Females translate male mRNA transferred during mating.

Although RNA is found in the seminal fluid of diverse organisms, it is unknown whether it is functional within females. We developed a proteomic method (VESPA, Variant Enabled SILAC Proteomic Analysis) to test the hypothesis that Drosophila male seminal fluid RNA is translated by females. We found 67 male-derived, female-translated proteins (mdFTPs) in female lower reproductive tracts, many with predicted functions relevant to reproduction. Knockout experiments indicate that mdFTPs play diverse roles in postmating interactions, affecting fertilization success, and the formation/persistence of the insemination reaction mass, a trait hypothesized to be involved in sexual conflict. These findings advance our understanding of reproduction by revealing a mechanism of postmating molecular interactions between the sexes that strengthens and extends male influences on reproduction in previously unrecognized ways. Given the diverse species that carry RNA in seminal fluid, this discovery has broad significance for understanding molecular mechanisms of cooperation and conflict during reproduction.

Targeting intracellular nontuberculous mycobacteria and M. tuberculosis with a bactericidal enzymatic cocktail.

To address intracellular mycobacterial infections, we developed a cocktail of four enzymes that catalytically attack three layers of the mycobacterial envelope. This cocktail is delivered to macrophages, through a targeted liposome presented here as ENTX_001. Endolytix Cocktail 1 (EC1) leverages mycobacteriophage lysin enzymes LysA and LysB, while also including α-amylase and isoamylase for degradation of the mycobacterial envelope from outside of the cell. The LysA family of proteins from mycobacteriophages has been shown to cleave the peptidoglycan layer, whereas LysB is an esterase that hydrolyzes the linkage between arabinogalactan and mycolic acids of the mycomembrane. The challenge of gaining access to the substrates of LysA and LysB provided exogenously was addressed by adding amylase enzymes that degrade the extracellular capsule shown to be present in Mycobacterium tuberculosis. This enzybiotic approach avoids antimicrobial resistance, specific receptor-mediated binding, and intracellular DNA surveillance pathways that limit many bacteriophage applications. We show this cocktail of enzymes is bactericidal in vitro against both rapid- and slow-growing nontuberculous mycobacteria (NTM) as well as M. tuberculosis strains. The EC1 cocktail shows superior killing activity when compared to previously characterized LysB alone. EC1 is also powerfully synergistic with standard-of-care antibiotics. In addition to in vitro killing of NTM, ENTX_001 demonstrates the rescue of infected macrophages from necrotic death by Mycobacteroides abscessus and Mycobacterium avium. Here, we demonstrate shredding of mycobacterial cells by EC1 into cellular debris as a mechanism of bactericide.IMPORTANCEThe world needs entirely new forms of antibiotics as resistance to chemical antibiotics is a critical problem facing society. We addressed this need by developing a targeted enzyme therapy for a broad range of species and strains within mycobacteria and highly related genera including nontuberculous mycobacteria such as Mycobacteroides abscessus, Mycobacterium avium, Mycobacterium intracellulare, as well as Mycobacterium tuberculosis. One advantage of this approach is the ability to drive our lytic enzymes through encapsulation into macrophage-targeted liposomes resulting in attack of mycobacteria in the cells that harbor them where they hide from the adaptive immune system and grow. Furthermore, this approach shreds mycobacteria independent of cell physiology as the drug targets the mycobacterial envelope while sidestepping the host range limitations observed with phage therapy and resistance to chemical antibiotics.

Females translate male mRNA transferred during mating.

Although RNA is found in the seminal fluid of diverse organisms, it is unknown whether this RNA is functional within females. Here, we develop an experimental proteomic method called VESPA (Variant Enabled SILAC Proteomic Analysis) to test the hypothesis that Drosophila male seminal fluid RNA is translated by females. We find strong evidence for 67 male-derived, female-translated proteins (mdFTPs) in female lower reproductive tracts at six hours postmating, many with predicted functions relevant to reproduction. Gene knockout experiments indicate that genes coding for mdFTPs play diverse roles in postmating interactions, with effects on fertilization efficiency, and the formation and persistence of the insemination reaction mass, a trait hypothesized to be involved in sexual conflict. These findings advance our understanding of reproduction by revealing a novel mechanism of postmating molecular interactions between the sexes that strengthens and extends male influences on reproductive outcomes in previously unrecognized ways. Given the diverse species known to carry RNA in seminal fluid, this discovery has broad significance for understanding molecular mechanisms of cooperation and conflict during reproduction.

Evaluation of a Delta-9-Tetrahydrocannabinol Carboxylic Acid (Δ9-THC-COOH) Immunoassay and a Gas Chromatography-Mass Spectrometry (GC-MS) Method for the Detection of Delta-8-Tetrahydrocannabinol Carboxylic Acid (Δ8-THC-COOH).

BACKGROUND: Delta-8 tetrahydrocannabinol (Δ8-THC) is a naturally occurring or synthetically prepared cannabinoid that elicits psychological and physiological experiences commonly reported for its more infamous isomer, delta-9 tetrahydrocannabinol (Δ9-THC). Unlike Δ9-THC, Δ8-THC products are generally legal under federal law and there has been a rise in their usage. One of the main targets for detection and quantitation of Δ9-THC is its inactive metabolite, 11-nor-9-carboxy-Δ9-tetrahydrocannabinol (Δ9-THC-COOH). METHODS: This study evaluated the ability of the currently used Δ9-THC-COOH immunoassay and gas chromatography-mass spectrometry (GC-MS) methods to detect 11-nor-9-carboxy-Δ8-tetrahydrocannabinol (Δ8-THC-COOH) and distinguish it from Δ9-THC-COOH. RESULTS: The EMIT II Plus® Cannabinoid immunoassay for Δ9-THC-COOH with a cutoff of 20 ng/mL showed positive results for Δ8-THC-COOH with concentrations of 30 ng/mL or higher. Although many of the ion fragments generated by mass spectrometry were found to overlap between the 2 compounds, the GC-MS method presently used to quantify Δ9-THC-COOH separated the 2 compounds sufficiently to identify them independently by relative retention time. CONCLUSION: Current immunoassays and GC-MS methods should be evaluated for the ability to detect and distinguish the presence of Δ8-THC-COOH.

Specialized Ribosomes in Health and Disease.

Ribosomal heterogeneity exists within cells and between different cell types, at specific developmental stages, and occurs in response to environmental stimuli. Mounting evidence supports the existence of specialized ribosomes, or specific changes to the ribosome that regulate the translation of a specific group of transcripts. These alterations have been shown to affect the affinity of ribosomes for certain mRNAs or change the cotranslational folding of nascent polypeptides at the exit tunnel. The identification of specialized ribosomes requires evidence of the incorporation of different ribosomal proteins or of modifications to rRNA and/or protein that lead(s) to physiologically relevant changes in translation. In this review, we summarize ribosomal heterogeneity and specialization in mammals and discuss their relevance to several human diseases.

Multicenter study of pediatric Epstein-Barr virus-negative monomorphic post solid organ transplant lymphoproliferative disorders.

BACKGROUND: Pediatric Epstein-Barr virus-negative monomorphic post solid organ transplant lymphoproliferative disorder [EBV(-)M-PTLD] comprises approximately 10% of M-PTLD. No large multi-institutional pediatric-specific reports on treatment and outcome are available. METHODS: A multi-institutional retrospective review of solid organ recipients diagnosed with EBV(-)M-PTLD aged ≤21 years between 2001 and 2020 in 12 centers in the United States and United Kingdom was performed, including demographics, staging, treatment, and outcomes data. RESULTS: Thirty-six patients were identified with EBV(-)M-PTLD. Twenty-three (63.9%) were male. Median age (range) at transplantation, diagnosis of EBV(-)M-PTLD, and interval from transplant to PTLD were 2.2 years (0.1-17), 14 years (3.0-20), and 8.5 years (0.6-18.3), respectively. Kidney (n = 17 [47.2%]) and heart (n = 13 [36.1%]) were the most commonly transplanted organs. Most were Murphy stage III (n = 25 [69.4%]). Lactate dehydrogenase was elevated in 22/34 (64.7%) and ≥2 times upper limit of normal in 11/34 (32.4%). Pathological diagnoses included diffuse large B-cell lymphoma (n = 31 [86.1%]) and B-non-Hodgkin lymphoma (B-NHL) not otherwise specified (NOS) (n = 5 [13.9%]). Of nine different regimens used, the most common were: pediatric mature B-NHL-specific regimen (n = 13 [36.1%]) and low-dose cyclophosphamide, prednisone, and rituximab (n = 9 [25%]). Median follow-up from diagnosis was 3.0 years (0.3-11.0 years). Three-year event-free survival (EFS) and overall survival (OS) were 64.8% and 79.9%, respectively. Of the seven deaths, six were from progressive disease. CONCLUSIONS: EFS and OS were comparable to pediatric EBV(+) PTLD, but inferior to mature B-NHL in immunocompetent pediatric patients. The wide range of therapeutic regimens used directs our work toward developing an active multi-institutional registry to design prospective studies. PLAIN LANGUAGE SUMMARY: Pediatric Epstein-Barr virus-negative monomorphic post solid organ transplant lymphoproliferative disorders (EBV(-)M-PTLD) have comparable outcomes to EBV(+) PTLD, but are inferior to diffuse large B-cell lymphoma in immunocompetent pediatric patients. The variety of treatment regimens used highlights the need to develop a pediatric PTLD registry to prospectively evaluate outcomes. The impact of treatment regimen on relapse risk could not be assessed because of small numbers. In the intensive pediatric B-non-Hodgkin lymphoma chemoimmunotherapy group, 11 of 13 patients remain alive in complete remission after 0.6 to 11 years.

Burkitt lymphoma after solid-organ transplant: Treatment and outcomes in the paediatric PTLD collaborative.

Burkitt lymphoma arising in paediatric post-solid-organ transplantation-Burkitt lymphoma (PSOT-BL) is a clinically aggressive malignancy and a rare form of post-transplant lymphoproliferative disorder (PTLD). We evaluated 35 patients diagnosed with PSOT-BL at 14 paediatric medical centres in the United States. Median age at organ transplantation was 2.0 years (range: 0.1-14) and age at PSOT-BL diagnosis was 8.0 years (range: 1-17). All but one patient had late onset of PSOT-BL (≥2 years post-transplant), with a median interval from transplant to PSOT-BL diagnosis of 4.0 years (range: 0.4-12). Heart (n = 18 [51.4%]) and liver (n = 13 [37.1%]) were the most frequently transplanted organs. No patients had loss of graft or treatment-related mortality. A variety of treatment regimens were used, led by intensive Burkitt lymphoma-specific French-American-British/Lymphomes Malins B (FAB/LMB), n = 13 (37.1%), and a low-intensity regimen consisting of cyclophosphamide, prednisone and rituximab (CPR) n = 12 (34.3%). Median follow-up was 6.7 years (range: 0.5-17). Three-year event-free and overall survival were 66.2% and 88.0%, respectively. Outcomes of PSOT-BL patients receiving BL-specific intensive regimens are comparable to reported BL outcomes in immunocompetent children. Multi-institutional collaboration is feasible and provides the basis of prospective data collection to determine the optimal treatment regimen for PSOT-BL.

Loss of G0/G1 switch gene 2 (G0S2) promotes disease progression and drug resistance in chronic myeloid leukaemia (CML) by disrupting glycerophospholipid metabolism.

Tyrosine kinase inhibitors (TKIs) targeting BCR::ABL1 have turned chronic myeloid leukaemia (CML) from a fatal disease into a manageable condition for most patients. Despite improved survival, targeting drug-resistant leukaemia stem cells (LSCs) remains a challenge for curative CML therapy. Aberrant lipid metabolism can have a large impact on membrane dynamics, cell survival and therapeutic responses in cancer. While ceramide and sphingolipid levels were previously correlated with TKI response in CML, the role of lipid metabolism in TKI resistance is not well understood. We have identified downregulation of a critical regulator of lipid metabolism, G0/G1 switch gene 2 (G0S2), in multiple scenarios of TKI resistance, including (1) BCR::ABL1 kinase-independent TKI resistance, (2) progression of CML from the chronic to the blast phase of the disease, and (3) in CML versus normal myeloid progenitors. Accordingly, CML patients with low G0S2 expression levels had a worse overall survival. G0S2 downregulation in CML was not a result of promoter hypermethylation or BCR::ABL1 kinase activity, but was rather due to transcriptional repression by MYC. Using CML cell lines, patient samples and G0s2 knockout (G0s2-/- ) mice, we demonstrate a tumour suppressor role for G0S2 in CML and TKI resistance. Our data suggest that reduced G0S2 protein expression in CML disrupts glycerophospholipid metabolism, correlating with a block of differentiation that renders CML cells resistant to therapy. Altogether, our data unravel a new role for G0S2 in regulating myeloid differentiation and TKI response in CML, and suggest that restoring G0S2 may have clinical utility.

Discovery of a novel type IIb RelBE toxin-antitoxin system in Mycobacterium tuberculosis defined by co-regulation with an antisense RNA.

Toxin-antitoxin loci regulate adaptive responses to stresses associated with the host environment and drug exposure. Phylogenomic studies have shown that Mycobacterium tuberculosis encodes a naturally expanded type II toxin-antitoxin system, including ParDE/RelBE superfamily members. Type II toxins are presumably regulated exclusively through protein-protein interactions with type II antitoxins. However, experimental observations in M. tuberculosis indicated that additional control mechanisms regulate RelBE2 type II loci under host-associated stress conditions. Herein, we describe for the first time a novel antisense RNA, termed asRelE2, that co-regulates RelE2 production via targeted processing by the Mtb RNase III, Rnc. We find that convergent expression of this coding-antisense hybrid TA locus, relBE2-asrelE2, is controlled in a cAMP-dependent manner by the essential cAMP receptor protein transcription factor, Crp, in response to the host-associated stresses of low pH and nutrient limitation. Ex vivo survival studies with relE2 and asrelE2 knockout strains showed that RelE2 contributes to Mtb survival in activated macrophages and low pH to nutrient limitation. To our knowledge, this is the first report of a novel tripartite type IIb TA loci and antisense post-transcriptional regulation of a type II TA loci.

Responses to Ixekizumab in Male and Female Patients with Psoriatic Arthritis: Results from Two Randomized, Phase 3 Clinical Trials.

INTRODUCTION: Differences in psoriatic arthritis (PsA) treatment response between sexes for ixekizumab, an interleukin-17A antagonist, are largely unexplored. This analysis used data from randomized clinical trials (RCTs) evaluating ixekizumab to study differences in treatment response between male and female patients with PsA. METHODS: We used pooled data from patients enrolled in SPIRIT-P1 and SPIRIT-P2 (NCT01695239 and NCT02349295, respectively), phase 3 RCTs evaluating ixekizumab every 4 and 2 weeks in patients with active PsA. Subgroups of patients were defined by sex (male, female). Efficacy was measured by the proportion of male and female patients achieving American College of Rheumatology 20%/50%/70% improvement criteria (ACR20/50/70), minimal disease activity or very low disease activity (MDA/VLDA), and Disease Activity Index for Psoriatic Arthritis (DAPSA) scores representing low disease activity (LDA) or remission through week 156. Changes from baseline in components of the above measures were also assessed through week 156. RESULTS: Compared to male patients at baseline, female patients were older, had higher body mass index and lower C-reactive protein levels, and had worse tender joint count, Health Assessment Questionnaire Disability Index, and Leeds Enthesitis Index scores. Through week 156, female patients in all treatment arms had lower response rates than male patients in all analyzed composite measures (ACR20/50/70; MDA/VLDA; DAPSA LDA/remission), with significant differences observed at multiple timepoints in both ixekizumab treatment arms. Female patients also had smaller numeric changes from baseline in the composite measures' individual components. CONCLUSION: Compared to female patients, male patients had greater response rates in ACR20/50/70, MDA/VLDA, and DAPSA LDA/remission and numerically larger improvements in these measures' individual components, although clinical significance is unclear. Continued efforts to understand sex differences in treatment response may provide insights that can help optimize clinical decision making. TRIAL REGISTRATION: ClinicalTrials.gov identifiers, NCT01695239 and NCT02349295.

Validation of an algorithm to identify incident interstitial lung disease in patients with rheumatoid arthritis.

BACKGROUND/PURPOSE: Interstitial lung disease (ILD) is an important problem for patients with rheumatoid arthritis (RA). However, current approaches to ILD case finding in real-world data have been evaluated only in limited settings and identify only prevalent ILD and not new-onset disease. Our objective was to develop, refine, and validate a claims-based algorithm to identify both prevalent and incident ILD in RA patients compared to the gold standard of medical record review. METHODS: We used administrative claims data 2006-2015 from Medicare to derive a cohort of RA patients. We then identified suspected ILD using variations of ILD algorithms to classify both prevalent and incident ILD based on features of the data that included hospitalization vs. outpatient setting, physician specialty, pulmonary-related diagnosis codes, and exclusions for potentially mimicking pulmonary conditions. Positive predictive values (PPV) of several ILD algorithm variants for both prevalent and incident ILD were evaluated. RESULTS: We identified 234 linkable RA patients with sufficient data to evaluate for ILD. Overall, 108 (46.2%) of suspected cases were confirmed as ILD. Most cases (64%) were diagnosed in the outpatient setting. The best performing algorithm for prevalent ILD had a PPV of 77% (95% CI 67-84%) and for incident ILD was 96% (95% CI 85-100%). CONCLUSION: Case finding in administrative data for both prevalent and incident interstitial lung disease in RA patients is feasible and has reasonable accuracy to support population-based research and real-world evidence generation.

Electrostatic Interactions between CSTF2 and pre-mRNA Drive Cleavage and Polyadenylation.

Nascent pre-mRNA 3'-end cleavage and polyadenylation (C/P) involves numerous proteins that recognize multiple RNA elements. Human CSTF2 binds to a downstream U- or G/U-rich sequence through its RNA recognition motif (RRM) regulating C/P. We previously reported the only known disease-related CSTF2 RRM mutant (CSTF2D50A) and showed that it changed the on-rate of RNA binding, leading to alternative polyadenylation in brains of mice carrying the same mutation. In this study, we further investigated the role of electrostatic interactions in the thermodynamics and kinetics of RNA binding for the CSTF2 RRM and the downstream consequences for regulation of C/P. By combining mutagenesis with NMR spectroscopy and biophysical assays, we confirmed that electrostatic attraction is the dominant factor in RRM binding to a naturally occurring U-rich RNA sequence. Moreover, we demonstrate that RNA binding is accompanied by an enthalpy-entropy compensation mechanism that is supported by changes in pico-to-nanosecond timescale RRM protein dynamics. We suggest that the dynamic binding of the RRM to U-rich RNA supports the diversity of sequences it encounters in the nucleus. Lastly, in vivo C/P assays demonstrate a competition between fast, high affinity RNA binding and efficient, correct C/P. These results highlight the importance of the surface charge of the RRM in RNA binding and the balance between nascent mRNA binding and C/P in vivo.

Findings from the Hispanic Community Health Study/Study of Latinos on the Importance of Sociocultural Environmental Interactors: Polygenic Risk Score-by-Immigration and Dietary Interactions.

Introduction: Hispanic/Latinos experience a disproportionate burden of obesity. Acculturation to US obesogenic diet and practices may lead to an exacerbation of innate genetic susceptibility. We examined the role of gene-environment interactions to better characterize the sociocultural environmental determinants and their genome-scale interactions, which may contribute to missing heritability of obesity. We utilized polygenic risk scores (PRSs) for body mass index (BMI) to perform analyses of PRS-by-acculturation and other environmental interactors among self-identified Hispanic/Latino adults from the Hispanic Community Health Study/Study of Latinos (HCHS/SOL). Methods: PRSs were derived using genome-wide association study (GWAS) weights from a publicly available, large meta-analysis of European ancestry samples. Generalized linear models were run using a set of a priori acculturation-related and environmental factors measured at visit 1 (2008-2011) and visit 2 (2014-2016) in an analytic subsample of 8,109 unrelated individuals with genotypic, phenotypic, and complete case data at both visits. We evaluated continuous measures of BMI and waist-to-hip ratio. All models were weighted for complex sampling design, combined, and sex-stratified. Results: Overall, we observed a consistent increase of BMI with greater PRS across both visits. We found the best-fitting model adjusted for top five principal components of ancestry, sex, age, study site, Hispanic/Latino background genetic ancestry group, sociocultural factors and PRS interactions with age at immigration, years since first arrival to the United States (p < 0.0104), and healthy diet (p < 0.0036) and explained 16% of the variation in BMI. For every 1-SD increase in PRS, there was a corresponding 1.10 kg/m2 increase in BMI (p < 0.001). When these results were stratified by sex, we observed that this 1-SD effect of PRS on BMI was greater for women than men (1.45 vs. 0.79 kg/m2, p < 0.001). Discussion: We observe that age at immigration and the adoption of certain dietary patterns may play a significant role in modifying the effect of genetic risk on obesity. Careful consideration of sociocultural and immigration-related factors should be evaluated. The role of nongenetic factors, including the social environment, should not be overlooked when describing the performance of PRS or for promoting population health in understudied populations in genomics.

Carfilzomib Enhances the Suppressive Effect of Ruxolitinib in Myelofibrosis.

As the first FDA-approved tyrosine kinase inhibitor for treatment of patients with myelofibrosis (MF), ruxolitinib improves clinical symptoms but does not lead to eradication of the disease or significant reduction of the mutated allele burden. The resistance of MF clones against the suppressive action of ruxolitinib may be due to intrinsic or extrinsic mechanisms leading to activity of additional pro-survival genes or signalling pathways that function independently of JAK2/STAT5. To identify alternative therapeutic targets, we applied a pooled-shRNA library targeting ~5000 genes to a JAK2V617F-positive cell line under a variety of conditions, including absence or presence of ruxolitinib and in the presence of a bone marrow microenvironment-like culture medium. We identified several proteasomal gene family members as essential to HEL cell survival. The importance of these genes was validated in MF cells using the proteasomal inhibitor carfilzomib, which also enhanced lethality in combination with ruxolitinib. We also showed that proteasome gene expression is reduced by ruxolitinib in MF CD34+ cells and that additional targeting of proteasomal activity by carfilzomib enhances the inhibitory action of ruxolitinib in vitro. Hence, this study suggests a potential role for proteasome inhibitors in combination with ruxolitinib for management of MF patients.

Interoperable genetic lab test reports: mapping key data elements to HL7 FHIR specifications and professional reporting guidelines.

OBJECTIVE: In many cases, genetic testing labs provide their test reports as portable document format files or scanned images, which limits the availability of the contained information to advanced informatics solutions, such as automated clinical decision support systems. One of the promising standards that aims to address this limitation is Health Level Seven International (HL7) Fast Healthcare Interoperability Resources Clinical Genomics Implementation Guide-Release 1 (FHIR CG IG STU1). This study aims to identify various data content of some genetic lab test reports and map them to FHIR CG IG specification to assess its coverage and to provide some suggestions for standard development and implementation. MATERIALS AND METHODS: We analyzed sample reports of 4 genetic tests and relevant professional reporting guidelines to identify their key data elements (KDEs) that were then mapped to FHIR CG IG. RESULTS: We identified 36 common KDEs among the analyzed genetic test reports, in addition to other unique KDEs for each genetic test. Relevant suggestions were made to guide the standard implementation and development. DISCUSSION AND CONCLUSION: The FHIR CG IG covers the majority of the identified KDEs. However, we suggested some FHIR extensions that might better represent some KDEs. These extensions may be relevant to FHIR implementations or future FHIR updates.The FHIR CG IG is an excellent step toward the interoperability of genetic lab test reports. However, it is a work-in-progress that needs informative and continuous input from the clinical genetics' community, specifically professional organizations, systems implementers, and genetic knowledgebase providers.

A qualitative investigation of biomedical informatics interoperability standards for genetic test reporting: benefits, challenges, and motivations from the testing laboratory's perspective.

PURPOSE: Genetic laboratory test reports can often be of limited computational utility to the receiving clinical information systems, such as clinical decision support systems. Many health-care interoperability (HC) standards aim to tackle this problem, but the perceived benefits, challenges, and motivations for implementing HC interoperability standards from the labs' perspective has not been systematically assessed. METHODS: We surveyed genetic testing labs across the United States and conducted a semistructured interview with responding lab representatives. We conducted a thematic analysis of the interview transcripts to identify relevant themes. A panel of experts discussed and validated the identified themes. RESULTS: Nine labs participated in the interview, and 24 relevant themes were identified within five domains. These themes included the challenge of complex and changing genetic knowledge, the motivation of competitive advantage, provided financial incentives, and the benefit of supporting the learning health system. CONCLUSION: Our study identified the labs' perspective on various aspects of implementing HC interoperability standards in producing and communicating genetic test reports. Interviewees frequently reported that increased adoption of HC standards may be motivated by competition and programs incentivizing and regulating the incorporation of interoperability standards for genetic test data, which could benefit quality control, research, and other areas.

Association of Combined Focal 22q11.22 Deletion and IKZF1 Alterations With Outcomes in Childhood Acute Lymphoblastic Leukemia.

IMPORTANCE: Alterations in the IKZF1 gene drive B-cell acute lymphoblastic leukemia (B-ALL) but are not routinely used to stratify patients by risk because of inconsistent associations with outcomes. We describe a novel deletion in 22q11.22 that was consistently associated with very poor outcomes in patients with B-ALL with IKZF1 alterations. OBJECTIVE: To determine whether focal deletions within the λ variable chain region in chromosome 22q11.22 were associated with patients with B-ALL with IKZF1 alterations with the highest risk of relapse and/or death. DESIGN, SETTING, AND PARTICIPANTS: This cohort study included 1310 primarily high-risk pediatric patients with B-ALL who were taken from 6 independent clinical cohorts, consisting of 3 multicenter cohorts (AALL0232 [2004-2011], P9906 [2000-2003], and patients with Down syndrome who were pooled from national and international studies) and 3 single-institution cohorts (University of Utah [Salt Lake City], Children's Hospital of Philadelphia [Philadelphia, Pennsylvania], and St. Jude Children's Hospital [Memphis, Tennessee]). Data analysis began in 2011 using patients from the older studies first, and data analysis concluded in 2021. EXPOSURES: Focal 22q11.22 deletions. MAIN OUTCOMES AND MEASURES: Event-free and overall survival was investigated. The hypothesis that 22q11.22 deletions stratified the prognostic effect of IKZF1 alterations was formulated while investigating nearby deletions in VPREB1 in 2 initial cohorts (n = 270). Four additional cohorts were then obtained to further study this association (n = 1040). RESULTS: This study of 1310 patients with B-ALL (717 male [56.1%] and 562 female patients [43.9%]) found that focal 22q11.22 deletions are frequent (518 of 1310 [39.5%]) in B-ALL and inconsistent with physiologic V(D)J recombination. A total of 299 of 1310 patients with B-ALL had IKZF1 alterations. Among patients with IKZF1 alterations, more than half shared concomitant focal 22q11.22 deletions (159 of 299 [53.0%]). Patients with combined IKZF1 alterations and 22q11.22 deletions had worse outcomes compared with patients with IKZF1 alterations and wild-type 22q11.22 alleles in every cohort examined (combined cohorts: 5-year event-free survival rates, 43.3% vs 68.5%; hazard ratio [HR], 2.18; 95% CI, 1.54-3.07; P < .001; 5-year overall survival rates, 66.9% vs 83.9%; HR, 2.05; 95% CI, 1.32-3.21; P = .001). While 22q11.22 deletions were not prognostic in patients with wild-type IKZF1 , concomitant 22q11.22 deletions in patients with IKZF1 alterations stratified outcomes across additional risk groups, including patients who met the IKZF1plus criteria, and maintained independent significance in multivariate analysis for event-free survival (HR, 2.05; 95% CI, 1.27-3.29; P = .003) and overall survival (HR, 1.83; 95% CI, 1.01-3.34; P = .05). CONCLUSIONS AND RELEVANCE: This cohort study suggests that 22q11.22 deletions identify patients with B-ALL and IKZF1 alterations who have very poor outcomes and may offer a new genetic biomarker to further refine B-ALL risk stratification and treatment strategies.

A qualitative study of prevalent laboratory information systems and data communication patterns for genetic test reporting.

PURPOSE: The availability of genetic test data within the electronic health record (EHR) is a pillar of the US vision for an interoperable health IT infrastructure and a learning health system. Although EHRs have been highly investigated, evaluation of the information systems used by the genetic labs has received less attention-but is necessary for achieving optimal interoperability. This study aimed to characterize how US genetic testing labs handle their information processing tasks. METHODS: We followed a qualitative research method that included interviewing lab representatives and a panel discussion to characterize the information flow models. RESULTS: Ten labs participated in the study. We identified three generic lab system models and their relevant characteristics: a backbone system with additional specialized systems for interpreting genetic results, a brokering system that handles housekeeping and communication, and a single primary system for results interpretation and report generation. CONCLUSION: Labs have heterogeneous workflows and generally have a low adoption of standards when sending genetic test reports back to EHRs. Core interpretations are often delivered as free text, limiting their computational availability for clinical decision support tools. Increased provision of genetic test data in discrete and standard-based formats by labs will benefit individual and public health.

The clear cell sarcoma functional genomic landscape.

Clear cell sarcoma (CCS) is a deadly malignancy affecting adolescents and young adults. It is characterized by reciprocal translocations resulting in expression of the chimeric EWSR1-ATF1 or EWSR1-CREB1 fusion proteins, driving sarcomagenesis. Besides these characteristics, CCS has remained genomically uncharacterized. Copy number analysis of human CCSs showed frequent amplifications of the MITF locus and chromosomes 7 and 8. Few alterations were shared with Ewing sarcoma or desmoplastic, small round cell tumors, which are other EWSR1-rearranged tumors. Exome sequencing in mouse tumors generated by expression of EWSR1-ATF1 from the Rosa26 locus demonstrated no other repeated pathogenic variants. Additionally, we generated a new CCS mouse by Cre-loxP-induced chromosomal translocation between Ewsr1 and Atf1, resulting in copy number loss of chromosome 6 and chromosome 15 instability, including amplification of a portion syntenic to human chromosome 8, surrounding Myc. Additional experiments in the Rosa26 conditional model demonstrated that Mitf or Myc can contribute to sarcomagenesis. Copy number observations in human tumors and genetic experiments in mice rendered, for the first time to our knowledge, a functional landscape of the CCS genome. These data advance efforts to understand the biology of CCS using innovative models that will eventually allow us to validate preclinical therapies necessary to achieve longer and better survival for young patients with this disease.

Large-Scale Identification of Clonal Hematopoiesis and Mutations Recurrent in Blood Cancers.

Clonal hematopoiesis of indeterminate potential (CHIP) is characterized by detectable hematopoietic-associated gene mutations in a person without evidence of hematologic malignancy. We sought to identify additional cancer-presenting mutations useable for CHIP detection by performing a data mining analysis of 48 somatic mutation studies reporting mutations at diagnoses of 7,430 adult and pediatric patients with hematologic malignancies. Following extraction of 20,141 protein-altering mutations, we identified 434 significantly recurrent mutation hotspots, 364 of which occurred at loci confidently assessable for CHIP. We then performed an additional large-scale analysis of whole exome sequencing data from 4,538 persons belonging to three non-cancer cohorts for clonal mutations. We found the combined cohort prevalence of CHIP with mutations identical to those reported at blood cancer mutation hotspots to be 1.8%, and that some of these CHIP mutations occurred in children. Our findings may help to improve CHIP detection and pre-cancer surveillance for both children and adults.