AML (n=390) was 54%, 60% and 31%. In multivariable analysis OS was similar for the two NPM1-mutated groups (HR 0.9, 95%CI 0.65-1.25, p=0.45) but better in t-NPM1 AML than t-AML (HR 1.86, 95%CI 1.30-2.68, p<0.001). Relapse-free survival did not differ between t-NPM1 and dn-NPM1 AML (HR 1.02, 95%CI 0.72-1.467, p=0.90) but was significantly higher in t-NPM1 AML than t-AML (HR 1.77, 95%CI 1.19-2.64, p=0.0045).t-NPM1 and dn-NPM1 AML have similar clinical, genomic and transcriptomic features, suggesting that they should be classified as a single disease entity.

DOI: https://doi.org/10.1182/blood.2022018108
Pathology. 2022 Nov 23. pii: S0031-3025(22)00312-9. [Epub ahead of print]

Applying molecular measurable residual disease testing in acute myeloid leukaemia.

Michael Krigstein, Harry J Iland, Andrew H Wei.

  Molecular testing in acute myeloid leukaemia (AML) has continued to dramatically advance in recent years, facilitating the ability to detect residual disease at exponentially lower levels. With the advent of the recently updated ELN consensus recommendations, there is increasing complexity to ordering and interpreting measurable residual disease (MRD) assays in AML. We outline the technology itself in conjunction with the relevant testing timepoints, clinically significant thresholds and potential prognostic and therapeutic significance of MRD testing for the major molecular targets in AML. This practical overview should assist haematologists in incorporating molecular MRD assays routinely into their personalised AML clinical management.

Keywords:  FLT3; Measurable residual disease; NPM1; acute myeloid leukaemia; clonal haematopoiesis

DOI:  https://doi.org/10.1016/j.pathol.2022.11.003
Leukemia. 2022 Dec 12.

A key role of the WEE1-CDK1 axis in mediating TKI-therapy resistance in FLT3-ITD positive acute myeloid leukemia patients.

Giorgia Massacci, Veronica Venafra, Sara Latini, Valeria Bica, Giusj Monia Pugliese, Simone Graziosi, Felix Klingelhuber, Natalie Krahmer, Thomas Fischer, Dimitrios Mougiakakos, Martin Boettcher, Livia Perfetto, Francesca Sacco.

The insertion site of the internal tandem duplications (ITDs) in the FLT3 gene affects the sensitivity to tyrosine kinase inhibitors (TKIs) therapy in acute myeloid leukemia (AML). Patients with the ITD in the tyrosine kinase domain lack effective therapeutic options. Here, to identify genotype-driven strategies increasing the TKI therapy efficacy, we developed SignalingProfiler, a strategy supporting the integration of high-sensitive mass spectrometry-based (phospho)proteomics, RNA sequencing datasets with literature-derived signaling networks. The approach generated FLT3-ITD genotype-specific predictive models and revealed a conserved role of the WEE1-CDK1 axis in TKIs resistance. Remarkably, pharmacological inhibition of the WEE1 kinase synergizes and strengthens the pro-apoptotic effect of TKIs therapy in cell lines and patient-derived primary blasts. Finally, we propose a new molecular mechanism of TKIs resistance in AML and suggest the combination of WEE1 inhibitor and TKI as a therapeutic option to improve patients clinical outcome.

DOI: https://doi.org/10.1038/s41375-022-01785-w
Cancers (Basel). 2022 Nov 24. pii: 5799. [Epub ahead of print]14(23):

Impact of FLT3-ITD Mutation Status and Its Ratio in a Cohort of 2901 Patients Undergoing Upfront Intensive Chemotherapy: A PETHEMA Registry Study.

Rosa Ayala, Gonzalo Carreño-Tarragona, Eva Barragán, Blanca Boluda, María J Larráyoz, María Carmen Chillón, Estrella Carrillo-Cruz, Cristina Bilbao, Joaquín Sánchez-García, Teresa Bernal, David Martinez-Cuadron, Cristina Gil, Josefina Serrano, Carlos Rodriguez-Medina, Juan Bergua, José A Pérez-Simón, María Calbacho, Juan M Alonso-Domínguez, Jorge Labrador, Mar Tormo, Maria Luz Amigo, Pilar Herrera-Puente, Inmaculada Rapado, Claudia Sargas, Iria Vazquez, María J Calasanz, Teresa Gomez-Casares, Ramón García-Sanz, Miguel A Sanz, Joaquín Martínez-López, Pau Montesinos.

  FLT3-ITD results in a poor prognosis in terms of overall survival (OS) and relapse-free survival (RFS) in acute myeloid leukemia (AML). However, the prognostic usefulness of the allelic ratio (AR) to select post-remission therapy remains controversial. Our study focuses on the prognostic impact of FLT3-ITD and its ratio in a series of 2901 adult patients treated intensively in the pre-FLT3 inhibitor era and reported in the PETHEMA registry. A total of 579 of these patients (20%) harbored FLT3-ITD mutations. In multivariate analyses, patients with an FLT3-ITD allele ratio (AR) of >0.5 showed a lower complete remission (CR rate) and OS (HR 1.47, p = 0.009), while AR > 0.8 was associated with poorer RFS (HR 2.1; p < 0.001). Among NPM1/FLT3-ITD-mutated patients, median OS gradually decreased according to FLT3-ITD status and ratio (34.3 months FLT3-ITD-negative, 25.3 months up to 0.25, 14.5 months up to 0.5, and 10 months ≥ 0.5, p < 0.001). Post-remission allogeneic transplant (allo-HSCT) resulted in better OS and RFS as compared to auto-HSCT in NPM1/FLT3-ITD-mutated AML regardless of pre-established AR cutoff (≤0.5 vs. >0.5). Using the maximally selected log-rank statistics, we established an optimal cutoff of FLT3-ITD AR of 0.44 for OS, and 0.8 for RFS. We analyzed the OS and RFS according to FLT3-ITD status in all patients, and we found that the group of FLT3-ITD-positive patients with AR < 0.44 had similar 5-year OS after allo-HSCT or auto-HSCT (52% and 41%, respectively, p = 0.86), but worse RFS after auto-HSCT (p = 0.01). Among patients with FLT3-ITD AR > 0.44, allo-HSCT was superior to auto-HSCT in terms of OS and RFS. This study provides more evidence for a better characterization of patients with AML harboring FLT3-ITD mutations.

Keywords:  FLT3–ITD mutation and ratio; acute myeloid leukemia (AML); death; outcome; prognosis; real-world outcomes; relapse; survival

DOI:  https://doi.org/10.3390/cancers14235799
Cancers (Basel). 2022 Nov 29. pii: 5906. [Epub ahead of print]14(23):

Targeting Menin and CD47 to Address Unmet Needs in Acute Myeloid Leukemia.

Andrew H Matthews, Keith W Pratz, Martin P Carroll.

  After forty years of essentially unchanged treatment in acute myeloid leukemia (AML), innovation over the past five years has been rapid, with nine drug approvals from 2016 to 2021. Increased understanding of the molecular changes and genetic ontology of disease have led to targeting mutations in isocitrate dehydrogenase, FMS-like tyrosine kinase 3 (FLT3), B-cell lymphoma 2 and hedgehog pathways. Yet outcomes remain variable; especially in defined molecular and genetic subgroups such as NPM1 (Nucleophosmin 1) mutations, 11q23/KMT2A rearranged and TP53 mutations. Emerging therapies seek to address these unmet needs, and all three of these subgroups have promising new therapeutic approaches. Here, we will discuss the normal biological roles of menin in acute leukemia, notably in KMT2A translocations and NPM1 mutation, as well as current drug development. We will also explore how CD47 inhibition may move immunotherapy into front-line settings and unlock new treatment strategies in TP53 mutated disease. We will then consider how these new therapeutic advances may change the management of AML overall.

Keywords:  CD47; KMT2A; acute myeloid leukemia (AML); menin; target therapy

DOI:  https://doi.org/10.3390/cancers14235906
Transplant Cell Ther. 2022 Dec 13. pii: S2666-6367(22)01832-2. [Epub ahead of print]

Outcomes in Patients With FLT3-Mutated R/R AML Who Underwent Transplantation in the Phase 3 ADMIRAL Trial of Gilteritinib Versus Salvage Chemotherapy.

Alexander E Perl, Richard A Larson, Nikolai A Podoltsev, Stephen Strickland, Eunice S Wang, Ehab Atallah, Gary J Schiller, Giovanni Martinelli, Andreas Neubauer, Jorge Sierra, Pau Montesinos, Christian Recher, Sung-Soo Yoon, Yoshinobu Maeda, Naoko Hosono, Masahiro Onozawa, Takayasu Kato, Hee-Je Kim, Nahla Hasabou, Rishita Nuthethi, Ramon Tiu, Mark J Levis.

   BACKGROUND: The fms-like tyrosine kinase 3 (FLT3) inhibitor gilteritinib improved the survival of patients with relapsed or refractory (R/R) FLT3-mutated acute myeloid leukemia (AML) in the phase 3 ADMIRAL trial.
OBJECTIVES: To assess survival and relapse rates among patients in the ADMIRAL trial who underwent hematopoietic stem cell transplantation (HSCT), as well as safety outcomes in patients who received posttransplant gilteritinib maintenance therapy.
STUDY DESIGN: ADMIRAL was a global, phase 3, randomized controlled trial, which enrolled adult patients with FLT3-mutated R/R AML. Patients with R/R AML who harbored FLT3 internal tandem duplication mutations in the juxtamembrane domain or D835/I836 point mutations in the tyrosine kinase domain were randomized (2:1) to gilteritinib (120 mg/day) or preselected high- or low-intensity salvage chemotherapy (1-2 cycles) as previously described (Perl AE, et al. N Engl J Med. 2019). Gilteritinib-arm patients who proceeded to HSCT could receive posttransplant gilteritinib maintenance therapy if they were within 30 to 90 days posttransplant and had achieved composite complete remission (CRc) with successful engraftment and no posttransplant complications. Adverse events during HSCT were collected in the gilteritinib arm only. Survival outcomes and the cumulative incidence of relapse were assessed in patients who underwent HSCT during the trial. Treatment-emergent adverse events were evaluated in patients who restarted gilteritinib as posttransplant maintenance therapy.
RESULTS: Gilteritinib-arm patients underwent HSCT more frequently (n=64; 26%) than chemotherapy-arm patients (n=19; 15%). For all transplanted patients, 12- and 24-month overall survival (OS) rates were 68% and 47%, respectively. Despite a trend toward longer OS after pretransplant CRc, posttransplant survival was comparable across arms. Patients who resumed gilteritinib after HSCT had low relapse rates after pretransplant CRc (20%) or CR (0%). The most common adverse events observed with posttransplant gilteritinib therapy were increased alanine aminotransferase (40%), pyrexia (43%), and diarrhea (40%); grade ≥3 adverse events were primarily related to myelosuppression. Incidences of grade ≥3 graft-versus-host disease and related mortality were low.
CONCLUSION: Posttransplant survival was similar across the two study arms in ADMIRAL, but higher remission rates with gilteritinib facilitated receipt of HSCT. Gilteritinib as posttransplant maintenance therapy had a stable safety and tolerability profile and was associated with low relapse rates. Taken together, these data support a preference for bridging therapy with gilteritinib over chemotherapy in transplant-eligible patients.

Keywords:  Acute myeloid leukemia; FLT3 inhibitor; FLT3 mutation; hematopoietic stem cell transplantation; posttransplant maintenance therapy

DOI:  https://doi.org/10.1016/j.jtct.2022.12.006
Blood Adv. 2022 Dec 15. pii: bloodadvances.2022008762. [Epub ahead of print]

IL3-zetakine combined with a CD33 costimulatory receptor as a Dual CAR approach for safer and selective targeting of AML.

Vincenzo Maria Perriello, Maria Caterina Rotiroti, Ilaria Pisani, Stefania Galimberti, Gaia Alberti, Giulia Pianigiani, Valerio Ciaurro, Andrea Marra, Marcella Sabino, Valentina Tini, Giulio Spinozzi, Federica Mezzasoma, Francesco Morena, Sabata Martino, Domenico Salerno, Julian François Ashby, Brittany Wingham, Marta Serafini, Maria Paola Martelli, Brunangelo Falini, Andrea Biondi, Sarah Tettamanti.

Acute Myeloid Leukemia (AML) still represents an unmet clinical need for adult and pediatric patients. Adoptive cell therapy by chimeric antigen receptor (CAR)-engineered T cells demonstrated a high therapeutic potential, but further development is required to ensure a safe and durable disease remission in AML, especially in elderly patients. To date, translation of CAR T cell therapy in AML is limited by the absence of an ideal tumor-specific antigen. CD123 and CD33 are the two most widely overexpressed LSCs biomarkers but their shared expression with endothelial and hematopoietic stem and progenitor cells (HSPCs) increases the risk of undesired vascular and hematologic toxicities. To counteract this issue, we established a balanced Dual CAR strategy aimed at reducing off-target toxicities while retaining full functionality against AML. Cytokine-Induced Killer (CIK) cells, co-expressing a first-generation low affinity anti-CD123 IL3-zetakine and an anti-CD33 as costimulatory receptor (CCR) without activation signaling domains, demonstrated a powerful antitumor efficacy against AML targets without any relevant toxicity on HSPCs and endothelial cells. The proposed optimized Dual CAR CIK strategy could offer the opportunity to unleash the potential of specifically target CD123+/CD33+ leukemic cells while minimizing toxicity against healthy cells.

DOI: https://doi.org/10.1182/bloodadvances.2022008762
NAR Cancer. 2022 Dec;4(4): zcac039

Heterogeneous nuclear ribonucleoprotein K is overexpressed in acute myeloid leukemia and causes myeloproliferation in mice via altered Runx1 splicing.

Marisa J L Aitken, Prerna Malaney, Xiaorui Zhang, Shelley M Herbrich, Lauren Chan, Oscar Benitez, Ashley G Rodriguez, Huaxian Ma, Rodrigo Jacamo, Ruizhi Duan, Todd M Link, Steven M Kornblau, Rashmi Kanagal-Shamanna, Carlos E Bueso-Ramos, Sean M Post.

Acute myeloid leukemia (AML) is driven by numerous molecular events that contribute to disease progression. Herein, we identify hnRNP K overexpression as a recurrent abnormality in AML that negatively correlates with patient survival. Overexpression of hnRNP K in murine fetal liver cells results in altered self-renewal and differentiation potential. Further, murine transplantation models reveal that hnRNP K overexpression results in myeloproliferation in vivo. Mechanistic studies expose a direct functional relationship between hnRNP K and RUNX1-a master transcriptional regulator of hematopoiesis often dysregulated in leukemia. Molecular analyses show that overexpression of hnRNP K results in an enrichment of an alternatively spliced isoform of RUNX1 lacking exon 4. Our work establishes hnRNP K's oncogenic potential in influencing myelogenesis through its regulation of RUNX1 splicing and subsequent transcriptional activity.

DOI: https://doi.org/10.1093/narcan/zcac039
Front Oncol. 2022 ;12 1054458

The transcriptomic landscape of elderly acute myeloid leukemia identifies B7H3 and BANP as a favorable signature in high-risk patients.

Sara Villar, Beñat Ariceta, Xabier Agirre, Aura Daniela Urribarri, Rosa Ayala, David Martínez-Cuadrón, Juan Miguel Bergua, Susana Vives, Lorenzo Algarra, Mar Tormo, Pilar Martínez, Josefina Serrano, Catia Simoes, Pilar Herrera, Maria José Calasanz, Ana Alfonso-Piérola, Bruno Paiva, Joaquín Martínez-López, Jesús F San Miguel, Felipe Prósper, Pau Montesinos.

  Acute myeloid leukemia (AML) in the elderly remains a clinical challenge, with a five-year overall survival rate below 10%. The current ELN 2017 genetic risk classification considers cytogenetic and mutational characteristics to stratify fit AML patients into different prognostic groups. However, this classification is not validated for elderly patients treated with a non-intensive approach, and its performance may be suboptimal in this context. Indeed, the transcriptomic landscape of AML in the elderly has been less explored and it might help stratify this group of patients. In the current study, we analyzed the transcriptome of 224 AML patients > 65 years-old at diagnosis treated in the Spanish PETHEMA-FLUGAZA clinical trial in order to identify new prognostic biomarkers in this population. We identified a specific transcriptomic signature for high-risk patients with mutated TP53 or complex karyotype, revealing that low expression of B7H3 gene with high expression of BANP gene identifies a subset of high-risk AML patients surviving more than 12 months. This result was further validated in the BEAT AML cohort. This unique signature highlights the potential of transcriptomics to identify prognostic biomarkers in in elderly AML.

Keywords:  acute myeloid leukemia; biomarkers; elderly; prognosis; transcriptomics

DOI:  https://doi.org/10.3389/fonc.2022.1054458
Leukemia. 2022 Dec 16.

Dual inhibition of CHK1/FLT3 enhances cytotoxicity and overcomes adaptive and acquired resistance in FLT3-ITD acute myeloid leukemia.

Kailong Jiang, Xuemei Li, Chang Wang, Xiaobei Hu, Peipei Wang, Lexian Tong, Yutong Tu, Beijing Chen, Tingting Jin, Tao Wang, Hanlin Wang, Yubing Han, Renzhao Gui, Jianmin Yang, Tao Liu, Jia Li, Yubo Zhou.

FLT3 inhibitors (FLT3i) are widely used for the treatment of acute myeloid leukemia (AML), but adaptive and acquired resistance remains a primary challenge. Inhibitors simultaneously blocking adaptive and acquired resistance are highly demanded. Here, we observed the potential of CHK1 inhibitors to synergistically improve the therapeutic effect of FLT3i in FLT3-mutated AML cells. Notably, the combination overcame adaptive resistance. The simultaneous targeting of FLT3 and CHK1 kinases may overcome acquired and adaptive resistance. A dual FLT3/CHK1 inhibitor 30 with a good oral PK profile was identified. Mechanistic studies indicated that 30 inhibited FLT3 and CHK1, downregulated the c-Myc pathway and further activated the p53 pathway. Functional studies showed that 30 was more selective against cells with various FLT3 mutants, overcame adaptive resistance in vitro, and effectively inhibited resistant FLT3-ITD AML in vivo. Moreover, 30 showed favorable druggability without significant blood toxicity or myelosuppression and exhibited a good oral PK profile with a T1/2 over 12 h in beagles. These findings support the targeting of FLT3 and CHK1 as a novel strategy for overcoming adaptive and acquired resistance to FLT3i therapy in AML and suggest 30 as a potential clinical candidate.

DOI: https://doi.org/10.1038/s41375-022-01795-8
Haematologica. 2022 Dec 15.

CCRL2 affects the sensitivity of myelodysplastic syndrome and secondary acute myeloid leukemia cells to azacitidine.

Theodoros Karantanos, Patric Teodorescu, Marios Arvanitis, Brandy Perkins, Tania Jain, Amy E DeZern, W Brian Dalton, Ilias Christodoulou, Bogdan C Paun, Ravi Varadhan, Christopher Esteb, Trivikram Rajkhowa, Challice Bonifant, Lukasz P Gondek, Mark J Levis, Srinivasan Yegnasubramanian, Gabriel Ghiaur, Richard J Jones.

Better understanding of the biology of resistance to DNA methyltransferase (DNMT) inhibitors (DNMTi) is required to identify therapies that can improve their efficacy for patients with highrisk myelodysplastic syndrome (MDS). CCRL2 is an atypical chemokine receptor that is upregulated in CD34+ cells from MDS patients and induces MDS and secondary AML (sAML) cell proliferation. In this study, we evaluated any role CCRL2 may have in the regulation of pathways associated with poor response or resistance to DNMTi. We found that CCRL2 KD in TF-1 cells downregulates DNA methylation and PRC2 activity pathways and increases DNA methyltransferases (DNMT) suppression by azacitidine in MDS/sAML vell lines (MDS92, MDS-L and TF-1). Consistently, CCRL2 deletion increased the sensitivity of these cells to azacitidine in vitro and the efficacy of azacitidine in an MDS-L xenograft model. Consistently, CCRL2 overexpression in MDS-L and TF-1 cells decreased their sensitivity to azacitidine. Finally, CCRL2 levels were higher in CD34+ cells from MDS and MDS/myeloproliferative neoplasm patients with poor response to DNMTi. In conclusion, we demonstrate that CCRL2 modulates epigenetic regulatory pathways, particularly DNMT levels, and affects MDS/sAML azacitidine sensitivity. These results support CCRL2 targeting as having MDS/sAML therapeutic potential.

DOI: https://doi.org/10.3324/haematol.2022.281444
Leuk Lymphoma. 2022 Dec 14. 1-5

A randomized phase II/III study of 'novel therapeutics' versus azacitidine in newly diagnosed patients with acute myeloid leukemia (AML), high-risk myelodysplastic syndrome (MDS), or chronic myelomonocytic leukemia (CMML), age 60 or older: a report of the comparison of azacitidine and nivolumab to azacitidine: SWOG S1612.

Sarit Assouline, Laura C Michaelis, Megan Othus, Annette E Hay, Roland B Walter, Meagan A Jacoby, Mark A Schroeder, Geoffrey L Uy, Lisa Y Law, Faisal Cheema, Kendra L Sweet, Adam S Asch, Jijun Jane Liu, Anna B Moseley, Tracy Maher, Laura L Kingsbury, Min Fang, Jerald Radich, Richard F Little, Harry P Erba.

DOI: https://doi.org/10.1080/10428194.2022.2148212
Blood. 2022 Dec 15. 140(24): 2529-2530

An unexpected partnership targeting FLT3wt AML.

Thorsten Braun.

DOI: https://doi.org/10.1182/blood.2022017344
ACS Med Chem Lett. 2022 Dec 08. 13(12): 1885-1891

Development of Gilteritinib-Based Chimeric Small Molecules that Potently Induce Degradation of FLT3-ITD Protein.

Nobumichi Ohoka, Masanori Suzuki, Takuya Uchida, Genichiro Tsuji, Yoshinori Tsukumo, Masayuki Yoshida, Takao Inoue, Yosuke Demizu, Hitoshi Ohki, Mikihiko Naito.

Internal tandem duplication (ITD) in the gene encoding FMS-like tyrosine kinase 3 (FLT3) (FLT3-ITD) is the most frequently observed mutation in acute myeloid leukemia (AML). Currently approved FLT3 kinase inhibitors have high efficacy, but drug resistance caused by reactivation of FLT3 kinase activity is often clinically observed. In this study, we developed novel FLT3 degraders by introducing gilteritinib, an FDA-approved FLT3 inhibitor, into targeted protein degradation technology. The most active compound, CRBN(FLT3)-8, potently degraded FLT3-ITD via the ubiquitin-proteasome system and inhibited the proliferation of FLT3-ITD mutant AML cells more effectively than gilteritinib. These findings provide a new lead compound for degradation-based drugs targeting FLT3-ITD-positive cancers.

DOI: https://doi.org/10.1021/acsmedchemlett.2c00402
Haematologica. 2022 Dec 15.

Ex vivo venetoclax sensitivity testing predicts treatment response in acute myeloid leukemia.

Heikki Kuusanmäki, Sari Kytölä, Ida Vänttinen, Tanja Ruokoranta, Amanda Ranta, Jani Huuhtanen, Minna Suvela, Alun Parsons, Annasofia Holopainen, Anu Partanen, Milla E L Kuusisto, Sirpa Koskela, Riikka Räty, Maija Itälä-Remes, Imre Västrik, Olli Dufva, Sanna Siitonen, Kimmo Porkka, Krister Wennerberg, Caroline A Heckman, Pia Ettala, Marja Pyörälä, Johanna Rimpiläinen, Timo Siitonen, Mika Kontro.

The BCL2 inhibitor venetoclax has revolutionized the treatment of acute myeloid leukemia (AML) patients not benefitting from intensive chemotherapy. Nevertheless, treatment failure remains a challenge, and predictive markers are needed, particularly for relapsed or refractory (R/R) AML. Ex vivo drug sensitivity testing may correlate with outcomes, but its prospective predictive value remains unexplored. Here we report the results of the first stage of the prospective Phase 2 VenEx trial evaluating the utility and predictiveness of venetoclax sensitivity testing using different cell culture conditions and cell viability assays in patients receiving venetoclax-azacitidine (NCT04267081). Participants with de novo AML ineligible for intensive chemotherapy, R/R AML, or secondary AML were included. The primary endpoint was the treatment response in ex vivo sensitive participants and the key secondary endpoints were the correlation of sensitivity with responses and survival. Venetoclax sensitivity testing was successful in 38/39 participants. Experimental conditions significantly influenced predictive accuracy. Blast-specific venetoclax sensitivity measured in conditioned medium most accurately correlated with treatment outcomes; 88% of sensitive participants achieved treatment response. Median survival was significantly longer for ex vivo sensitive participants (14. 6 months for s ensitive, 3. 5 for insensitive, p < 0 . 001). T his analysis illustrates the feasibility of integrating drug-response profiling into clinical practice and demonstrates excellent predictivity.

DOI: https://doi.org/10.3324/haematol.2022.281692
PLoS One. 2022 ;17(12): e0278209

NUC-7738 regulates β-catenin signalling resulting in reduced proliferation and self-renewal of AML cells.

Akbar Muhammed Shahid, In Hwa Um, Mustafa Elshani, Ying Zhang, David James Harrison.

Acute myeloid leukemia (AML) stem cells are required for the initiation and maintenance of the disease. Activation of the Wnt/β-catenin pathway is required for the survival and development of AML leukaemia stem cells (LSCs) and therefore, targeting β-catenin is a potential therapeutic strategy. NUC-7738, a phosphoramidate transformation of 3'-deoxyadenosine (3'-dA) monophosphate, is specifically designed to generate the active anti-cancer metabolite 3'-deoxyadenosine triphosphate (3'-dATP) intracellularly, bypassing key limitations of breakdown, transport, and activation. NUC-7738 is currently in a Phase I/II clinical study for the treatment of patients with advanced solid tumors. Protein expression and immunophenotypic profiling revealed that NUC-7738 caused apoptosis in AML cell lines through reducing PI3K-p110α, phosphorylated Akt (Ser473) and phosphorylated GSK3β (Ser9) resulting in reduced β-catenin, c-Myc and CD44 expression. NUC-7738 reduced β-catenin nuclear expression in AML cells. NUC-7738 also decreased the percentage of CD34+ CD38- CD123+ (LSC-like cells) from 81% to 47% and reduced the total number and size of leukemic colonies. These results indicate that therapeutic targeting of the PI3K/Akt/GSK3β axis can inhibit β-catenin signalling, resulting in reduced clonogenicity and eventual apoptosis of AML cells.

DOI: https://doi.org/10.1371/journal.pone.0278209
Leukemia. 2022 Dec 14.

A pro B cell population forms the apex of the leukemic hierarchy in Hoxa9/Meis1-dependent AML.

Anna Lieske, Eric Agyeman-Duah, Anton Selich, Nicole Dörpmund, Steven R Talbot, Axel Schambach, Tobias Maetzig.

Relapse is a major challenge to therapeutic success in acute myeloid leukemia (AML) and can be partly associated with heterogeneous leukemic stem cell (LSC) properties. In the murine Hoxa9/Meis1-dependent (H9M) AML model, LSC potential lies in three defined immunophenotypes, including Lin-cKit+ progenitor cells (Lin-), Gr1+CD11b+cKit+ myeloid cells, and lymphoid cells (Lym+). Previous reports demonstrated their interconversion and distinct drug sensitivities. In contrast, we here show that H9M AML is hierarchically organized. We, therefore, tracked the developmental potential of LSC phenotypes. This unexpectedly revealed a substantial fraction of Lin- LSCs that failed to regenerate Lym+ LSCs, and that harbored reduced leukemogenic potential. However, Lin- LSCs capable of producing Lym+ LSCs as well as Lym+ LSCs triggered rapid disease development suggestive of their high relapse-driving potential. Transcriptional analyses revealed that B lymphoid master regulators, including Sox4 and Bach2, correlated with Lym+ LSC development and presumably aggressive disease. Lentiviral overexpression of Sox4 and Bach2 induced dedifferentiation of H9M cells towards a lineage-negative state in vitro as the first step of lineage conversion. This work suggests that the potency to initiate a partial B lymphoid primed transcriptional program as present in infant AML correlates with aggressive disease and governs the H9M LSC hierarchy.

DOI: https://doi.org/10.1038/s41375-022-01775-y
Cell Rep. 2022 Dec 13. pii: S2211-1247(22)01717-X. [Epub ahead of print]41(11): 111825

Splicing factor deficits render hematopoietic stem and progenitor cells sensitive to STAT3 inhibition.

Kathryn S Potts, Rosannah C Cameron, Amina Metidji, Noura Ghazale, LaShanale Wallace, Ana I Leal-Cervantes, Reid Palumbo, Juan Martin Barajas, Varun Gupta, Srinivas Aluri, Kith Pradhan, Jacquelyn A Myers, Mia McKinstry, Xiaoying Bai, Gaurav S Choudhary, Aditi Shastri, Amit Verma, Esther A Obeng, Teresa V Bowman.

  Hematopoietic stem and progenitor cells (HSPCs) sustain lifelong hematopoiesis. Mutations of pre-mRNA splicing machinery, especially splicing factor 3b, subunit 1 (SF3B1), are early lesions found in malignancies arising from HSPC dysfunction. However, why splicing factor deficits contribute to HSPC defects remains incompletely understood. Using zebrafish, we show that HSPC formation in sf3b1 homozygous mutants is dependent on STAT3 activation. Clinically, mutations in SF3B1 are heterozygous; thus, we explored if targeting STAT3 could be a vulnerability in these cells. We show that SF3B1 heterozygosity confers heightened sensitivity to STAT3 inhibition in zebrafish, mouse, and human HSPCs. Cells carrying mutations in other splicing factors or treated with splicing modulators are also more sensitive to STAT3 inhibition. Mechanistically, we illustrate that STAT3 inhibition exacerbates aberrant splicing in SF3B1 mutant cells. Our findings reveal a conserved vulnerability of splicing factor mutant HSPCs that could allow for their selective targeting in hematologic malignancies.

Keywords:  CP: Molecular biology; CP: Stem cell research; SF3B1; STAT3; hematopoietic stem and progenitor cell; myelodysplastic syndrome; splicing factor; zebrafish

DOI:  https://doi.org/10.1016/j.celrep.2022.111825
Leukemia. 2022 Dec 12.

Infectious complications after intensive chemotherapy with CLAG-M versus 7+3 for AML and other high-grade myeloid neoplasms.

Carla S Walti, Anna B Halpern, Hu Xie, Erika S Kiem, E Lisa Chung, Kelda G Schonhoff, Emily M Huebner, Colleen Delaney, Catherine Liu, Steven A Pergam, Guang-Shing Cheng, Louise E Kimball, Wendy M Leisenring, Michael Boeckh, Roland B Walter, Joshua A Hill.

Contemporary data on infections after intensive chemotherapy for acute myeloid leukemia (AML) are scarce. Cladribine, high-dose cytarabine, G-CSF, and dose-escalated mitoxantrone ("CLAG-M") may result in higher remission rates than standard-dose cytarabine plus anthracycline ("7 + 3") but may result in more infections. We compared moderate to severe infections occurring up to 90 days after the first induction cycle for AML or other high-grade myeloid neoplasms in patients receiving CLAG-M for newly diagnosed (n = 196) or relapsed/refractory disease (n = 131) or 7 + 3 for newly diagnosed disease (n = 115). For newly diagnosed disease, microbiologically documented infections were more frequent after CLAG-M compared to 7 + 3 (adjusted rate ratio, 1.65 [95% CI, 1.06-2.58]; P = 0.03), with a cumulative incidence of 27.8% and 16.5% by day 90, respectively. Patients receiving CLAG-M for relapsed/refractory disease had the highest cumulative incidence of 50.7%. Bacterial bloodstream infections were the most frequent followed by respiratory tract infections. Among 29 patients (7%) who died, infection was a primary or contributing cause of death in 59%. These data indicate that infections continue to cause substantial morbidity in patients treated for AML, especially those treated for relapsed/refractory disease, and are more common with newer, more myelosuppressive regimens such as CLAG-M. Improved strategies for infection prevention are needed.

DOI: https://doi.org/10.1038/s41375-022-01786-9
Cancers (Basel). 2022 Dec 04. pii: 5989. [Epub ahead of print]14(23):

Viewing AML through a New Lens: Technological Advances in the Study of Epigenetic Regulation.

Laura C Godfrey, Alba Rodriguez-Meira.

  Epigenetic modifications, such as histone modifications and DNA methylation, are essential for ensuring the dynamic control of gene regulation in every cell type. These modifications are associated with gene activation or repression, depending on the genomic context and specific type of modification. In both cases, they are deposited and removed by epigenetic modifier proteins. In acute myeloid leukemia (AML), the function of these proteins is perturbed through genetic mutations (i.e., in the DNA methylation machinery) or translocations (i.e., MLL-rearrangements) arising during leukemogenesis. This can lead to an imbalance in the epigenomic landscape, which drives aberrant gene expression patterns. New technological advances, such as CRISPR editing, are now being used to precisely model genetic mutations and chromosomal translocations. In addition, high-precision epigenomic editing using dCas9 or CRISPR base editing are being used to investigate the function of epigenetic mechanisms in gene regulation. To interrogate these mechanisms at higher resolution, advances in single-cell techniques have begun to highlight the heterogeneity of epigenomic landscapes and how these impact on gene expression within different AML populations in individual cells. Combined, these technologies provide a new lens through which to study the role of epigenetic modifications in normal hematopoiesis and how the underlying mechanisms can be hijacked in the context of malignancies such as AML.

Keywords:  AML; CRISPR; DNA methylation; epigenomic editing; hematopoiesis; histone modifications; leukemia; single-cell technology

DOI:  https://doi.org/10.3390/cancers14235989
Nat Immunol. 2022 Dec 15.

A genetic disorder reveals a hematopoietic stem cell regulatory network co-opted in leukemia.

Richard A Voit, Liming Tao, Fulong Yu, Liam D Cato, Blake Cohen, Travis J Fleming, Mateusz Antoszewski, Xiaotian Liao, Claudia Fiorini, Satish K Nandakumar, Lara Wahlster, Kristian Teichert, Aviv Regev, Vijay G Sankaran.

The molecular regulation of human hematopoietic stem cell (HSC) maintenance is therapeutically important, but limitations in experimental systems and interspecies variation have constrained our knowledge of this process. Here, we have studied a rare genetic disorder due to MECOM haploinsufficiency, characterized by an early-onset absence of HSCs in vivo. By generating a faithful model of this disorder in primary human HSCs and coupling functional studies with integrative single-cell genomic analyses, we uncover a key transcriptional network involving hundreds of genes that is required for HSC maintenance. Through our analyses, we nominate cooperating transcriptional regulators and identify how MECOM prevents the CTCF-dependent genome reorganization that occurs as HSCs differentiate. We show that this transcriptional network is co-opted in high-risk leukemias, thereby enabling these cancers to acquire stem cell properties. Collectively, we illuminate a regulatory network necessary for HSC self-renewal through the study of a rare experiment of nature.

DOI: https://doi.org/10.1038/s41590-022-01370-4
Leukemia. 2022 Dec 16.

Asxl1 deletion disrupts MYC and RNA polymerase II function in granulocyte progenitors.

Theodore P Braun, Joseph Estabrook, Zachary Schonrock, Brittany M Curtiss, Lucie Darmusey, Jommel Macaraeg, Trevor Enright, Cody Coblentz, Rowan Callahan, William Yashar, Akram Taherinasab, Hisham Mohammed, Daniel J Coleman, Brian J Druker, Emek Demir, Theresa A Lusardi, Julia E Maxson.

Mutations in the gene Additional Sex-Combs Like 1 (ASXL1) are recurrent in myeloid malignancies as well as the pre-malignant condition clonal hematopoiesis, where they are universally associated with poor prognosis. However, the role of ASXL1 in myeloid lineage maturation is incompletely described. To define the role of ASXL1 in myelopoiesis, we employed single cell RNA sequencing and a murine model of hematopoietic-specific Asxl1 deletion. In granulocyte progenitors, Asxl1 deletion leads to hyperactivation of MYC and a quantitative decrease in neutrophil production. This loss of granulocyte production was not accompanied by significant changes in the landscape of covalent histone modifications. However, Asxl1 deletion results in a decrease in RNAPII promoter-proximal pausing in granulocyte progenitors, indicative of a global increase in productive transcription. These results suggest that ASXL1 inhibits productive transcription in granulocyte progenitors, identifying a new role for this epigenetic regulator in myeloid development.

DOI: https://doi.org/10.1038/s41375-022-01792-x
Am J Cancer Res. 2022 ;12(11): 5140-5159

Deciphering the immuno-pathological role of FLT, and evaluation of a novel dual inhibitor of topoisomerases and mutant-FLT3 for treating leukemia.

Bashir Lawal, Yu-Cheng Kuo, Harshita Khedkar, Ntlotlang Mokgautsi, Maryam Rachmawati Sumitra, Alexander Th Wu, Hsu-Shan Huang.

Acute myeloid leukemia (AML) is a type of leukemia with an aggressive phenotype, that commonly occurs in adults and with disappointing treatment outcomes. Genetic alterations were implicated in the etiology of cancers and form the basis for defining patient prognoses and guiding targeted therapies. In the present study, we leveraged bulk and single-cell RNA sequencing datasets from AML patients to determine the clinical significance of Fms-related receptor tyrosine kinase 3 (FLT3) alterations on the T-cell phenotype and immune response of AML patients. Subsequently, we evaluated the therapeutic potential of Lwk-n019, a novel small-molecule derivative of thiochromeno[2,3-c]quinolin-12-one. Our results suggested that FLT3 plays an important role in the progression, aggressive phenotype, and worse immune response of patients. An FLT3 mutation was associated with dysfunctional T-cell phenotypes, and high risk and shorter survival of AML patients. Our findings further suggested that the aggressiveness of AML and the prognostic role of FLT3 are associated with the co-occurrence of NPM1 and DNMT3A mutations. Lwk-n019 demonstrated dose-dependent anticancer activities against various leukemia cancer cell lines. Lwk-n019 demonstrated highly selective kinase inhibitory activities against the wild-type FLT3 (D835V) and mutant FLT3 (internal tandem duplication (ITD), D835V) with >95% and 99% inhibitory levels, respectively. Moreover, the compound demonstrated the best binding constant (Kd value) of 0.77 µM against FLT3 (ITD, 835V). In addition, Lwk-n019 significantly inhibited the activities of both the topoisomerase I (TOPI) and TOPII enzymes, with higher TOPI inhibitory activity than camptothecin, a clinical inhibitor. While the jejunum, duodenum, cecum, and colon were prime sites of absorption, Lwk-n019 achieved maximum concentration (Cmax), Vd, blood/plasma ratio, time to maximum concentration (Tmax), area under the receiver operating concentration curve (AUC)(0-24), and AUC(0-∞) values of 0.665 µg/mL, 5.21 Vc, L/kg, 1.5 h, 6634.7, and 6909.2, respectively. In conclusion, Lwk-n019 demonstrated anticancer activities via multi-target inhibition of TOPs and kinases with high inhibition preference for mutant ITD-FLT3. The present pioneer study provides a basis for advanced optimization of drug potency, selectivity, specificity, and other properties desired of anticancer drug leads. Studies are ongoing to determine the full therapeutic properties of Lwk-n019 and the detailed mechanisms of FLT3 in TOP inhibition.

Keywords: Acute myeloid leukemia; FLT3; Lwk-n019; immune infiltration; topoisomerase
Int J Mol Sci. 2022 Nov 25. pii: 14706. [Epub ahead of print]23(23):

Heterogeneity of Patient-Derived Acute Myeloid Leukemia Cells Subjected to SYK In Vitro Inhibition.

Marte Karen Brattås, Anette Lodvir Hemsing, Kristin Paulsen Rye, Kimberley Joanne Hatfield, Håkon Reikvam.

  Acute myeloid leukemia (AML) is an aggressive hematological malignancy with a dismal prognosis. The cytoplasmic spleen tyrosine kinase (SYK) is highly expressed by hematopoietic cells and has emerged as a potential therapeutic target. In this study, we evaluated the in vitro antileukemic effects of five SYK inhibitors, fostamatinib, entospletinib, cerdulatinib, TAK-659, and RO9021, in a consecutive AML patient cohort. All inhibitors demonstrated a concentration-dependent antiproliferative effect, although there was considerable heterogeneity among patients. For fostamatinib and TAK-659, the antiproliferative effects were significantly higher in FLT3 mutated patients compared to nonmutated patients. Fostamatinib, entospletinib, TAK-659, and RO9021 induced significant apoptosis in primary AML cells, although the proapoptotic effects of the SYK inhibitors were less pronounced than the antiproliferative effects. Finally, most of the SYK inhibitors caused a significant decrease in the release of cytokines and chemokines from primary AML cells, indicating a potent inhibitory effect on the release of these leukemic signaling molecules. We concluded that the SYK inhibitors had antileukemic effects in AML, although larger studies are strongly needed to identify which patient subsets will benefit most from such a treatment.

Keywords:  acute myeloid leukemia (AML); apoptosis; cytokines; proliferation; spleen tyrosine kinase (SYK)

DOI:  https://doi.org/10.3390/ijms232314706
Future Oncol. 2022 Dec 12.

Indirect treatment comparison of oral versus injectable azacitidine as maintenance therapy for acute myeloid leukemia.

Ashley Tabah, David Huggar, Si-Tien Wang, Scott J Johnson, Ronda M Copher, Thomas O'Connell, Ali McBride, Thomas W LeBlanc.

Aim: Evaluate the relative efficacy of oral versus injectable azacitidine (AZA) maintenance therapy in acute myeloid leukemia (AML) after complete remission. Materials & methods: Systematic literature review identified QUAZAR AML-001, HOVON 97 AML, UK NCRI AML16 and QoLESS-AZA-AMLE (sensitivity analysis) trials. Network meta-analysis and matching-adjusted indirect comparisons assessed survival outcomes. Results: In the network meta-analysis, combining the HOVON 97 and UK NCRI trials, oral AZA (QUAZAR) was associated with significantly improved overall survival (OS) versus injectable AZA (hazard ratio: 0.744; 95% credible interval: 0.557-0.998). After matching-adjusted indirect comparisons, to address differences in patient characteristics across trials, OS improvements were maintained with oral versus injectable AZA (hazard ratio: 0.753; credible interval: 0.563-0.998). Conclusion: In AML, maintenance therapy with oral AZA was associated with improved OS versus injectable AZA.

DOI: https://doi.org/10.2217/fon-2022-0820
Nat Commun. 2022 Dec 10. 13(1): 7657

Myeloid cells promote interferon signaling-associated deterioration of the hematopoietic system.

Jacqueline Feyen, Zhen Ping, Lanpeng Chen, Claire van Dijk, Tim V D van Tienhoven, Paulina M H van Strien, Remco M Hoogenboezem, Michiel J W Wevers, Mathijs A Sanders, Ivo P Touw, Marc H G P Raaijmakers.

Innate and adaptive immune cells participate in the homeostatic regulation of hematopoietic stem cells (HSCs). Here, we interrogate the contribution of myeloid cells, the most abundant cell type in the mammalian bone marrow, in a clinically relevant mouse model of neutropenia. Long-term genetic depletion of neutrophils and eosinophils results in activation of multipotent progenitors but preservation of HSCs. Depletion of myeloid cells abrogates HSC expansion, loss of serial repopulation and lymphoid reconstitution capacity and remodeling of HSC niches, features previously associated with hematopoietic aging. This is associated with mitigation of interferon signaling in both HSCs and their niches via reduction of NK cell number and activation. These data implicate myeloid cells in the functional decline of hematopoiesis, associated with activation of interferon signaling via a putative neutrophil-NK cell axis. Innate immunity may thus come at the cost of system deterioration through enhanced chronic inflammatory signaling to stem cells and their niches.

DOI: https://doi.org/10.1038/s41467-022-35318-x
Blood Adv. 2022 Dec 15. pii: bloodadvances.2022008996. [Epub ahead of print]

Adaptation of Serious Illness Care Program to be Delivered via Telehealth for Older Patients with Hematologic Malignancy.

Marissa Locastro, Chandrika Sanapala, Jason H Mendler, Sally A Norton, Rachelle Bernacki, Thomas Carroll, Heidi D Klepin, Erin Elizabeth Watson, Jane L Liesveld, Eric J Huselton, Kristen M O'Dwyer, Andrea M Baran, Marie Flannery, Benzi Kluger, Kah Poh Loh.

Older patients with acute myeloid leukemia (AML) and myelodysplastic syndromes (MDS) experience intense inpatient healthcare at the end-of-life (EOL) . Early advance care planning (ACP) may improve care at EOL for patients with AML and MDS. The Serious Illness Care Program (SICP) is a multicomponent, communication intervention developed to improve conversations about values for patients with serious illnesses. The SICP has been shown to improve the quality and frequency of ACP discussions. We adapted the SICP for delivery via telehealth to older patients with AML and MDS. We conducted a single-center qualitative study of 45 participants (25 clinicians, 15 older patients with AML and MDS, and 5 caregivers). Participants, whether clinicians, patients, or caregivers, agreed that the SICP would help older patients with AML and MDS to share their personal values with their care team. Four qualitative themes emerged from our data: 1) Serious illness conversations can be conducted via telehealth, 2) Older patients have limited experience using technology but are willing and able to learn, 3) Patients feel that serious illness conversations will help them understand their AML or MDS diagnosis and prognosis better, and 4) Serious illness conversations should be common and routine, not extra-ordinary. The adapted SICP may provide older patients with AML and MDS an opportunity to share what matters most to them with their care team and may assist oncologists in aligning patient care with patient values. The adapted SICP is the subject of an ongoing single-arm pilot study at the Wilmot Cancer Institute.

DOI: https://doi.org/10.1182/bloodadvances.2022008996
Exp Hematol. 2022 Nov 26. pii: S0301-472X(22)00809-8. [Epub ahead of print]

Establishment of isogenic induced pluripotent stem cells with or without pathogenic mutation for understanding the pathogenesis of myeloproliferative neoplasms.

Chang Liu, Misa Imai, Yoko Edahiro, Shuichi Mano, Hiraku Takei, Mai Nudejima, Akira Kurose, Soji Morishita, Miki Ando, Satoshi Tsuneda, Marito Araki, Norio Komatsu.

Identification and functional characterization of disease-associated genetic traits are crucial for understanding the pathogenesis of hematologic malignancies. Various in vitro and in vivo models, including cell lines, primary cells, and animal models, have been established to examine these genetic alterations. However, their nonphysiologic conditions, diverse genetic backgrounds, and species-specific differences often limit data interpretation. To evaluate somatic mutations in myeloproliferative neoplasms (MPNs), we used CRISPR/Cas9 combined with the piggyBac transposon system to establish isogenic induced pluripotent stem (iPS) cell lines with or without JAK2V617F mutation, a driver mutation of MPNs. We induced hematopoietic stem/progenitor cells (HSPCs) from these iPS cells and observed phenotypic differences during hematopoiesis using fluorescence-activated cell sorting analysis. HSPCs with pathogenic mutations exhibited cell-autonomous erythropoiesis and megakaryopoiesis, which are hallmarks in the bone marrow of patients with MPNs. Furthermore, we used these HSPCs as a model to validate therapeutic compounds and showed that interferon alpha selectively inhibited erythropoiesis and megakaryopoiesis in mutant HSPCs. These results demonstrate that genome editing is feasible for establishing isogenic iPS cells, studying genetic elements to understand the pathogenesis of MPNs, and evaluating therapeutic compounds against MPNs.

DOI: https://doi.org/10.1016/j.exphem.2022.11.005
Future Oncol. 2022 Dec 16.

Asciminib monotherapy for newly diagnosed chronic myeloid leukemia in chronic phase: the ASC4FIRST phase III trial.

Jorge E Cortes, Andreas Hochhaus, Naoto Takahashi, Richard A Larson, Ghayas C Issa, Felice Bombaci, Nicholas Ramscar, Sophie Ifrah, Timothy P Hughes.

  Asciminib is the first BCR::ABL1 inhibitor that works by Specifically Targeting the ABL Myristoyl Pocket (STAMP). Asciminib has shown favorable efficacy and safety in patients with chronic myeloid leukemia in chronic phase without the T315I mutation who have received ≥2 prior tyrosine kinase inhibitors (TKIs) in phase I and III clinical trials and in patients with the T315I mutation who have received ≥1 prior TKI in Phase I. ASC4FIRST (NCT04971226) is a phase III, multicenter, open-label, randomized study of asciminib versus investigator-selected TKI in patients with newly diagnosed chronic myeloid leukemia in chronic phase. The primary end point is major molecular response at week 48. Secondary end points include responses at and by scheduled time points, safety, pharmacokinetics and patient-reported outcomes. Clinical Trial Registration: NCT04971226 (ClinicalTrials.gov).

Keywords:  BCR::ABL1; asciminib; chronic myeloid leukemia; chronic phase; first line; newly diagnosed; ongoing clinical trial

DOI:  https://doi.org/10.2217/fon-2022-0923
Ann Hematol. 2022 Dec 17.

Integrated single-cell transcriptome analysis of CD34 + enriched leukemic stem cells revealed intra- and inter-patient transcriptional heterogeneity in pediatric acute myeloid leukemia.

Deepshi Thakral, Vivek Kumar Singh, Ritu Gupta, Nitu Jha, Aafreen Khan, Gurvinder Kaur, Sandeep Rai, Vijay Kumar, Manisha Supriya, Sameer Bakhshi, Rachna Seth.

  To gain insights into the idiosyncrasies of CD34 + enriched leukemic stem cells, we investigated the nature and extent of transcriptional heterogeneity by single-cell sequencing in pediatric AML. Whole transcriptome analysis of 28,029 AML single cells was performed using the nanowell cartridge-based barcoding technology. Integrated transcriptional analysis identified unique leukemic stem cell clusters of each patient and intra-patient heterogeneity was revealed by multiple LSC-enriched clusters differing in their cell cycle processes and BCL2 expression. All LSC-enriched clusters exhibited gene expression profile of dormancy and self-renewal. Upregulation of genes involved in non-coding RNA processing and ribonucleoprotein assembly were observed in LSC-enriched clusters relative to HSC. The genes involved in regulation of apoptotic processes, response to cytokine stimulus, and negative regulation of transcription were upregulated in LSC-enriched clusters as compared to the blasts. Validation of top altered genes in LSC-enriched clusters confirmed upregulation of TCF7L2, JUP, ARHGAP25, LPAR6, and PRDX1 genes, and serine/threonine kinases (STK24, STK26). Upregulation of LPAR6 showed trend towards MRD positive status (Odds ratio = 0.126; 95% CI = 0.0144-1.10; p = 0.067) and increased expression of STK26 significantly correlated with higher RFS (HR = 0.231; 95% CI = 0.0506-1.052; p = 0.04). Our findings addressed the inter- and intra-patient diversity within AML LSC and potential signaling and chemoresistance-associated targets that warrant investigation in larger cohort that may guide precision medicine in the near future.

Keywords:  Acute myeloid leukemia; DEGs; Gene expression profile; Leukemic stem cells; Pediatric AML; Single-cell sequencing; Transcriptional heterogeneity

DOI:  https://doi.org/10.1007/s00277-022-05021-4
Sci Adv. 2022 Dec 16. 8(50): eabp8293

In vivo characterization of glutamine metabolism identifies therapeutic targets in clear cell renal cell carcinoma.

Akash K Kaushik, Amy Tarangelo, Lindsey K Boroughs, Mukundan Ragavan, Yuanyuan Zhang, Cheng-Yang Wu, Xiangyi Li, Kristen Ahumada, Jui-Chung Chiang, Vanina T Tcheuyap, Faeze Saatchi, Quyen N Do, Cissy Yong, Tracy Rosales, Christina Stevens, Aparna D Rao, Brandon Faubert, Panayotis Pachnis, Lauren G Zacharias, Hieu Vu, Feng Cai, Thomas P Mathews, Giannicola Genovese, Barbara S Slusher, Payal Kapur, Xiankai Sun, Matthew Merritt, James Brugarolas, Ralph J DeBerardinis.

Targeting metabolic vulnerabilities has been proposed as a therapeutic strategy in renal cell carcinoma (RCC). Here, we analyzed the metabolism of patient-derived xenografts (tumorgrafts) from diverse subtypes of RCC. Tumorgrafts from VHL-mutant clear cell RCC (ccRCC) retained metabolic features of human ccRCC and engaged in oxidative and reductive glutamine metabolism. Genetic silencing of isocitrate dehydrogenase-1 or isocitrate dehydrogenase-2 impaired reductive labeling of tricarboxylic acid (TCA) cycle intermediates in vivo and suppressed growth of tumors generated from tumorgraft-derived cells. Glutaminase inhibition reduced the contribution of glutamine to the TCA cycle and resulted in modest suppression of tumorgraft growth. Infusions with [amide-15N]glutamine revealed persistent amidotransferase activity during glutaminase inhibition, and blocking these activities with the amidotransferase inhibitor JHU-083 also reduced tumor growth in both immunocompromised and immunocompetent mice. We conclude that ccRCC tumorgrafts catabolize glutamine via multiple pathways, perhaps explaining why it has been challenging to achieve therapeutic responses in patients by inhibiting glutaminase.

DOI: https://doi.org/10.1126/sciadv.abp8293
J Biol Chem. 2022 Dec 09. pii: S0021-9258(22)01230-3. [Epub ahead of print] 102787

DNA damage to bone marrow stromal cells by anti-leukemia drugs induces chemo-resistance in acute myeloid leukemia via paracrine FGF10-FGFR2 signaling.

Shuang Yu, Jingjing Ye, Yingqiao Wang, Ting Lu, Yan Liu, Na Liu, Jingru Zhang, Fei Lu, Daoxin Ma, Robert Peter Gale, Chunyan Ji.

  Chemo-resistance remains a major challenge in the current treatment of acute myeloid leukemia (AML). The bone marrow microenvironment (BMM) plays a complex role in protecting leukemia cells from chemotherapeutics, and the mechanisms involved are not fully understood. Anti-leukemia drugs kill AML cells directly but also damage the BMM. Here, we determined anti-leukemia drugs induce DNA damage in bone marrow stromal cells (BMSCs), resulting in resistance of AML cell lines to adriamycin and idarubicin killing. Damaged BMSCs induced an inflammatory microenvironment through NF-κB; suppressing NF-κB with small molecule inhibitor Bay11-7082 attenuated the pro-survival effects of BMSCs on AML cell lines. Furthermore, we used an ex vivo functional screen of 507 chemokines and cytokines to identify 44 proteins secreted from damaged BMSCs. Fibroblast growth factor-10 (FGF10) was most strongly associated with chemo-resistance in AML cell lines. Additionally, expression of FGF10 and its receptors, FGFR1 and FGFR2, was increased in AML patients after chemotherapy. FGFR1 and FGFR2 were also widely expressed by AML cell lines. FGF10-induced FGFR2 activation in AML cell lines operates by increasing P38 MAPK, AKT, ERK1/2, and STAT3 phosphorylation. FGFR2 inhibition with small molecules, or gene silencing of FGFR2 inhibited proliferation and reverses drug-resistance of AML cells by inhibiting P38 MAPK, AKT, and ERK1/2 signaling pathways. Finally, release of FGF10 was mediated by β-catenin signaling in damaged BMSCs. Our data indicate FGF10-FGFR2 signaling acts as an effector of damaged BMSC-mediated chemo-resistance in AML cells, and FGFR2 inhibition can reverse stromal protection and AML cell chemo-resistance in the BMM.

Keywords:  DNA damage; FGF10; acute myeloid leukemia; bone marrow stromal cells; chemo-resistance

DOI:  https://doi.org/10.1016/j.jbc.2022.102787
Nat Genet. 2022 Dec 15.

Human genetic diversity alters off-target outcomes of therapeutic gene editing.

Samuele Cancellieri, Jing Zeng, Linda Yingqi Lin, Manuel Tognon, My Anh Nguyen, Jiecong Lin, Nicola Bombieri, Stacy A Maitland, Marioara-Felicia Ciuculescu, Varun Katta, Shengdar Q Tsai, Myriam Armant, Scot A Wolfe, Rosalba Giugno, Daniel E Bauer, Luca Pinello.

CRISPR gene editing holds great promise to modify DNA sequences in somatic cells to treat disease. However, standard computational and biochemical methods to predict off-target potential focus on reference genomes. We developed an efficient tool called CRISPRme that considers single-nucleotide polymorphism (SNP) and indel genetic variants to nominate and prioritize off-target sites. We tested the software with a BCL11A enhancer targeting guide RNA (gRNA) showing promise in clinical trials for sickle cell disease and β-thalassemia and found that the top candidate off-target is produced by an allele common in African-ancestry populations (MAF 4.5%) that introduces a protospacer adjacent motif (PAM) sequence. We validated that SpCas9 generates strictly allele-specific indels and pericentric inversions in CD34+ hematopoietic stem and progenitor cells (HSPCs), although high-fidelity Cas9 mitigates this off-target. This report illustrates how genetic variants should be considered as modifiers of gene editing outcomes. We expect that variant-aware off-target assessment will become integral to therapeutic genome editing evaluation and provide a powerful approach for comprehensive off-target nomination.

DOI: https://doi.org/10.1038/s41588-022-01257-y
Blood Adv. 2022 Dec 15. pii: bloodadvances.2022008221. [Epub ahead of print]

Venetoclax treatment in cancer patients has limited impact on circulating T and NK cells.

Charis E Teh, Hongke Peng, Mengxiao Luo, Tania Tan, Marie Trussart, Lauren J Howson, Chong Chyn Chua, Christine Muttiah, Fiona C Brown, Matthew E Ritchie, Andrew H Wei, Andrew W Roberts, Vanessa L Bryant, Mary Ann Anderson, Geoffrey J Lindeman, David Ching Siang Huang, Rachel Thijssen, Daniel H D Gray.

Venetoclax is an effective treatment for certain blood cancers, such as chronic lymphocytic leukemia (CLL) and acute myeloid leukemia (AML). However, most patients relapse while on venetoclax and further treatment options become limited. Combining venetoclax with immunotherapies is an attractive approach; however, a detailed understanding of how venetoclax treatment impacts normal immune cells in patients is lacking. In this study, we performed deep profiling of peripheral blood cells from CLL and AML patients before and after short-term treatment with venetoclax using mass cytometry (CyTOF) and found no impact on the concentrations of key T cell subsets nor their expression of checkpoint molecules. We also analyzed peripheral blood from breast cancer patients receiving venetoclax long-term using a single-cell multi-omics approach (CITE-seq) and functional assays. We found significant depletion of B cell populations with low expression of MCL-1 relative to other immune cells, attended by extensive transcriptomic changes. By contrast, there was less impact on circulating T cells and natural killer (NK) cells, with no changes in their subset composition, transcriptome or function following venetoclax treatment. Our data indicate that venetoclax has minimal impact on circulating T or NK cells, supporting the rationale of combining this BH3 mimetic drug with cancer immunotherapies for more durable anti-tumour responses.

DOI: https://doi.org/10.1182/bloodadvances.2022008221
Front Endocrinol (Lausanne). 2022 ;13 1001210

Raman microspectroscopy reveals unsaturation heterogeneity at the lipid droplet level and validates an in vitro model of bone marrow adipocyte subtypes.

Josefine Tratwal, Guillaume Falgayrac, Alexandrine During, Nicolas Bertheaume, Charles Bataclan, Daniel N Tavakol, Vasco Campos, Ludovic Duponchel, George Q Daley, Guillaume Penel, Christophe Chauveau, Olaia Naveiras.

  Bone marrow adipocytes (BMAds) constitute the most abundant stromal component of adult human bone marrow. Two subtypes of BMAds have been described, the more labile regulated adipocytes (rBMAds) and the more stable constitutive adipocytes (cBMAds), which develop earlier in life and are more resilient to environmental and metabolic disruptions. In vivo, rBMAds are enriched in saturated fatty acids, contain smaller lipid droplets (LDs) and more readily provide hematopoietic support than their cBMAd counterparts. Mouse models have been used for BMAds research, but isolation of primary BMAds presents many challenges, and thus in vitro models remain the current standard to study nuances of adipocyte differentiation. No in vitro model has yet been described for the study of rBMAds/cBMAds. Here, we present an in vitro model of BM adipogenesis with differential rBMAd and cBMAd-like characteristics. We used OP9 BM stromal cells derived from a (C57BL/6xC3H)F2-op/op mouse, which have been extensively characterized as feeder layer for hematopoiesis research. We observed similar canonical adipogenesis transcriptional signatures for spontaneously-differentiated (sOP9) and induced (iOP9) cultures, while fatty acid composition and desaturase expression of Scd1 and Fads2 differed at the population level. To resolve differences at the single adipocyte level we tested Raman microspectroscopy and show it constitutes a high-resolution method for studying adipogenesis in vitro in a label-free manner, with resolution to individual LDs. We found sOP9 adipocytes have lower unsaturation ratios, smaller LDs and higher hematopoietic support than iOP9 adipocytes, thus functionally resembling rBMAds, while iOP9 more closely resembled cBMAds. Validation in human primary samples confirmed a higher unsaturation ratio for lipids extracted from stable cBMAd-rich sites (femoral head upon hip-replacement surgery) versus labile rBMAds (iliac crest after chemotherapy). As a result, the 16:1/16:0 fatty acid unsaturation ratio, which was already shown to discriminate BMAd subtypes in rabbit and rat marrow, was validated to discriminate cBMAds from rBMAd in both the OP9 model in vitro system and in human samples. We expect our model will be useful for cBMAd and rBMAd studies, particularly where isolation of primary BMAds is a limiting step.

Keywords:  OP9 cell; bone marrow stromal cell (BMSC); constitutive bone marrow adiposity; fatty acid; lipid droplet diameter; regulated bone marrow adiposity; stroma; unsaturation (fatty acid)

DOI:  https://doi.org/10.3389/fendo.2022.1001210
Haematologica. 2022 Dec 15.

A phase 2, prospective, randomized, open-label study of defibrotide added to standard-of-care prophylaxis for the prevention of acute graft-versus-host disease after allogeneic hematopoietic cell transplantation.

Michelle Hudspeth, Shahram Mori, David Nachbaur, José Antonio Perez-Simon, Friedrich Stölzel, Marcie Riches, Wendy Wu, Peixin Zhang, Shirali Agarwal, Ibrahim Yakoub-Agha.

Acute graft-versus-host disease (aGvHD) is a life-threatening complication typically occurring within 100 days after allogeneic hematopoietic cell transplantation (allo-HCT). This hypothesisgenerating, phase 2, prospective, open-label, randomized study compared defibrotide added to standard-of-care (SOC) GvHD prophylaxis (defibrotide prophylaxis arm) versus SOC alone (SOC arm) to prevent aGvHD post-transplant. This study estimated incidences of aGvHD and was not statistically powered to assess differences among treatment arms. Patients were randomized 1:1 to defibrotide prophylaxis arm (n=79; median age: 57 years, range: 2-69 years) or SOC arm (n=73; median age: 56 years, range: 2-72 years). Patient demographics in the two arms were similar except for conditioning regimen type (myeloablative: defibrotide, 76% vs. SOC, 61%) and stem cell source for allo-HCT (bone marrow: defibrotide, 34% vs. SOC, 26%). In the intent-to-treat primary endpoint analysis, the cumulative incidence of grade B-D aGvHD at Day 100 post-transplant was 38.4% in the defibrotide prophylaxis arm versus 47.1% in the SOC arm (difference: -8.8% [90% confidence interval (CI): -22.5, 4.9]). The difference noted at Day 100 became more pronounced in a subgroup analysis of patients who received antithymocyte globulin (defibrotide: 30.4%, SOC: 47.6%; difference: -17.2% [90% CI: -41.8, 7.5]). Overall survival rates at Day 180 post-transplant were similar between arms, as were the rates of serious treatment-emergent adverse events (defibrotide: 42%, SOC: 44%). While the observed differences in endpoints between the two arms were not substantial, these results suggest defibrotide prophylaxis may add a benefit to currently available SOC to prevent aGvHD following allo-HCT without adding significant toxicities.

DOI: https://doi.org/10.3324/haematol.2022.281471