The Lineage-Defining Transcription Factors SOX2 and NKX2-1 Determine Lung Cancer Cell Fate and Shape the Tumor Immune Microenvironment.

The major types of non-small-cell lung cancer (NSCLC)-squamous cell carcinoma and adenocarcinoma-have distinct immune microenvironments. We developed a genetic model of squamous NSCLC on the basis of overexpression of the transcription factor Sox2, which specifies lung basal cell fate, and loss of the tumor suppressor Lkb1 (SL mice). SL tumors recapitulated gene-expression and immune-infiltrate features of human squamous NSCLC; such features included enrichment of tumor-associated neutrophils (TANs) and decreased expression of NKX2-1, a transcriptional regulator that specifies alveolar cell fate. In Kras-driven adenocarcinomas, mis-expression of Sox2 or loss of Nkx2-1 led to TAN recruitment. TAN recruitment involved SOX2-mediated production of the chemokine CXCL5. Deletion of Nkx2-1 in SL mice (SNL) revealed that NKX2-1 suppresses SOX2-driven squamous tumorigenesis by repressing adeno-to-squamous transdifferentiation. Depletion of TANs in SNL mice reduced squamous tumors, suggesting that TANs foster squamous cell fate. Thus, lineage-defining transcription factors determine the tumor immune microenvironment, which in turn might impact the nature of the tumor.

The origin and evolution of mutations in acute myeloid leukemia.

Most mutations in cancer genomes are thought to be acquired after the initiating event, which may cause genomic instability and drive clonal evolution. However, for acute myeloid leukemia (AML), normal karyotypes are common, and genomic instability is unusual. To better understand clonal evolution in AML, we sequenced the genomes of M3-AML samples with a known initiating event (PML-RARA) versus the genomes of normal karyotype M1-AML samples and the exomes of hematopoietic stem/progenitor cells (HSPCs) from healthy people. Collectively, the data suggest that most of the mutations found in AML genomes are actually random events that occurred in HSPCs before they acquired the initiating mutation; the mutational history of that cell is "captured" as the clone expands. In many cases, only one or two additional, cooperating mutations are needed to generate the malignant founding clone. Cells from the founding clone can acquire additional cooperating mutations, yielding subclones that can contribute to disease progression and/or relapse.

Acute myeloid leukemias with UBTF tandem duplications are sensitive to menin inhibitors.

ABSTRACT: UBTF tandem duplications (UBTF-TDs) have recently emerged as a recurrent alteration in pediatric and adult acute myeloid leukemia (AML). UBTF-TD leukemias are characterized by a poor response to conventional chemotherapy and a transcriptional signature that mirrors NUP98-rearranged and NPM1-mutant AMLs, including HOX-gene dysregulation. However, the mechanism by which UBTF-TD drives leukemogenesis remains unknown. In this study, we investigated the genomic occupancy of UBTF-TD in transformed cord blood CD34+ cells and patient-derived xenograft models. We found that UBTF-TD protein maintained genomic occupancy at ribosomal DNA loci while also occupying genomic targets commonly dysregulated in UBTF-TD myeloid malignancies, such as the HOXA/HOXB gene clusters and MEIS1. These data suggest that UBTF-TD is a gain-of-function alteration that results in mislocalization to genomic loci dysregulated in UBTF-TD leukemias. UBTF-TD also co-occupies key genomic loci with KMT2A and menin, which are known to be key partners involved in HOX-dysregulated leukemias. Using a protein degradation system, we showed that stemness, proliferation, and transcriptional signatures are dependent on sustained UBTF-TD localization to chromatin. Finally, we demonstrate that primary cells from UBTF-TD leukemias are sensitive to the menin inhibitor SNDX-5613, resulting in markedly reduced in vitro and in vivo tumor growth, myeloid differentiation, and abrogation of the UBTF-TD leukemic expression signature. These findings provide a viable therapeutic strategy for patients with this high-risk AML subtype.

Activation of human STING by a molecular glue-like compound.

Stimulator of interferon genes (STING) is a dimeric transmembrane adapter protein that plays a key role in the human innate immune response to infection and has been therapeutically exploited for its antitumor activity. The activation of STING requires its high-order oligomerization, which could be induced by binding of the endogenous ligand, cGAMP, to the cytosolic ligand-binding domain. Here we report the discovery through functional screens of a class of compounds, named NVS-STGs, that activate human STING. Our cryo-EM structures show that NVS-STG2 induces the high-order oligomerization of human STING by binding to a pocket between the transmembrane domains of the neighboring STING dimers, effectively acting as a molecular glue. Our functional assays showed that NVS-STG2 could elicit potent STING-mediated immune responses in cells and antitumor activities in animal models.

TXNIP loss expands Myc-dependent transcriptional programs by increasing Myc genomic binding.

The c-Myc protooncogene places a demand on glucose uptake to drive glucose-dependent biosynthetic pathways. To meet this demand, c-Myc protein (Myc henceforth) drives the expression of glucose transporters, glycolytic enzymes, and represses the expression of thioredoxin interacting protein (TXNIP), which is a potent negative regulator of glucose uptake. A Mychigh/TXNIPlow gene signature is clinically significant as it correlates with poor clinical prognosis in triple-negative breast cancer (TNBC) but not in other subtypes of breast cancer, suggesting a functional relationship between Myc and TXNIP. To better understand how TXNIP contributes to the aggressive behavior of TNBC, we generated TXNIP null MDA-MB-231 (231:TKO) cells for our study. We show that TXNIP loss drives a transcriptional program that resembles those driven by Myc and increases global Myc genome occupancy. TXNIP loss allows Myc to invade the promoters and enhancers of target genes that are potentially relevant to cell transformation. Together, these findings suggest that TXNIP is a broad repressor of Myc genomic binding. The increase in Myc genomic binding in the 231:TKO cells expands the Myc-dependent transcriptome we identified in parental MDA-MB-231 cells. This expansion of Myc-dependent transcription following TXNIP loss occurs without an apparent increase in Myc's intrinsic capacity to activate transcription and without increasing Myc levels. Together, our findings suggest that TXNIP loss mimics Myc overexpression, connecting Myc genomic binding and transcriptional programs to the nutrient and progrowth signals that control TXNIP expression.

tRNA shape is an identity element for an archaeal pyrrolysyl-tRNA synthetase from the human gut.

Protein translation is orchestrated through tRNA aminoacylation and ribosomal elongation. Among the highly conserved structure of tRNAs, they have distinguishing features which promote interaction with their cognate aminoacyl tRNA synthetase (aaRS). These key features are referred to as identity elements. In our study, we investigated the tRNA:aaRS pair that installs the 22nd amino acid, pyrrolysine (tRNAPyl:PylRS). Pyrrolysyl-tRNA synthetases (PylRSs) are naturally encoded in some archaeal and bacterial genomes to acylate tRNAPyl with pyrrolysine. Their large amino acid binding pocket and poor recognition of the tRNA anticodon have been instrumental in incorporating >200 noncanonical amino acids. PylRS enzymes can be divided into three classes based on their genomic structure. Two classes contain both an N-terminal and C-terminal domain, however the third class (ΔpylSn) lacks the N-terminal domain. In this study we explored the tRNA identity elements for a ΔpylSn tRNAPyl from Candidatus Methanomethylophilus alvus which drives the orthogonality seen with its cognate PylRS (MaPylRS). From aminoacylation and translation assays we identified five key elements in ΔpylSn tRNAPyl necessary for MaPylRS activity. The absence of a base (position 8) and a G-U wobble pair (G28:U42) were found to affect the high-resolution structure of the tRNA, while molecular dynamic simulations led us to acknowledge the rigidity imparted from the G-C base pairs (G3:C70 and G5:C68).

Enzymes known as PylRS offer the remarkable ability to expand the natural genetic code of a living cell with unnatural amino acids. Currently, over 200 unnatural amino acids can be genetically encoded with the help of PylRS and its partner tRNAPyl, enabling us to endow proteins with novel properties, or regulate protein activity using light or inducible cross-linking. One intriguing feature of PylRS enzymes is their ability to avoid cross-reactivity when two PylRS homologs from different organisms-such as those from the archaea Methanosarcina mazei and Methanomethylophilus alvus-are co-expressed in a single cell. This makes it possible to simultaneously encode two unnatural amino acids in a single protein. This study illuminates the elusive mechanism of PylRS specificity by using cryo-electron microscopy, biochemistry and molecular simulations. The interaction of PylRS from M. alvus with its tRNAPyl is best described as two pieces of a jigsaw puzzle; in which PylRS recognizes the unique shape of its cognate tRNA instead of specific nucleotides in the tRNA sequence like other tRNA-binding enzymes. This finding may streamline the rational design of tools for simultaneous genetic incorporation of multiple unnatural amino acids, thereby facilitating the development of valuable proteins for research, medicine, and biotechnology.

Split aminoacyl-tRNA synthetases for proximity-induced stop codon suppression.

Synthetic biology tools for regulating gene expression have many useful biotechnology and therapeutic applications. Most tools developed for this purpose control gene expression at the level of transcription, and relatively few methods are available for regulating gene expression at the translational level. Here, we design and engineer split orthogonal aminoacyl-tRNA synthetases (o-aaRS) as unique tools to control gene translation in bacteria and mammalian cells. Using chemically induced dimerization domains, we developed split o-aaRSs that mediate gene expression by conditionally suppressing stop codons in the presence of the small molecules rapamycin and abscisic acid. By activating o-aaRSs, these molecular switches induce stop codon suppression, and in their absence stop codon suppression is turned off. We demonstrate, in Escherichia coli and in human cells, that split o-aaRSs function as genetically encoded AND gates where stop codon suppression is controlled by two distinct molecular inputs. In addition, we show that split o-aaRSs can be used as versatile biosensors to detect therapeutically relevant protein-protein interactions, including those involved in cancer, and those that mediate severe acute respiratory syndrome-coronavirus-2 infection.

Proactive M2 blockade prevents cognitive decline in GRK5-deficient APP transgenic mice via enhancing cholinergic neuronal resilience.

Alzheimer's disease (AD) poses an immense challenge in healthcare, lacking effective therapies. This study investigates the potential of anthranilamide derivative (AAD23), a selective M2 receptor antagonist, in proactively preventing cognitive impairments and cholinergic neuronal degeneration in G protein-coupled receptor kinase-5-deficient Swedish APP (GAP) mice. GAP mice manifest cognitive deficits by 7 months and develop senile plaques by 9 months. A 6-month AAD23 treatment was initiated at 5 months and stopped at 11 months before behavioral assessments without the treatment. AAD23-treated mice exhibited preserved cognitive abilities and improved cholinergic axonal health in the nucleus basalis of Meynert akin to wildtype mice. Conversely, vehicle-treated GAP mice displayed memory deficits and pronounced cholinergic axonal swellings in the nucleus basalis of Meynert. Notably, AAD23 treatment did not alter senile plaques and microgliosis. These findings highlight AAD23's efficacy in forestalling AD-related cognitive decline in G protein-coupled receptor kinase-5-deficient subjects, attributing its success to restoring cholinergic neuronal integrity and resilience, enhancing resistance against diverse degenerative insults.

OX40/OX40 ligand and its role in precision immune oncology.

Immune checkpoint inhibitors have changed the treatment landscape for various malignancies; however, their benefit is limited to a subset of patients. The immune machinery includes both mediators of suppression/immune evasion, such as PD-1, PD-L1, CTLA-4, and LAG-3, all of which can be inhibited by specific antibodies, and immune-stimulatory molecules, such as T-cell co-stimulatory receptors that belong to the tumor necrosis factor receptor superfamily (TNFRSF), including OX40 receptor (CD134; TNFRSF4), 4-1BB (CD137; TNFRSF9), and glucocorticoid-induced TNFR-related (GITR) protein (CD357; TNFRSF18). In particular, OX40 and its binding ligand OX40L (CD134L; TNFSF4; CD252) are critical for immunoregulation. When OX40 on activated T cells binds OX40L on antigen-presenting cells, T-cell activation and immune stimulation are initiated via enhanced T-cell survival, proliferation and cytotoxicity, memory T-cell formation, and abrogation of regulatory T cell (Treg) immunosuppressive functions. OX40 agonists are in clinical trials both as monotherapy and in combination with other immunotherapy agents, in particular specific checkpoint inhibitors, for cancer treatment. To date, however, only a minority of patients respond. Transcriptomic profiling reveals that OX40 and OX40L expression vary between and within tumor types, and that only ~ 17% of cancer patients have high OX40 and low OX40L, one of the expression patterns that might be theoretically amenable to OX40 agonist enhancement. Taken together, the data suggest that the OX40/OX40L machinery is a critical part of the immune stimulatory system and that understanding endogenous expression patterns of these molecules and co-existing checkpoints merits further investigation in the context of a precision immunotherapy strategy for cancer therapy.

Report of the APOE4 National Institute on Aging/Alzheimer Disease Sequencing Project Consortium Working Group: Reducing APOE4 in Carriers is a Therapeutic Goal for Alzheimer's Disease.

Alzheimer's disease (AD) is the most common neurodegenerative disorder and one of the leading causes of disability worldwide. The apolipoprotein E4 gene (APOE4) is the strongest genetic risk factor for AD. In 2023, the APOE4 National Institute on Aging/Alzheimer's Disease Sequencing Project working group came together to gather data and discuss the question of whether to reduce or increase APOE4 as a therapeutic intervention for AD. It was the unanimous consensus that cumulative data from multiple studies in humans and animal models support that lowering APOE4 should be a target for therapeutic approaches for APOE4 carriers. ANN NEUROL 2024;95:625-634.

A locus at 19q13.31 significantly reduces the ApoE ε4 risk for Alzheimer's Disease in African Ancestry.

African descent populations have a lower Alzheimer disease risk from ApoE ε4 compared to other populations. Ancestry analysis showed that the difference in risk between African and European populations lies in the ancestral genomic background surrounding the ApoE locus (local ancestry). Identifying the mechanism(s) of this protection could lead to greater insight into the etiology of Alzheimer disease and more personalized therapeutic intervention. Our objective is to follow up the local ancestry finding and identify the genetic variants that drive this risk difference and result in a lower risk for developing Alzheimer disease in African ancestry populations. We performed association analyses using a logistic regression model with the ApoE ε4 allele as an interaction term and adjusted for genome-wide ancestry, age, and sex. Discovery analysis included imputed SNP data of 1,850 Alzheimer disease and 4,331 cognitively intact African American individuals. We performed replication analyses on 63 whole genome sequenced Alzheimer disease and 648 cognitively intact Ibadan individuals. Additionally, we reproduced results using whole-genome sequencing of 273 Alzheimer disease and 275 cognitively intact admixed Puerto Rican individuals. A further comparison was done with SNP imputation from an additional 8,463 Alzheimer disease and 11,365 cognitively intact non-Hispanic White individuals. We identified a significant interaction between the ApoE ε4 allele and the SNP rs10423769_A allele, (ß = -0.54,SE = 0.12,p-value = 7.50x10-6) in the discovery data set, and replicated this finding in Ibadan (ß = -1.32,SE = 0.52,p-value = 1.15x10-2) and Puerto Rican (ß = -1.27,SE = 0.64,p-value = 4.91x10-2) individuals. The non-Hispanic Whites analyses showed an interaction trending in the "protective" direction but failing to pass a 0.05 significance threshold (ß = -1.51,SE = 0.84,p-value = 7.26x10-2). The presence of the rs10423769_A allele reduces the odds ratio for Alzheimer disease risk from 7.2 for ApoE ε4/ε4 carriers lacking the A allele to 2.1 for ApoE ε4/ε4 carriers with at least one A allele. This locus is located approximately 2 mB upstream of the ApoE locus, in a large cluster of pregnancy specific beta-1 glycoproteins on chromosome 19 and lies within a long noncoding RNA, ENSG00000282943. This study identified a new African-ancestry specific locus that reduces the risk effect of ApoE ε4 for developing Alzheimer disease. The mechanism of the interaction with ApoEε4 is not known but suggests a novel mechanism for reducing the risk for ε4 carriers opening the possibility for potential ancestry-specific therapeutic intervention.

Pediatric Erythroid Sarcoma Diagnostically Confirmed by Identification of a Recurrent NFIA::CBFA2T3 Fusion.

Erythroid sarcoma (ES) is exceedingly rare in the pediatric population with only a handful of reports of de novo cases, mostly occurring in the central nervous system (CNS) or orbit. It is clinically and pathologically challenging and can masquerade as a nonhematopoietic small round blue cell tumor. Clinical presentation of ES without bone marrow involvement makes diagnosis particularly difficult. We describe a 22-month-old female with ES who presented with a 2-cm mass involving the left parotid region and CNS. The presence of crush/fixation artifact from the initial biopsy made definitive classification of this highly proliferative and malignant neoplasm challenging despite an extensive immunohistochemical workup. Molecular studies including RNA-sequencing revealed a NFIA::CBFA2T3 fusion. This fusion has been identified in several cases of de novo acute erythroid leukemia (AEL) and gene expression analysis comparing this case to other AELs revealed a similar transcriptional profile. Given the diagnostically challenging nature of this tumor, clinical RNA-sequencing was essential for establishing a diagnosis.

Genetic architecture of RNA editing regulation in Alzheimer's disease across diverse ancestral populations.

Most Alzheimer's disease (AD)-associated genetic variants do not change protein coding sequence and thus likely exert their effects through regulatory mechanisms. RNA editing, the post-transcriptional modification of RNA bases, is a regulatory feature that is altered in AD patients that differs across ancestral backgrounds. Editing QTLs (edQTLs) are DNA variants that influence the level of RNA editing at a specific site. To study the relationship of DNA variants genome-wide, and particularly in AD-associated loci, with RNA editing, we performed edQTL analyses in self-reported individuals of African American (AF) or White (EU) race with corresponding global genetic ancestry averaging 82.2% African ancestry (AF) and 96.8% European global ancestry (EU) in the two groups, respectively. We used whole-genome genotyping array and RNA sequencing data from peripheral blood of 216 AD cases and 212 age-matched, cognitively intact controls. We identified 2144 edQTLs in AF and 3579 in EU, of which 1236 were found in both groups. Among these, edQTLs in linkage disequilibrium (r2 > 0.5) with AD-associated genetic variants in the SORL1, SPI1 and HLA-DRB1 loci were associated with sites that were differentially edited between AD cases and controls. While there is some shared RNA editing regulatory architecture, most edQTLs had distinct effects on the rate of RNA editing in different ancestral populations suggesting a complex architecture of RNA editing regulation. Altered RNA editing may be one possible mechanism for the functional effect of AD-associated variants and may contribute to observed differences in the genetic etiology of AD between ancestries.

Baseline characteristics including blood and urine metal levels in the Trial to Assess Chelation Therapy 2 (TACT2).

BACKGROUND: The reduction in cardiovascular disease (CVD) events with edetate disodium (EDTA) in the Trial to Assess Chelation Therapy (TACT) suggested that chelation of toxic metals might provide novel opportunities to reduce CVD in patients with diabetes. Lead and cadmium are vasculotoxic metals chelated by EDTA. We present baseline characteristics for participants in TACT2, a randomized, double-masked, placebo-controlled trial designed as a replication of the TACT trial limited to patients with diabetes. METHODS: TACT2 enrolled 1,000 participants with diabetes and prior myocardial infarction, age 50 years or older between September 2016 and December 2020. Among 959 participants with at least one infusion, 933 had blood and/or urine metals measured at the Centers for Diseases Control and Prevention using the same methodology as in the National Health and Nutrition Examination Survey (NHANES). We compared metal levels in TACT2 to a contemporaneous subset of NHANES participants with CVD, diabetes and other inclusion criteria similar to TACT2's participants. RESULTS: At baseline, the median (interquartile range, IQR) age was 67 (60, 72) years, 27% were women, 78% reported white race, mean (SD) BMI was 32.7 (6.6) kg/m2, 4% reported type 1 diabetes, 46.8% were treated with insulin, 22.3% with GLP1-receptor agonists or SGLT-2 inhibitors, 90.2% with aspirin, warfarin or P2Y12 inhibitors, and 86.5% with statins. Blood lead was detectable in all participants; median (IQR) was 9.19 (6.30, 13.9) µg/L. Blood and urine cadmium were detectable in 97% and median (IQR) levels were 0.28 (0.18, 0.43) µg/L and 0.30 (0.18, 0.51) µg/g creatinine, respectively. Metal levels were largely similar to those in the contemporaneous NHANES subset. CONCLUSIONS: TACT2 participants were characterized by high use of medication to treat CVD and diabetes and similar baseline metal levels as in the general US population. TACT2 will determine whether chelation therapy reduces the occurrence of subsequent CVD events in this high-risk population. CLINICAL TRIALS REGISTRATION: ClinicalTrials.gov. Identifier: NCT02733185. https://clinicaltrials.gov/study/NCT02733185.

Evaluation of Bayesian Point-Based System on the Variant Classification of Hereditary Cancer Predisposition Genes.

PURPOSE: To assess the differences in variant classifications using the ACMG/AMP 2015 guidelines and the Bayesian point-based classification system (here referred to as the point system) in 115 hereditary cancer predisposition genes and explore variant sub-tiering by the point system. METHODS: Germline variant classifications for 721 pediatric patients from an in-house panel were retrospectively evaluated using the two scoring systems. RESULTS: 2376 unique variants were identified, with ∼23.5% exhibiting discordant classifications. Unique variants classified by the point system demonstrated a lower rate of variants of uncertain significance (VUS; ∼15%) compared to ACMG/AMP 2015 (∼36%). This change is attributed to unique variants with one benign supporting evidence (∼12%) or one benign strong evidence (∼4%) being classified as likely benign by the point system. Additionally, variants with conflicting/modified evidence (∼5% of 2376) are also resolved by the point system. Sub-tiering unique variants classified by the point system as VUS (n=354) indicates ∼77.4% were VUS-Low (0-1 points), while the remaining ∼22.6% were VUS-Mid (2-3 points) and VUS-High (4-5 points). CONCLUSION: The point system reduces the VUS rate and facilitates their sub-tiering. Future large-scale studies are warranted to explore the impact of the point system on improving VUS reporting and/or VUS clinical management.

Engineering naturally occurring CD7- T cells for the immunotherapy of hematological malignancies.

Chimeric antigen receptor (CAR) T-cell therapy targeting T-cell acute lymphoblastic leukemia (T-ALL) faces limitations such as antigen selection and limited T-cell persistence. CD7 is an attractive antigen for targeting T-ALL, but overlapping expression on healthy T cells leads to fratricide of CD7-CAR T cells, requiring additional genetic modification. We took advantage of naturally occurring CD7- T cells to generate CD7-CAR (CD7-CARCD7-) T cells. CD7-CARCD7- T cells exhibited a predominantly CD4+ memory phenotype and had significant antitumor activity upon chronic antigen exposure in vitro and in xenograft mouse models. Based on these encouraging results, we next explored the utility of CD7- T cells for the immunotherapy of CD19+ hematological malignancies. Direct comparison of nonselected (bulk) CD19-CAR and CD19-CARCD7- T cells revealed that CD19-CARCD7- T cells had enhanced antitumor activity compared with their bulk counterparts in vitro and in vivo. Lastly, to gain insight into the behavior of CD19-CAR T cells with low levels of CD7 gene expression (CD7lo) in humans, we mined single-cell gene and T-cell receptor (TCR) expression data sets from our institutional CD19-CAR T-cell clinical study. CD19-CARCD7lo T cells were present in the initial CD19-CAR T-cell product and could be detected postinfusion. Intriguingly, the only functional CD4+ CD19-CAR T-cell cluster observed postinfusion exhibited CD7lo expression. Additionally, samples from patients responsive to therapy had a higher proportion of CD7lo T cells than nonresponders (NCT03573700). Thus, CARCD7- T cells have favorable biological characteristics and may present a promising T-cell subset for adoptive cell therapy of T-ALL and other hematological malignancies.

MS4A3 promotes differentiation in chronic myeloid leukemia by enhancing common ß-chain cytokine receptor endocytosis.

The chronic phase of chronic myeloid leukemia (CP-CML) is characterized by the excessive production of maturating myeloid cells. As CML stem/progenitor cells (LSPCs) are poised to cycle and differentiate, LSPCs must balance conservation and differentiation to avoid exhaustion, similar to normal hematopoiesis under stress. Since BCR-ABL1 tyrosine kinase inhibitors (TKIs) eliminate differentiating cells but spare BCR-ABL1-independent LSPCs, understanding the mechanisms that regulate LSPC differentiation may inform strategies to eliminate LSPCs. Upon performing a meta-analysis of published CML transcriptomes, we discovered that low expression of the MS4A3 transmembrane protein is a universal characteristic of LSPC quiescence, BCR-ABL1 independence, and transformation to blast phase (BP). Several mechanisms are involved in suppressing MS4A3, including aberrant methylation and a MECOM-C/EBPε axis. Contrary to previous reports, we find that MS4A3 does not function as a G1/S phase inhibitor but promotes endocytosis of common ß-chain (ßc) cytokine receptors upon GM-CSF/IL-3 stimulation, enhancing downstream signaling and cellular differentiation. This suggests that LSPCs downregulate MS4A3 to evade ßc cytokine-induced differentiation and maintain a more primitive, TKI-insensitive state. Accordingly, knockdown (KD) or deletion of MS4A3/Ms4a3 promotes TKI resistance and survival of CML cells ex vivo and enhances leukemogenesis in vivo, while targeted delivery of exogenous MS4A3 protein promotes differentiation. These data support a model in which MS4A3 governs response to differentiating myeloid cytokines, providing a unifying mechanism for the differentiation block characteristic of CML quiescence and BP-CML. Promoting MS4A3 reexpression or delivery of ectopic MS4A3 may help eliminate LSPCs in vivo.

Genomic profiling for clinical decision making in myeloid neoplasms and acute leukemia.

Myeloid neoplasms and acute leukemias derive from the clonal expansion of hematopoietic cells driven by somatic gene mutations. Although assessment of morphology plays a crucial role in the diagnostic evaluation of patients with these malignancies, genomic characterization has become increasingly important for accurate diagnosis, risk assessment, and therapeutic decision making. Conventional cytogenetics, a comprehensive and unbiased method for assessing chromosomal abnormalities, has been the mainstay of genomic testing over the past several decades and remains relevant today. However, more recent advances in sequencing technology have increased our ability to detect somatic mutations through the use of targeted gene panels, whole-exome sequencing, whole-genome sequencing, and whole-transcriptome sequencing or RNA sequencing. In patients with myeloid neoplasms, whole-genome sequencing represents a potential replacement for both conventional cytogenetic and sequencing approaches, providing rapid and accurate comprehensive genomic profiling. DNA sequencing methods are used not only for detecting somatically acquired gene mutations but also for identifying germline gene mutations associated with inherited predisposition to hematologic neoplasms. The 2022 International Consensus Classification of myeloid neoplasms and acute leukemias makes extensive use of genomic data. The aim of this report is to help physicians and laboratorians implement genomic testing for diagnosis, risk stratification, and clinical decision making and illustrates the potential of genomic profiling for enabling personalized medicine in patients with hematologic neoplasms.

UBTF tandem duplications in pediatric myelodysplastic syndrome and acute myeloid leukemia: implications for clinical screening and diagnosis.

Recent genomic studies in adult and pediatric acute myeloid leukemia (AML) demonstrated recurrent in-frame tandem duplications (TD) in exon 13 of upstream binding transcription factor (UBTF). These alterations, which account for approximately 4.3% of AML in childhood and about 3% in adult AML aged <60 years of age, are subtype-defining and associated with poor outcomes. Here, we provide a comprehensive investigation into the clinicopathological features of UBTF-TD myeloid neoplasms in childhood, including 89 unique pediatric AML and 6 myelodysplastic syndrome (MDS) cases harboring a tandem duplication in exon 13 of UBTF. We demonstrate that UBTF-TD myeloid tumors are associated with dysplastic features, low bone marrow blast infiltration, and low white blood cell count. Furthermore, using bulk and single-cell analyses, we confirm that UBTF-TD is an early and clonal event associated with a distinct transcriptional profile, whereas the acquisition of FLT3 or WT1 mutations is associated with more stem cell-like programs. Lastly, we report rare duplications within exon 9 of UBTF that phenocopy exon 13 duplications, expanding the spectrum of UBTF alterations in pediatric myeloid tumors. Collectively, we comprehensively characterize pediatric AML and MDS with UBTF-TD, and highlight key clinical and pathologic features that distinguish this new entity from other molecular subtypes of AML.

Comparison of treatment strategies based on clinical and pathological nodal status in resectable gastric adenocarcinoma.

BACKGROUND: To determine the optimal multimodal treatment strategy between perioperative chemotherapy (PEC), postoperative chemoradiation therapy (POCR), and postoperative chemotherapy (POC) in resected gastric cancer (GC) patients based on nodal status. METHODS: In this retrospective analysis, the National Cancer Database was used to identify resected non-metastatic GC (2006-2016). Patients were stratified by clinical nodal status-negative (cLN-) and positive (cLN+). In patients with cLN- disease who underwent upfront resection and were upstaged to pathological LN+, overall survival (OS) was compared between POC and POCR. In patients with cLN- and cLN+ disease, OS was compared between PEC, POCR, and POC. Kaplan-Meier survival estimate, log-rank test, and multivariable Cox proportional hazards analysis were performed. RESULTS: We identified 7827 patients (cLN- 4828; cLN+ 2999). On multivariable analysis in patients with cLN- disease who underwent upfront resection (n = 4314) and were upstaged to pLN+ disease (70%), POCR (n = 2300, aHR 0.78, 95% CI 0.70-0.87, p < 0.001) was associated with improved OS compared to POC (n = 907). No significant difference was noted between POCR (n = 766, aHR 1.11, 95% CI 0.88-1.40, p = 0.39) and POC (n = 341) in patients with pLN- disease. On multivariable analysis in all patients with cLN- disease, POCR (n = 3066) was significantly associated with improved OS (aHR 0.84, 95% CI 0.75-0.92, p < 0.01) compared to POC (n = 1248). No significant difference was noted between POCR (aHR 1.0, 95% CI 0.70-1.01, p = 0.958) and PEC (n = 514). These results remained consistent in patients with cLN+ disease (POCR = 1602, POC = 720, PEC = 677). CONCLUSION: Postoperative chemoradiation is associated with improved survival in GC patients upstaged from clinically node-negative disease to pathologically node-positive disease. Negative clinical nodal disease status is not a reliable indicator of pathological nodal disease.