bims-tremyl 2022-02-27 papers

on Therapy resistance biology in myeloid leukemia

Issue of 2022‒02‒27
twenty-two papers selected by
Paolo Gallipoli
Barts Cancer Institute, Queen Mary University of London

Activation of RAS/MAPK pathway confers MCL-1 mediated acquired resistance to BCL-2 inhibitor venetoclax in acute myeloid leukemia.
Pseudo-mutant P53 is a unique phenotype of DNMT3A-mutated pre-leukemia.
Validation of ALFA 1200 score in patients with AML >60 years treated with double nucleoside-based low-intensity therapy.
Osteopontin is required for the maintenance of leukemia stem cells in acute myeloid leukemia.
A PRC2-Kdm5b axis sustains tumorigenicity of acute myeloid leukemia.
IDH1/2 mutations in acute myeloid leukemia.
The m6A reader IGF2BP3 promotes acute myeloid leukemia progression by enhancing RCC2 stability.
Hematopoietic cell transplantation donor-derived memory-like NK cells functionally persist after transfer into patients with leukemia.
Identification of novel STAT5B mutations and characterization of TCRβ signatures in CD4+ T-cell large granular lymphocyte leukemia.
What Clonal Hematopoiesis Can Teach Us About MDS.
Lasting shift in the gut microbiota in patients with acute myeloid leukemia.
Deneddylation of PML/RARα reconstructs functional PML nuclear bodies via orchestrating phase separation to eradicate APL.
Changes in body mass index, weight, and height in children with acute myeloid leukemia and the associations with outcome.
EZH2 noncanonically binds cMyc and p300 through a cryptic transactivation domain to mediate gene activation and promote oncogenesis.
Next-Generation Sequencing Revealed a Distinct Immunoglobulin Repertoire with Specific Mutation Hotspots in Acute Myeloid Leukemia.
Deep learning identifies Acute Promyelocytic Leukemia in bone marrow smears.
Phosphoproteomic profiling of T cell acute lymphoblastic leukemia reveals targetable kinases and combination treatment strategies.
Mechanisms of Resistance to Noncovalent Bruton's Tyrosine Kinase Inhibitors.
VCAM1 confers innate immune tolerance on haematopoietic and leukaemic stem cells.
Combining CRISPRi and metabolomics for functional annotation of compound libraries.
Gilteritinib monotherapy as a transplant bridging option for high risk FLT3-mutated AML with t(6;9)(p23;q34.1);DEK-NUP214 in morphological but not cytogenetic or molecular remission following standard induction chemotherapy.
A predictive scoring system for therapy-failure in persons with chronic myeloid leukemia receiving initial imatinib therapy.

Signal Transduct Target Ther. 2022 02 21. 7(1): 51

Activation of RAS/MAPK pathway confers MCL-1 mediated acquired resistance to BCL-2 inhibitor venetoclax in acute myeloid leukemia.

Qi Zhang, Bridget Riley-Gillis, Lina Han, Yanan Jia, Alessia Lodi, Haijiao Zhang, Saravanan Ganesan, Rongqing Pan, Sergej N Konoplev, Shannon R Sweeney, Jeremy A Ryan, Yulia Jitkova, Kenneth Dunner, Shaun E Grosskurth, Priyanka Vijay, Sujana Ghosh, Charles Lu, Wencai Ma, Stephen Kurtz, Vivian R Ruvolo, Helen Ma, Connie C Weng, Cassandra L Ramage, Natalia Baran, Ce Shi, Tianyu Cai, Richard Eric Davis, Venkata L Battula, Yingchang Mi, Jing Wang, Courtney D DiNardo, Michael Andreeff, Jeffery W Tyner, Aaron Schimmer, Anthony Letai, Rose Ann Padua, Carlos E Bueso-Ramos, Stefano Tiziani, Joel Leverson, Relja Popovic, Marina Konopleva.

Despite high initial response rates, acute myeloid leukemia (AML) treated with the BCL-2-selective inhibitor venetoclax (VEN) alone or in combinations commonly acquires resistance. We performed gene/protein expression, metabolomic and methylation analyses of isogenic AML cell lines sensitive or resistant to VEN, and identified the activation of RAS/MAPK pathway, leading to increased stability and higher levels of MCL-1 protein, as a major acquired mechanism of VEN resistance. MCL-1 sustained survival and maintained mitochondrial respiration in VEN-RE cells, which had impaired electron transport chain (ETC) complex II activity, and MCL-1 silencing or pharmacologic inhibition restored VEN sensitivity. In support of the importance of RAS/MAPK activation, we found by single-cell DNA sequencing rapid clonal selection of RAS-mutated clones in AML patients treated with VEN-containing regimens. In summary, these findings establish RAS/MAPK/MCL-1 and mitochondrial fitness as key survival mechanisms of VEN-RE AML and provide the rationale for combinatorial strategies effectively targeting these pathways.

DOI: https://doi.org/10.1038/s41392-021-00870-3
Haematologica. 2022 Feb 24.

Pseudo-mutant P53 is a unique phenotype of DNMT3A-mutated pre-leukemia.

Amos Tuval, Yardena Brilon, Hadas Azogy, Yoni Moskovitz, Dena Leshkowitz, Tomer M Salame, Mark D Minden, Perry Tal, Varda Rotter, Moshe Oren, Nathali Kaushansky, Liran I Shlush.

Pre-leukemic clones carrying DNMT3A mutations have a selective advantage and an inherent chemo-resistance, however the basis for this phenotype has not been fully elucidated. Mutations affecting the gene TP53 occur in pre-leukemic hematopoietic stem/progenitor cells (preL-HSPCs) and lead to chemo-resistance. Many of these mutations cause a conformational change and some of them were shown to enhance self-renewal capacity of preL-HSPCs. Intriguingly, a misfolded P53 was described in AML blasts that do not harbor mutations in TP53, emphasizing the dynamic equilibrium between wild-type (WT) and "pseudo-mutant" conformations of P53. By combining single cell analyses and P53 conformation-specific monoclonal antibodies we studied preL-HSPCs from primary human DNMT3A-mutated AML samples. We found that while leukemic blasts express mainly the WT conformation, in preL-HSPCs the pseudomutant conformation is the dominant. HSPCs from non-leukemic samples expressed both conformations to a similar extent. In a mouse model we found a small subset of HSPCs with a dominant pseudo-mutant P53. This subpopulation was significantly larger among DNMT3AR882H-mutated HSPCs, suggesting that while a pre-leukemic mutation can predispose for P53 misfolding, additional factors are involved as well. Treatment with a short peptide that can shift the dynamic equilibrium favoring the WT conformation of P53, specifically eliminated preL-HSPCs that had dysfunctional canonical P53 pathway activity as reflected by single cell RNA sequencing. Our observations shed light upon a possible targetable P53 dysfunction in human preLHSPCs carrying DNMT3A mutations. This opens new avenues for leukemia prevention.

DOI: https://doi.org/10.3324/haematol.2021.280329
Blood Adv. 2022 Feb 24. pii: bloodadvances.2021006689. [Epub ahead of print]

Validation of ALFA 1200 score in patients with AML >60 years treated with double nucleoside-based low-intensity therapy.

Sangeetha Venugopal, Gautam Borthakur, Naval G Daver, Courtney D DiNardo, Naveen Pemmaraju, Nicholas J Short, Hussein A Abbas, Guillermo Garcia-Manero, Marina Y Konopleva, Farhad Ravandi, Tapan M Kadia.

DOI: https://doi.org/10.1182/bloodadvances.2021006689
Biochem Biophys Res Commun. 2022 Feb 10. pii: S0006-291X(22)00205-4. [Epub ahead of print]600 29-34

Osteopontin is required for the maintenance of leukemia stem cells in acute myeloid leukemia.

Jing Zhou, Xing Chen, Pan Zhou, Xiaolu Sun, Yangpeng Chen, Mengke Li, Yajing Chu, Jianfeng Zhou, Xuelian Hu, Yi Luo, Weiping Yuan, Gaoxiang Wang.

  Acute myeloid leukemia (AML) is a heterogeneous hematopoietic disorder with a poor prognosis. The clinical significance of Leukemia stem cells (LSCs) plays an important role in the generation of AML and is the main cause of the recurrence after remission. Osteopontin (OPN), an extracellular matrix protein, has been implicated in hematopoietic malignancies. However, the specific role and the underlying mechanism of AML cell autocrined OPN in leukemia maintenance remain unknown. Here, we showed that knockdown of Opn expression significantly prolonged the survival of mice with MLL-AF9 cell-induced AML and markedly reduced the tumor burden. The LSCs from the Opn-knockdown groups exhibited decreased numbers and impaired function as determined by immunophenotype, colony-forming and limiting dilution assays. Further analysis revealed that Opn prevents LSCs from undergoing apoptosis and cell cycle arrest. Repression of OPN in human AML cell lines in vitro mimics the phenotypes observed in the mouse model. Overall, our data indicated that OPN is a potent therapeutic target for eradicating LSCs in AML.

Keywords:  Acute myeloid leukemia; Apoptosis; Leukemia stem cell; Osteopontin; Proliferation

DOI:  https://doi.org/10.1016/j.bbrc.2022.02.022
Proc Natl Acad Sci U S A. 2022 Mar 01. pii: e2122940119. [Epub ahead of print]119(9):

A PRC2-Kdm5b axis sustains tumorigenicity of acute myeloid leukemia.

Zhihong Ren, Arum Kim, Yu-Ting Huang, Wen-Chieh Pi, Weida Gong, Xufen Yu, Jun Qi, Jian Jin, Ling Cai, Robert G Roeder, Wei-Yi Chen, Gang Greg Wang.

  Acute myeloid leukemias (AMLs) with the NUP98-NSD1 or mixed lineage leukemia (MLL) rearrangement (MLL-r) share transcriptomic profiles associated with stemness-related gene signatures and display poor prognosis. The molecular underpinnings of AML aggressiveness and stemness remain far from clear. Studies with EZH2 enzymatic inhibitors show that polycomb repressive complex 2 (PRC2) is crucial for tumorigenicity in NUP98-NSD1+ AML, whereas transcriptomic analysis reveal that Kdm5b, a lysine demethylase gene carrying "bivalent" chromatin domains, is directly repressed by PRC2. While ectopic expression of Kdm5b suppressed AML growth, its depletion not only promoted tumorigenicity but also attenuated anti-AML effects of PRC2 inhibitors, demonstrating a PRC2-|Kdm5b axis for AML oncogenesis. Integrated RNA sequencing (RNA-seq), chromatin immunoprecipitation followed by sequencing (ChIP-seq), and Cleavage Under Targets & Release Using Nuclease (CUT&RUN) profiling also showed that Kdm5b directly binds and represses AML stemness genes. The anti-AML effect of Kdm5b relies on its chromatin association and/or scaffold functions rather than its demethylase activity. Collectively, this study describes a molecular axis that involves histone modifiers (PRC2-|Kdm5b) for sustaining AML oncogenesis.

Keywords:  AML; Kdm5b; PRC2; stemness; tumorigenicity

DOI:  https://doi.org/10.1073/pnas.2122940119
Blood Res. 2022 Feb 24.

IDH1/2 mutations in acute myeloid leukemia.

Ja Min Byun, Seung-Joo Yoo, Hyeong-Joon Kim, Jae-Sook Ahn, Youngil Koh, Jun Ho Jang, Sung-Soo Yoon.

  The mutational and epigenetic landscape of acute myeloid leukemia (AML) has become increasingly well understood in recent years, informing on biological targets for precision medicine. Among the most notable findings was the recognition of mutational hot-spots in the isocitrate dehydrogenase (IDH) genes. In this review, we provide an overview on the IDH1/2 mutation landscape in Korean AML patients, and compare it with available public data. We also discuss the role of IDH1/2 mutations as biomarkers and drug targets. Taken together, occurrence of IDH1/2 mutations is becoming increasingly important in AML treatment, thus requiring thorough examination and follow-up throughout the clinical course of the disease.

Keywords:  Acute myeloid leukemia; Biomarker; Drug target; IDH; Korean

DOI:  https://doi.org/10.5045/br.2021.2021152
Exp Mol Med. 2022 Feb 25.

The m6A reader IGF2BP3 promotes acute myeloid leukemia progression by enhancing RCC2 stability.

Nan Zhang, Yan Shen, Huan Li, Ying Chen, Ping Zhang, Shifeng Lou, Jianchuan Deng.

N6-methyladenosine (m6A) is the most abundant posttranscriptional modification of mRNA in eukaryotes. Recent evidence suggests that dysregulated m6A-associated proteins and m6A modifications play a pivotal role in the initiation and progression of diseases such as cancer. Here, we identified that IGF2BP3 is specifically overexpressed in acute myeloid leukemia (AML), a subtype of leukemia associated with poor prognosis and high genetic risk. IGF2BP3 is required for maintaining AML cell survival in an m6A-dependent manner, and knockdown of IGF2BP3 dramatically suppresses the apoptosis, reduces the proliferation, and impairs the leukemic capacity of AML cells in vitro and in vivo. Mechanistically, IGF2BP3 interacts with RCC2 mRNA and stabilizes the expression of m6A-modified RNA. Thus, we provided compelling evidence demonstrating that the m6A reader IGF2BP3 contributes to tumorigenesis and poor prognosis in AML and can serve as a target for the development of cancer therapeutics.

DOI: https://doi.org/10.1038/s12276-022-00735-x
Sci Transl Med. 2022 Feb 23. 14(633): eabm1375

Hematopoietic cell transplantation donor-derived memory-like NK cells functionally persist after transfer into patients with leukemia.

Melissa M Berrien-Elliott, Jennifer A Foltz, David A Russler-Germain, Carly C Neal, Jennifer Tran, Margery Gang, Pamela Wong, Bryan Fisk, Celia C Cubitt, Nancy D Marin, Alice Y Zhou, Miriam T Jacobs, Mark Foster, Timothy Schappe, Ethan McClain, Samantha Kersting-Schadek, Sweta Desai, Patrick Pence, Michelle Becker-Hapak, Jeremy Eisele, Matthew Mosior, Lynne Marsala, Obi L Griffith, Malachi Griffith, Saad M Khan, David H Spencer, John F DiPersio, Rizwan Romee, Geoffrey L Uy, Camille N Abboud, Armin Ghobadi, Peter Westervelt, Keith Stockerl-Goldstein, Mark A Schroeder, Fei Wan, Wen-Rong Lie, Patrick Soon-Shiong, Allegra A Petti, Amanda F Cashen, Todd A Fehniger.

Natural killer (NK) cells are innate lymphoid cells that eliminate cancer cells, produce cytokines, and are being investigated as a nascent cellular immunotherapy. Impaired NK cell function, expansion, and persistence remain key challenges for optimal clinical translation. One promising strategy to overcome these challenges is cytokine-induced memory-like (ML) differentiation, whereby NK cells acquire enhanced antitumor function after stimulation with interleukin-12 (IL-12), IL-15, and IL-18. Here, reduced-intensity conditioning (RIC) for HLA-haploidentical hematopoietic cell transplantation (HCT) was augmented with same-donor ML NK cells on day +7 and 3 weeks of N-803 (IL-15 superagonist) to treat patients with relapsed/refractory acute myeloid leukemia (AML) in a clinical trial (NCT02782546). In 15 patients, donor ML NK cells were well tolerated, and 87% of patients achieved a composite complete response at day +28, which corresponded with clearing high-risk mutations, including TP53 variants. NK cells were the major blood lymphocytes for 2 months after HCT with 1104-fold expansion (over 1 to 2 weeks). Phenotypic and transcriptional analyses identified donor ML NK cells as distinct from conventional NK cells and showed that ML NK cells persisted for over 2 months. ML NK cells expressed CD16, CD57, and high granzyme B and perforin, along with a unique transcription factor profile. ML NK cells differentiated in patients had enhanced ex vivo function compared to conventional NK cells from both patients and healthy donors. Overall, same-donor ML NK cell therapy with 3 weeks of N-803 support safely augmented RIC haplo-HCT for AML.

DOI: https://doi.org/10.1126/scitranslmed.abm1375
Blood Cancer J. 2022 Feb 24. 12(2): 31

Identification of novel STAT5B mutations and characterization of TCRβ signatures in CD4+ T-cell large granular lymphocyte leukemia.

Dipabarna Bhattacharya, Antonella Teramo, Vanessa Rebecca Gasparini, Jani Huuhtanen, Daehong Kim, Jason Theodoropoulos, Gianluca Schiavoni, Gregorio Barilà, Cristina Vicenzetto, Giulia Calabretto, Monica Facco, Toru Kawakami, Hideyuki Nakazawa, Brunangelo Falini, Enrico Tiacci, Fumihiro Ishida, Gianpietro Semenzato, Tiina Kelkka, Renato Zambello, Satu Mustjoki.

CD4+ T-cell large granular lymphocyte leukemia (T-LGLL) is a rare subtype of T-LGLL with unknown etiology. In this study, we molecularly characterized a cohort of patients (n = 35) by studying their T-cell receptor (TCR) repertoire and the presence of somatic STAT5B mutations. In addition to the previously described gain-of-function mutations (N642H, Y665F, Q706L, S715F), we discovered six novel STAT5B mutations (Q220H, E433K, T628S, P658R, P702A, and V712E). Multiple STAT5B mutations were present in 22% (5/23) of STAT5B mutated CD4+ T-LGLL cases, either coexisting in one clone or in distinct clones. Patients with STAT5B mutations had increased lymphocyte and LGL counts when compared to STAT5B wild-type patients. TCRβ sequencing showed that, in addition to large LGL expansions, non-leukemic T cell repertoires were more clonal in CD4+ T-LGLL compared to healthy. Interestingly, 25% (15/59) of CD4+ T-LGLL clonotypes were found, albeit in much lower frequencies, in the non-leukemic CD4+ T cell repertoires of the CD4+ T-LGLL patients. Additionally, we further confirmed the previously reported clonal dominance of TRBV6-expressing clones in CD4+ T-LGLL. In conclusion, CD4+ T-LGLL patients have a typical TCR and mutation profile suggestive of aberrant antigen response underlying the disease.

DOI: https://doi.org/10.1038/s41408-022-00630-8
Front Oncol. 2022 ;12 794021

What Clonal Hematopoiesis Can Teach Us About MDS.

Irenaeus C C Chan, Brian J Wiley, Kelly L Bolton.

  Clonal hematopoiesis (CH), defined as the clonal expansion of mutated hematopoietic stem and progenitor cells (HSPCs), is a common aging process. CH is a risk factor for the development of hematologic malignancies, most commonly myeloid neoplasms (MNs) including acute myeloid leukemia (AML), myelodysplastic syndrome (MDS), and myeloproliferative neoplasm (MPN). Recent work has elucidated how the development and cellular fitness of CH is shaped by aging, environmental exposures, and the germline (inherited) genetic background of an individual. This in turn has provided valuable insights into the pathogenesis of MNs including MDS. Here, in this review, we discuss the genetic origins of CH, the environmental stressors that influence CH, and the implications of CH on health outcomes including MDS. Since MNs have shared risk factors and underlying biology, most of our discussion regarding the implications of CH surrounds MN in general rather than focusing specifically on MDS. We conclude with future directions and areas of investigation including how intervention studies of CH might inform future therapeutic approaches to MN including MDS.

Keywords:  clonal hematopoiesis (CH); environmental risk; genetic predisposition; myelodysplastic syndrome (MDS); myeloid neoplasm

DOI:  https://doi.org/10.3389/fonc.2022.794021
Blood Adv. 2022 Feb 22. pii: bloodadvances.2021006783. [Epub ahead of print]

Lasting shift in the gut microbiota in patients with acute myeloid leukemia.

Armin Rashidi, Maryam Ebadi, Tauseef Ur Rehman, Heba Elhusseini, Hossam Fathi Halaweish, Thomas Kaiser, Shernan G Holtan, Alexander Khoruts, Daniel J Weisdorf, Christopher Staley.

Previous studies have shown that the gut microbiota of patients with acute myeloid leukemia (AML) is disrupted during induction chemotherapy. However, the durability of microbiota changes is unknown. This is an important knowledge gap because reduced microbiota diversity at the time of stem cell transplantation weeks to months after the initial chemotherapy has been associated with higher mortality after transplantation. By sequencing the gut microbiota in 410 longitudinal stool samples from 52 patients with AML, we find that during inpatient chemotherapy, the gut microbiota is stressed beyond its ability to recover its original state. Despite major reductions in antibiotic pressure and other disturbances to the microbiota after hospital discharge, the trajectory of microbiota recovery yields new communities that are highly dissimilar to baseline. This lasting shift in the gut microbiota is relevant for subsequent phases of curative therapy and is a potential target for novel microbiota protective/restorative interventions. This trial is registered at www.clinicaltrials.gov as # NCT03316456.

DOI: https://doi.org/10.1182/bloodadvances.2021006783
Cell Death Differ. 2022 Feb 22.

Deneddylation of PML/RARα reconstructs functional PML nuclear bodies via orchestrating phase separation to eradicate APL.

Xuejing Shao, Yingqian Chen, Aixiao Xu, Danyan Xiang, Wei Wang, Wenxin Du, Yunpeng Huang, Xingya Zhang, Minyi Cai, Zhimei Xia, Yi Wang, Ji Cao, Yan Zhang, Bo Yang, Qiaojun He, Meidan Ying.

Acute promyelocytic leukemia (APL) is driven by the oncoprotein PML/RARα, which destroys the architecture of PML nuclear bodies (NBs). PML NBs are critical to tumor suppression, and their disruption mediated by PML/RARα accelerates APL pathogenesis. However, the mechanisms of PML NB disruption remain elusive. Here, we reveal that the failure of NB assembly in APL results from neddylation-induced aberrant phase separation of PML/RARα. Mechanistically, PML/RARα is neddylated in the RARα moiety, and this neddylation enhances its DNA-binding ability and further impedes the phase separation of the PML moiety, consequently disrupting PML NB construction. Accordingly, deneddylation of PML/RARα restores its phase separation process to reconstruct functional NBs and activates RARα signaling, thereby suppressing PML/RARα-driven leukemogenesis. Pharmacological inhibition of neddylation by MLN4924 eradicates APL cells both in vitro and in vivo. Our work elucidates the neddylation-destroyed phase separation mechanism for PML/RARα-driven NB disruption and highlights targeting neddylation for APL eradication.

DOI: https://doi.org/10.1038/s41418-022-00955-8
Blood Adv. 2022 Feb 23. pii: bloodadvances.2021006090. [Epub ahead of print]

Changes in body mass index, weight, and height in children with acute myeloid leukemia and the associations with outcome.

Mayuko Iijima, Melanie Stall, Lei Wang, John Carl Panetta, Brandon M Triplett, Ching-Hon Pui, Raul C Ribeiro, Jeffrey E Rubnitz, Stanley B Pounds, Hiroto Inaba.

Little is known about body composition changes in patients with acute myeloid leukemia (AML) during and after treatment or their associations with outcomes. Z-scores for body mass index (BMI), weight, and height at diagnosis; their longitudinal changes from diagnosis to 5 years off-therapy; and their associations with adverse effects and outcomes were evaluated in 227 pediatric patients with AML enrolled in the AML02 and AML08 trials at St. Jude Children's Research Hospital between 2002 and 2017. The median Z-scores for baseline weight, height, and BMI were 0.193, 0.209, and 0.170, respectively, and those for weight and height decreased significantly during therapy to -0.038 and -0.163, respectively, at off-therapy (P < .001 for both). At 5 years off-therapy, the Z-scores for weight and BMI had increased significantly to 0.492 (P = .003) and 0.911 (P < .001), respectively, whereas the height Z-score remained significantly lower at -0.066 (P < .001) compared to baseline. The height Z-score of transplant recipients decreased further from -0.211 at transplant to -0.617 12 months later (P < .001). Baseline BMI category and Z-score were not associated with outcomes, but higher weight Z-scores were associated with lower incidences of refractory or relapsed disease (HR, 0.82; 95% confidence interval [CI], 0.67-0.99) and higher incidences of death in remission (HR, 1.31; 95% CI, 1.01-1.70). Furthermore, weight Z-score decrease during induction therapy was associated with gastrointestinal, hepatic, and infection toxicities during subsequent therapy and with death in remission (hazard ratio, 2.66; 95% CI, 1.11-6.45). Multidisciplinary monitoring for weight changes and short stature is required from diagnosis to the off-therapy period.

DOI: https://doi.org/10.1182/bloodadvances.2021006090
Nat Cell Biol. 2022 Feb 24.

EZH2 noncanonically binds cMyc and p300 through a cryptic transactivation domain to mediate gene activation and promote oncogenesis.

Jun Wang, Xufen Yu, Weida Gong, Xijuan Liu, Kwang-Su Park, Anqi Ma, Yi-Hsuan Tsai, Yudao Shen, Takashi Onikubo, Wen-Chieh Pi, David F Allison, Jing Liu, Wei-Yi Chen, Ling Cai, Robert G Roeder, Jian Jin, Gang Greg Wang.

Canonically, EZH2 serves as the catalytic subunit of PRC2, which mediates H3K27me3 deposition and transcriptional repression. Here, we report that in acute leukaemias, EZH2 has additional noncanonical functions by binding cMyc at non-PRC2 targets and uses a hidden transactivation domain (TAD) for (co)activator recruitment and gene activation. Both canonical (EZH2-PRC2) and noncanonical (EZH2-TAD-cMyc-coactivators) activities of EZH2 promote oncogenesis, which explains the slow and ineffective antitumour effect of inhibitors of the catalytic function of EZH2. To suppress the multifaceted activities of EZH2, we used proteolysis-targeting chimera (PROTAC) to develop a degrader, MS177, which achieved effective, on-target depletion of EZH2 and interacting partners (that is, both canonical EZH2-PRC2 and noncanonical EZH2-cMyc complexes). Compared with inhibitors of the enzymatic function of EZH2, MS177 is fast-acting and more potent in suppressing cancer growth. This study reveals noncanonical oncogenic roles of EZH2, reports a PROTAC for targeting the multifaceted tumorigenic functions of EZH2 and presents an attractive strategy for treating EZH2-dependent cancers.

DOI: https://doi.org/10.1038/s41556-022-00850-x
Biology (Basel). 2022 Jan 19. pii: 161. [Epub ahead of print]11(2):

Next-Generation Sequencing Revealed a Distinct Immunoglobulin Repertoire with Specific Mutation Hotspots in Acute Myeloid Leukemia.

Miaoran Xia, Lina Wu, Xiaoping Sun, Xin Han, Huige Yan, Jing Huang, Youhui Zhang, Zhihong Hu, Youli Zu, C Cameron Yin, Xiaoyan Qiu.

  Immunoglobulin (Ig) is known as a hallmark of B-lymphocytes exerting antibody functions. However, our previous studies demonstrated that myeloblasts from acute myeloid leukemia (AML) patients could also express Ig with distinct roles. Here, we quantified Ig (IGHG and IGK) transcripts by real-time PCR and performed a comprehensive analysis of Ig repertoire (both heavy chains and light chains) in AML blasts. We found that Ig was frequently expressed by AML blasts. A higher level of AML-derived IGHG expression correlated with a significantly shorter disease-free survival. Next-generation sequencing revealed dysregulated transcripts of all five Ig classes (IGHA, IGHD, IGHE, IGHG, and IGHM) and two Ig types (IGK and IGL) in AML. VH-D-JH rearrangements in myeloblasts were biased with individual specificity rather than generally diverse as in B-cells. Compared to AML-derived IgH, AML-derived IGK was more conserved among different AML samples. The frequently shared Vκ-Jκ patterns were IGKV3-20*01/IGKJ1*01, IGKV2D-28*01/IGKJ1*01, and IGKV4-1*01/IGKJ1*01. Moreover, AML-derived IGK was different from classical IGK in B-cells for the high mutation rates and special mutation hotspots at serine codons. Findings of the distinct Ig repertoire in myeloblasts may facilitate the discovery of a new molecular marker for disease monitoring and target therapy.

Keywords:  Ig; V(D)J rearrangement; acute myeloid leukemia; next-generation sequencing; somatic hypermutation

DOI:  https://doi.org/10.3390/biology11020161
BMC Cancer. 2022 Feb 22. 22(1): 201

Deep learning identifies Acute Promyelocytic Leukemia in bone marrow smears.

Jan-Niklas Eckardt, Tim Schmittmann, Sebastian Riechert, Michael Kramer, Anas Shekh Sulaiman, Katja Sockel, Frank Kroschinsky, Johannes Schetelig, Lisa Wagenführ, Ulrich Schuler, Uwe Platzbecker, Christian Thiede, Friedrich Stölzel, Christoph Röllig, Martin Bornhäuser, Karsten Wendt, Jan Moritz Middeke.

  BACKGROUND: Acute promyelocytic leukemia (APL) is considered a hematologic emergency due to high risk of bleeding and fatal hemorrhages being a major cause of death. Despite lower death rates reported from clinical trials, patient registry data suggest an early death rate of 20%, especially for elderly and frail patients. Therefore, reliable diagnosis is required as treatment with differentiation-inducing agents leads to cure in the majority of patients. However, diagnosis commonly relies on cytomorphology and genetic confirmation of the pathognomonic t(15;17). Yet, the latter is more time consuming and in some regions unavailable.METHODS: In recent years, deep learning (DL) has been evaluated for medical image recognition showing outstanding capabilities in analyzing large amounts of image data and provides reliable classification results. We developed a multi-stage DL platform that automatically reads images of bone marrow smears, accurately segments cells, and subsequently predicts APL using image data only. We retrospectively identified 51 APL patients from previous multicenter trials and compared them to 1048 non-APL acute myeloid leukemia (AML) patients and 236 healthy bone marrow donor samples, respectively.
RESULTS: Our DL platform segments bone marrow cells with a mean average precision and a mean average recall of both 0.97. Further, it achieves high accuracy in detecting APL by distinguishing between APL and non-APL AML as well as APL and healthy donors with an area under the receiver operating characteristic of 0.8575 and 0.9585, respectively, using visual image data only.
CONCLUSIONS: Our study underlines not only the feasibility of DL to detect distinct morphologies that accompany a cytogenetic aberration like t(15;17) in APL, but also shows the capability of DL to abstract information from a small medical data set, i. e. 51 APL patients, and infer correct predictions. This demonstrates the suitability of DL to assist in the diagnosis of rare cancer entities. As our DL platform predicts APL from bone marrow smear images alone, this may be used to diagnose APL in regions were molecular or cytogenetic subtyping is not routinely available and raise attention to suspected cases of APL for expert evaluation.

Keywords:  Acute myeloid leukemia; Acute promyelocytic leukemia; Artificial intelligence; Bone marrow smear; Deep learning

DOI:  https://doi.org/10.1186/s12885-022-09307-8
Nat Commun. 2022 Feb 25. 13(1): 1048

Phosphoproteomic profiling of T cell acute lymphoblastic leukemia reveals targetable kinases and combination treatment strategies.

Valentina Cordo', Mariska T Meijer, Rico Hagelaar, Richard R de Goeij-de Haas, Vera M Poort, Alex A Henneman, Sander R Piersma, Thang V Pham, Koichi Oshima, Adolfo A Ferrando, Guido J R Zaman, Connie R Jimenez, Jules P P Meijerink.

Protein kinase inhibitors are amongst the most successful cancer treatments, but targetable kinases activated by genomic abnormalities are rare in T cell acute lymphoblastic leukemia. Nevertheless, kinases can be activated in the absence of genetic defects. Thus, phosphoproteomics can provide information on pathway activation and signaling networks that offer opportunities for targeted therapy. Here, we describe a mass spectrometry-based global phosphoproteomic profiling of 11 T cell acute lymphoblastic leukemia cell lines to identify targetable kinases. We report a comprehensive dataset consisting of 21,000 phosphosites on 4,896 phosphoproteins, including 217 kinases. We identify active Src-family kinases signaling as well as active cyclin-dependent kinases. We validate putative targets for therapy ex vivo and identify potential combination treatments, such as the inhibition of the INSR/IGF-1R axis to increase the sensitivity to dasatinib treatment. Ex vivo validation of selected drug combinations using patient-derived xenografts provides a proof-of-concept for phosphoproteomics-guided design of personalized treatments.

DOI: https://doi.org/10.1038/s41467-022-28682-1
N Engl J Med. 2022 02 24. 386(8): 735-743

Mechanisms of Resistance to Noncovalent Bruton's Tyrosine Kinase Inhibitors.

Eric Wang, Xiaoli Mi, Meghan C Thompson, Skye Montoya, Ryan Q Notti, Jumana Afaghani, Benjamin H Durham, Alex Penson, Matthew T Witkowski, Sydney X Lu, Jessie Bourcier, Simon J Hogg, Caroline Erickson, Dan Cui, Hana Cho, Michael Singer, Tulasigeri M Totiger, Sana Chaudhry, Mark Geyer, Alvaro Alencar, Adam J Linley, M Lia Palomba, Catherine C Coombs, Jae H Park, Andrew Zelenetz, Lindsey Roeker, Mary Rosendahl, Donald E Tsai, Kevin Ebata, Barbara Brandhuber, David M Hyman, Iannis Aifantis, Anthony Mato, Justin Taylor, Omar Abdel-Wahab.

BACKGROUND: Covalent (irreversible) Bruton's tyrosine kinase (BTK) inhibitors have transformed the treatment of multiple B-cell cancers, especially chronic lymphocytic leukemia (CLL). However, resistance can arise through multiple mechanisms, including acquired mutations in BTK at residue C481, the binding site of covalent BTK inhibitors. Noncovalent (reversible) BTK inhibitors overcome this mechanism and other sources of resistance, but the mechanisms of resistance to these therapies are currently not well understood.METHODS: We performed genomic analyses of pretreatment specimens as well as specimens obtained at the time of disease progression from patients with CLL who had been treated with the noncovalent BTK inhibitor pirtobrutinib. Structural modeling, BTK-binding assays, and cell-based assays were conducted to study mutations that confer resistance to noncovalent BTK inhibitors.
RESULTS: Among 55 treated patients, we identified 9 patients with relapsed or refractory CLL and acquired mechanisms of genetic resistance to pirtobrutinib. We found mutations (V416L, A428D, M437R, T474I, and L528W) that were clustered in the kinase domain of BTK and that conferred resistance to both noncovalent BTK inhibitors and certain covalent BTK inhibitors. Mutations in BTK or phospholipase C gamma 2 (PLCγ2), a signaling molecule and downstream substrate of BTK, were found in all 9 patients. Transcriptional activation reflecting B-cell-receptor signaling persisted despite continued therapy with noncovalent BTK inhibitors.
CONCLUSIONS: Resistance to noncovalent BTK inhibitors arose through on-target BTK mutations and downstream PLCγ2 mutations that allowed escape from BTK inhibition. A proportion of these mutations also conferred resistance across clinically approved covalent BTK inhibitors. These data suggested new mechanisms of genomic escape from established covalent and novel noncovalent BTK inhibitors. (Funded by the American Society of Hematology and others.).

DOI: https://doi.org/10.1056/NEJMoa2114110
Nat Cell Biol. 2022 Feb 24.

VCAM1 confers innate immune tolerance on haematopoietic and leukaemic stem cells.

Sandra Pinho, Qiaozhi Wei, Maria Maryanovich, Dachuan Zhang, Juan Carlos Balandrán, Halley Pierce, Fumio Nakahara, Anna Di Staulo, Boris A Bartholdy, Jianing Xu, Daniel K Borger, Amit Verma, Paul S Frenette.

Haematopoietic stem cells (HSCs) home to the bone marrow via, in part, interactions with vascular cell adhesion molecule-1 (VCAM1)1-3. Once in the bone marrow, HSCs are vetted by perivascular phagocytes to ensure their self-integrity. Here we show that VCAM1 is also expressed on healthy HSCs and upregulated on leukaemic stem cells (LSCs), where it serves as a quality-control checkpoint for entry into bone marrow by providing 'don't-eat-me' stamping in the context of major histocompatibility complex class-I (MHC-I) presentation. Although haplotype-mismatched HSCs can engraft, Vcam1 deletion, in the setting of haplotype mismatch, leads to impaired haematopoietic recovery due to HSC clearance by mononuclear phagocytes. Mechanistically, VCAM1 'don't-eat-me' activity is regulated by β2-microglobulin MHC presentation on HSCs and paired Ig-like receptor-B (PIR-B) on phagocytes. VCAM1 is also used by cancer cells to escape immune detection as its expression is upregulated in multiple cancers, including acute myeloid leukaemia (AML), where high expression associates with poor prognosis. In AML, VCAM1 promotes disease progression, whereas VCAM1 inhibition or deletion reduces leukaemia burden and extends survival. These results suggest that VCAM1 engagement regulates a critical immune-checkpoint gate in the bone marrow, and offers an alternative strategy to eliminate cancer cells via modulation of the innate immune tolerance.

DOI: https://doi.org/10.1038/s41556-022-00849-4
Nat Chem Biol. 2022 Feb 22.

Combining CRISPRi and metabolomics for functional annotation of compound libraries.

Miquel Anglada-Girotto, Gabriel Handschin, Karin Ortmayr, Adrian I Campos, Ludovic Gillet, Pablo Manfredi, Claire V Mulholland, Michael Berney, Urs Jenal, Paola Picotti, Mattia Zampieri.

Molecular profiling of small molecules offers invaluable insights into the function of compounds and allows for hypothesis generation about small-molecule direct targets and secondary effects. However, current profiling methods are limited in either the number of measurable parameters or throughput. Here we developed a multiplexed, unbiased framework that, by linking genetic to drug-induced changes in nearly a thousand metabolites, allows for high-throughput functional annotation of compound libraries in Escherichia coli. First, we generated a reference map of metabolic changes from CRISPR interference (CRISPRi) with 352 genes in all major essential biological processes. Next, on the basis of the comparison of genetic changes with 1,342 drug-induced metabolic changes, we made de novo predictions of compound functionality and revealed antibacterials with unconventional modes of action (MoAs). We show that our framework, combining dynamic gene silencing with metabolomics, can be adapted as a general strategy for comprehensive high-throughput analysis of compound functionality from bacteria to human cell lines.

DOI: https://doi.org/10.1038/s41589-022-00970-3
Leuk Res Rep. 2022 ;17 100291

Gilteritinib monotherapy as a transplant bridging option for high risk FLT3-mutated AML with t(6;9)(p23;q34.1);DEK-NUP214 in morphological but not cytogenetic or molecular remission following standard induction chemotherapy.

James W Day, Thomas A Fox, Rajeev Gupta, Asim Khwaja, Andrew J Wilson, Panagiotis D Kottaridis.

We report a case of FLT3-mutated AML with t(6;9) in which induction chemotherapy with DA and midostaurin failed to achieve complete cytogenetic or molecular remission. Due to the COVID-19 pandemic and co-existing cellulitis, monotherapy with the selective FLT3-inhibitor gilteritinib was used as an alternative consolidation treatment option rather than further intensive chemotherapy. Gilteritinib was able to achieve complete molecular and cytogenetic remission despite the additional cytogenetic abnormality. This case provides supporting evidence for the use of single agent gilteritinib in high-risk primary refractory FLT3-mutated AML with t(6;9) prior to transplantation.

DOI: https://doi.org/10.1016/j.lrr.2022.100291
Leukemia. 2022 Feb 22.

A predictive scoring system for therapy-failure in persons with chronic myeloid leukemia receiving initial imatinib therapy.

Xiao-Shuai Zhang, Robert Peter Gale, Mei-Jie Zhang, Xiao-Jun Huang, Qian Jiang.

Data from 1,364 consecutive subjects with chronic-phase chronic myeloid leukemia (CML) receiving initial imatinib-therapy were interrogated to identify co-variates predicting therapy failure. Subjects were randomly divided into training (n = 908) and validation datasets (n = 456). In the training dataset, WBC count ≥120 × 10E + 9/L, haemoglobin concentration <115 g/L, blood basophils ≥12% and European Treatment and Outcome Study for CML Long-Term Survival (ELTS) risk score were significantly-associated with failure-free survival (FFS). Each co-variate was assigned 1 point to develop the imatinib-therapy failure (IMTF) model except ELTS high-risk category which was assigned 2 points based on multi-variable regression coefficients. Area under receiver-operator characteristic curve values in the IMTF model for 1-, 3- and 5-year FFS were 0.79-0.84 in the training dataset and 0.78-0.85 in the validation dataset. Calibration plots showed high agreement between predicted and observed outcomes. Decision curve analyses indicated subjects benefited from clinical use of this model. Cumulative incidences of imatinib-therapy failure and probabilities of FFS among the 5 risk cohorts (very low-, low-, intermediate-, high- and very high-risk) using the IMTF model were significantly different (all p values < 0.001). The IMTF model also correlated with probabilities of progression-free survival and survival (all p values < 0.001). These data should help physicians optimize TKI-therapy strategy at diagnosis in persons with chronic phase CML.

DOI: https://doi.org/10.1038/s41375-022-01527-y