Mutant U2AF1-Induced Mis-Splicing of mRNA Translation Genes Confers Resistance to Chemotherapy in Acute Myeloid Leukemia.

Patients with primary refractory acute myeloid leukemia (AML) have a dismal long-term prognosis. Elucidating the resistance mechanisms to induction chemotherapy could help identify strategies to improve AML patient outcomes. Herein, we retrospectively analyzed the multiomics data of more than 1,500 AML cases and found that patients with spliceosome mutations had a higher risk of developing refractory disease. RNA splicing analysis revealed that the mis-spliced genes in refractory patients converged on translation-associated pathways, promoted mainly by U2AF1 mutations. Integrative analyses of binding and splicing in AML cell lines substantiated that the splicing perturbations of mRNA translation genes originated from both the loss and gain of mutant U2AF1 binding. In particular, the U2AF1S34F and U2AF1Q157R mutants orchestrated the inclusion of exon 11 (encoding a premature termination codon) in the eukaryotic translation initiation factor 4A2 (EIF4A2). This aberrant inclusion led to reduced eIF4A2 protein expression via nonsense-mediated mRNA decay. Consequently, U2AF1 mutations caused a net decrease in global mRNA translation that induced the integrated stress response (ISR) in AML cells, which was confirmed by single-cell RNA sequencing. The induction of ISR enhanced the ability of AML cells to respond and adapt to stress, contributing to chemoresistance. A pharmacologic inhibitor of ISR, ISRIB, sensitized U2AF1 mutant cells to chemotherapy. These findings highlight a resistance mechanism by which U2AF1 mutations drive chemoresistance and provide a therapeutic approach for AML through targeting the ISR pathway. SIGNIFICANCE: U2AF1 mutations induce the integrated stress response by disrupting splicing of mRNA translation genes that improves AML cell fitness to enable resistance to chemotherapy, which can be targeted to improve AML treatment.

TIGIT axis: novel immune checkpoints in anti-leukemia immunity.

Hematologic malignancy evades immune-mediated recognition through upregulating various checkpoint inhibitory receptors (IRs) on several types of lymphocytes. Immunotherapies targeting IRs have provided ample evidence supporting regulating innate and adaptive immunity and obtaining clinical benefits. Newly described IRs have received considerable attention and are under investigation in cancer immunotherapy. Specifically, T cell immunoglobulin and ITIM domain is a novel inhibitory checkpoint receptor, and its immune checkpoint axis includes additional receptors such as CD96 and CD226, which are very promising targets. However, how the dynamics and functions of these receptor networks remain unknown, this review addresses the recent findings of the relevance of this complex receptor-ligand system and discusses their potential approaches in translating these preclinical findings into novel clinical agents in anti-leukemia immunotherapy.

High TRGV 9 Subfamily Expression Marks an Improved Overall Survival in Patients With Acute Myeloid Leukemia.

Background: Heterogeneous T cells in acute myeloid leukemia (AML) have the combinatorial variety generated by different T cell receptors (TCRs). γδ T cells are a distinct subgroup of T cells containing TCRγ (TRGV) and TCRδ (TRDV) subfamilies with diverse structural and functional heterogeneity. Our previous study showed that clonally expanded TRDV T cells might benefit the immune response directed against AML. However, the features of the TRGV repertoire in AML remain unknown. To fully characterize the features of γδ T cells, we analyzed the distribution and clonality of TRGV I-III subfamilies (TRGV II is also termed TRVG 9), the proportions of γδ T cell subsets, and their effects on the overall survival (OS) of patients with AML. Methods: In this study, the complementarity-determining region 3 (CDR3) size of TRGV subfamilies in γδ T cells of peripheral blood (PB) from de novo AML patients were analyzed by Genescan analysis. Expression levels of TRGV subfamilies were performed by real-time quantitative PCR. The proportions of total γδ T cells and their Vγ9+ Vδ2+ T cells subsets were detected by multicolor flow cytometry assay. We further compared the correlation among the TRGV gene expression levels, the proportion of Vγ9+ Vδ2+ T cells, and OS in AML. Results: We first found that the distribution pattern and clonality of TRGV subfamilies were changed. The expression frequencies and gene expression levels of three TRGV subfamilies in AML samples were significantly lower than those in healthy individuals (HIs). Compared with HIs, the proportions of total γδ T cells and Vγ9+ Vδ2+ T cells were also significantly decreased in patients with AML. In addition, patients with AML who had higher expression levels of the TRGV gene and higher proportion of Vγ9+ Vδ2+ T cells showed better OS than their counterparts. Furthermore, high expression levels of TRGV 9 and proportion of Vγ9+ Vδ2+ T cells were identified as independent protective factors for complete remission in patients with AML. Conclusions: The restriction of TRGV usage might be related to the preference of usage of γδ T cells. Higher expression of TRGV subfamilies might be associated with better OS in AML. Higher TRGV 9 expression and increased Vγ9+ Vδ2+ T cells subfamilies might indicate a better prognosis in patients with AML.

Characteristic of TIGIT and DNAM-1 Expression on Foxp3+ γδ T Cells in AML Patients.

Foxp3+ γδ regulatory T (γδ Treg) cells promote tumor growth by various mechanisms and induce immuno-senescence. The novel immune checkpoint coinhibitory receptor T cell Ig and ITIM domain (TIGIT) shares similar ligands as the costimulatory receptor DNAX accessory molecule 1 (DNAM-1) and suppresses T cell responses in tumor patients. This study is aimed at characterizing whether the TIGIT/DNAM-1 axis is involved in the distribution and expression of Foxp3+ γδ Treg cell subsets in acute myeloid leukemia (AML) patients of different clinical statuses: de novo AML (27 patients), AML in nonremission (NR) (7 patients), and AML in complete remission (CR) (12 patients). Our data demonstrated that the proportions of Foxp3+, TIGIT+Foxp3+, and DNAM-1+Foxp3+ γδ T cells are significantly higher in de novo and NR patients. High levels of TIGIT and DNAM-1 on Foxp3+ γδ T cells correlated with increased Foxp3+ γδ T cell frequencies. In addition, a high TIGIT/DNAM-1 ratio was observed in de novo AML patients and healthy individuals (HIs). Furthermore, the phenotypic abnormalities in Foxp3+, TIGIT+Foxp3+, and DNAM-1+Foxp3+ γδ T cells were restored when the patients achieved CR after chemotherapy. Moreover, higher TIGIT+Foxp3+ γδ T cells were associated with AML patients who had poor overall survival and were an independent risk factor for prognosis. In conclusion, our study reveals for the first time that the TIGIT/DNAM-1 axis may be involved in Foxp3+ γδ Treg cells and indicates the clinical progression and prognosis of AML patients of different clinical statuses, which is considered beneficial for efficient AML immunotherapy.

Tonsillectomy discharge information - An improvement in both patient safety and satisfaction.

Approximately 20,000 adult and 25,000 paediatric tonsillectomies are performed each year in England. 0.9% of these patients return to theatre for post-tonsillectomy bleeding. The Royal College of Surgeons of England (RCSEng) have produced guidelines regarding emergency surgery, with standards for tonsillectomy discharge information. We audited our compliance with these guidelines and patient satisfaction regarding the information currently provided. Theatre records identified all tonsillectomies carried out between December 2012 and February 2013. 71 patients and their electronic discharge information were reviewed for post-operative bleeding information. Each patient was contacted, with a second call made to those who did not answer. 35 patients took part in our telephone questionnaire. Only 35% of patients had post-operative bleeding information on their discharge summary. 51% received no written information either in clinic or on the day of surgery, 66% recalled a verbal explanation. Only 54% knew to go to A&E if they experienced bleeding. 40% were not satisfied with their discharge information, stating that they wanted to know about bleeding, recovery expectations, and information regarding oral intake. A focus group was formed to discuss potential solutions to the audit outcomes and a tonsillectomy leaflet was produced inline with the trust patient information template. It contained specific instructions regarding bleeding and the nearest A&E contact details. It was reviewed by three tonsillectomy patients and their feedback regarding further information on post-operative diet, throat appearance and pain expectations was incorporated. A second cycle of the audit took place between August and September 2013. Results showed improvement, with 83% receiving an information leaflet and 100% a verbal explanation. 100% of patients were satisfied with their discharge information and 100% knew what to do if they bled. As a result we now meet the standards set out by the RCSEng and have increased our patient safety and satisfaction rates.