Enigmatic missense mutations can cause disease via creation of de novo nuclear export signals.

Disease-causing missense mutations that occur within structurally and functionally unannotated protein regions can guide researchers to new mechanisms of protein regulation and dysfunction. Here, we report that the thrombocytopenia-, myelodysplastic syndromes-, and leukemia-associated P214L mutation in the transcriptional regulator ETV6 creates an XPO1-dependent nuclear export signal to cause protein mislocalization. Strategies to disrupt XPO1 activity fully restore ETV6 P214L protein nuclear localization and transcription regulation activity. Mechanistic insight inspired the design of a 'humanized' ETV6 mice, which we employ to demonstrate that the germline P214L mutation is sufficient to elicit severe defects in thrombopoiesis and hematopoietic stem cell maintenance. Beyond ETV6, we employed computational methods to uncover rare disease-associated missense mutations in unrelated proteins that create a nuclear export signal to disrupt protein function. Thus, missense mutations that operate through this mechanism should be predictable and may suggest rational therapeutic strategies for associated diseases.

Update on chromium speciation analysis in foods: a review of advances in analytical methods and dietary exposure assessment.

Chromium occurs naturally in different oxidation states. Amongst them, hexavalent chromium is classified as both genotoxic and carcinogenic while trivalent chromium can be considered as an essential element. Therefore, speciation analysis is essential when conducting dietary exposure assessment. Several critical reviews have been published on chromium speciation analysis in foodstuffs in the last decade. However, a method that can account for species interconversion during the extraction procedure has not been reported in the reviews. In recent years, an online method using species-specific isotope dilution mass spectrometry has been developed for the simultaneous determination of trivalent and hexavalent chromium in foodstuffs. Apart from that, new methods based on offline analytical techniques, to analyse trivalent and hexavalent chromium separately, are still under development. Therefore, one of the objectives of this paper is to review these recently published analytical methods and assess whether they are fit for chromium speciation analysis in foodstuffs. Additionally, an objective is also to assess whether their limits of detection are sufficiently low for dietary exposure assessment with respect to the neoplastic effects of hexavalent chromium. Moreover, possible future research gaps are identified based on the current knowledge and existing literature.

Racial and ethnic disparities in Acute Myeloid Leukemia: 15-year experience at a safety net hospital system.

Despite recent therapeutic advances, ethnic minorities in the U.S. continue to have disproportionately poor outcomes in many hematologic malignancies including AML. We identified 162 adult AML patients treated at a non-transplant safety net hospital from 2007 to 2022 and evaluated differences in disease characteristics, treatment and clinical outcomes based on race and ethnicity. Our cohort consisted of 82 (50.6%) Hispanic, 36 (22.2%) non-Hispanic black and 44 (27.2%) non-Hispanic white and Asian patients. Median age at diagnosis was 42.5, 49.0 and 52.5 years respectively (p=0.025). Hispanics had higher rates of intermediate and high-risk disease (p=0.699) and received high intensity induction and consolidation chemotherapy at lower rates (p=0.962), although differences did not reach statistical significance. Despite this, similar remission rates were achieved. Hispanics with high-risk disease had longer overall survival (OS) than the combined non-Hispanic cohort (mOS 14 m vs 7 m, p=0.030). Multivariate regression analysis showed that OS was negatively associated with age (HR 1.023, p=0.006), intermediate (HR 3.431, p=0.0003) and high-risk disease (HR 4.689, p<0.0001) and positively associated with Hispanic ethnicity (HR 0.614, p=0.026). This report suggests that contrary to other studies, Hispanics, particularly those with high-risk AML, may have improved OS compared to other ethnic groups. These results are unique to our safety net hospital setting where common barriers to medical care and healthcare disparities are largely mitigated.

Genomic heterogeneity within B/T mixed phenotype acute leukemia in a context of an immunophenotype.

B/T mixed phenotype acute leukemia (MPAL) is a rare aggressive leukemia. Three cases of B/T MPAL were identified with comprehensive immunophenotypic, cytogenetic, and molecular studies. T-lineage predominant B/T MPAL shares a genetic signature with T-ALL whereas B/T lineage co-dominant B/T MPAL lacks such a T-ALL signature. All three patients were treated with lineage-matched-ALL therapy and alive at the last follow-up. Our study is the first to demonstrate molecular heterogeneity within B/T MPAL in a context of an immunophenotype of T-lineage versus B-lineage predominance. The implication of such a phenotype-genotype association on diagnostic classification is briefly discussed.

Identifying patients at risk for hereditary myeloid malignancy syndromes incorporating a novel, self-administered questionnaire to an initial screening platform.

INTRODUCTION: Four to 10% of cases of myeloid malignancies are inherited. We report our experience on hereditary myeloid malignancy syndromes (HMMS) incorporating a novel questionnaire in the screening platform for patients with myeloid malignancies and aplastic anemia. METHODS: The questionnaire was sent via electronic patient portal prior to clinic visits. Patients screened positive based on responses to questionnaire items, presence of suspicion disease characteristics (young age, family history, monosomy 7 etc.) and/or presence of signs of HMMS. Those deemed at-risk based on questionnaire responses, clinical features and/or somatic mutation profile were offered germline testing. RESULTS: A total of 408 patients were screened, 141 (35%) were deemed at-risk. Fifty-four (38%) of at-risk patients were seen in the genetics clinic. Forty-one (76%) of the patients seen agreed to germline testing and 13 declined due to cost or personal decision. Twenty pathogenic (P)/likely-pathogenic (LP) germline mutations were identified in 16 (39%) of the tested patients. Five patients also had a variant of uncertain significance (VUS) and an additional 13 had at least 1 VUS without P/LP mutations (total 29 VUS's were found in 18 (44%) of tested patients). The median age of diagnosis for patients with P/LP mutations was 56 years versus 66 years in the entire cohort. CONCLUSION: Incorporating an electronic questionnaire is an effective screening method for HMMS. Many patients declined testing due to cost. These results highlight the importance of germline testing in patients with myeloid malignancies, further research in HMMS, and coverage by healthcare plans.

SARS-CoV-2 Vaccination IgG Antibody Responses in Patients with Hematologic Malignancies in a Myeloid Enriched Cohort: A Single Center Observation.

OBJECTIVE: Patients diagnosed with hematologic malignancies are at increased risk for severe SARS-CoV-2 infection. We evaluated the serological IgG response following two doses of the SARS-CoV-2 vaccine in patients with hematologic malignancies. METHODS: Patients treated at UT Southwestern Medical Center with a diagnosis of a myeloid or lymphoid neoplasm were included. SARS-CoV-2 vaccination response was defined as a positive quantifiable spike IgG antibody titer. RESULTS: Sixty patients were included in the study and 60% were diagnosed with a myeloid neoplasm. The majority (85%) of the patients with a myeloid malignancy and 50% of the patients with a lymphoid malignancy mounted a serological response after receiving two doses of the vaccine. CONCLUSION: Vaccination should be offered irrespective of ongoing treatment or active disease. Findings require validation in a larger cohort of patients.

Ferrofluid drop impacts and Rosensweig peak formation in a non-uniform magnetic field.

Vertical drop impacts of ferrofluids onto glass slides in a non-uniform magnetic field have been studied using high-speed photography. Outcomes have been classified based on the motion of the fluid-surface contact lines, and formation of peaks (Rosensweig instabilities) which affect the height of the spreading drop. The largest peaks are nucleated at the edge of a spreading drop, similarly to crown-rim instabilities in drop impacts with conventional fluids, and remain there for an extended time. Impact Weber numbers ranged from 18.0 to 489, and the vertical component of the B-field was varied between 0 and 0.37 T at the surface by changing the vertical position of a simple disc magnet placed below the surface. The falling drop was aligned with the vertical cylindrical axis of the 25 mm diameter magnet, and the impacts produced Rosensweig instabilities without splashing. At high magnetic flux densities a stationary ring of ferrofluid forms approximately above the outer edge of the magnet.

Neutrophil metabolomics in severe COVID-19 reveal GAPDH as a suppressor of neutrophil extracellular trap formation.

Severe COVID-19 is characterized by an increase in the number and changes in the function of innate immune cells including neutrophils. However, it is not known how the metabolome of immune cells changes in patients with COVID-19. To address these questions, we analyzed the metabolome of neutrophils from patients with severe or mild COVID-19 and healthy controls. We identified widespread dysregulation of neutrophil metabolism with disease progression including in amino acid, redox, and central carbon metabolism. Metabolic changes in neutrophils from patients with severe COVID-19 were consistent with reduced activity of the glycolytic enzyme GAPDH. Inhibition of GAPDH blocked glycolysis and promoted pentose phosphate pathway activity but blunted the neutrophil respiratory burst. Inhibition of GAPDH was sufficient to cause neutrophil extracellular trap (NET) formation which required neutrophil elastase activity. GAPDH inhibition increased neutrophil pH, and blocking this increase prevented cell death and NET formation. These findings indicate that neutrophils in severe COVID-19 have an aberrant metabolism which can contribute to their dysfunction. Our work also shows that NET formation, a pathogenic feature of many inflammatory diseases, is actively suppressed in neutrophils by a cell-intrinsic mechanism controlled by GAPDH.

Post-transplant cyclophosphamide and early mixed donor Chimerism in myeloid malignancies; a single-center experience.

BACKGROUND: Allogeneic hematopoietic stem cell transplantation (allo-HSCT) remains the only curative option for high-risk myeloid malignancies. Post-transplant cyclophosphamide (PT-Cy) has proven to be effective for graft versus host disease (GVHD) prophylaxis. Given that graft-versus-tumor (GVT) effect plays a major role in reducing the risk of disease relapse, the application of PT-Cy must balance the risk of relapse. Mixed chimerism (MC) refers to a state of concurrent presence of recipient and donor cells post allo-HSCT which may precede relapse disease. OBJECTIVE: We investigated the impact of PT-Cy on early MC (EMC) and disease relapse in patients with a myeloid malignancy post allo-HSCT. STUDY DESIGN: This retrospective single-center study included patients that underwent allo-HSCT between 2015 and 2021. Patient and disease characteristics were collected from the electronic health records. EMC was defined as <95% donor cells at day 90-120 post allo-HSCT. RESULTS: A total of 144 patient that received an allo-HSCT were included in the study. One hundred and eight (75%) patients received PT-Cy as part of the GVHD prophylaxis regimen. The majority underwent allo-HSCT for acute myeloid leukemia (62%) or myelodysplastic syndrome (31%). Sixty-five percent received allo-HSCT from a matched unrelated donor transplant and 65% received a myeloablative conditioning regimen. A lower rate of chronic GVHD (p = 0.03) and a higher rate of EMC (p = 0.04) were observed in patients that received PT-Cy. PT-Cy was not associated with overall survival (OS) and relapse-free survival (RFS). Multivariable analysis identified measurable residual disease status (p = 0.003), hematopoietic cell transplantation-specific comorbidity index (p = 0.012) and chronic GVHD (p = 0.006) as independent prognostic variables for OS. AML-adverse risk (p = 0.004) and EMC (p = 0.018) were independently prognostic for RFS. While EMC overall was not significantly associated with higher risk of relapse, EMC was associated with shorter RFS within adverse-risk AML patients. CONCLUSION: Our study shows that PT-Cy was associated with an increased risk of EMC. The predictive value of EMC for relapse remains unclear and may depend on the underlying disease, which should be validated in a larger cohort.

Clinical and molecular characteristics associated with Vitamin C deficiency in myeloid malignancies; real world data from a prospective cohort.

Vitamin C is an essential vitamin that acts as a co-factor for many enzymes involved in epigenetic regulation in humans. Low vitamin C levels in hematopoietic stem cells (HSC) promote self-renewal and vitamin C supplementation retards leukaemogenesis in vitamin C-deficient mouse models. Studies on vitamin C levels in patients with myeloid malignancies are limited. We thus conducted a retrospective analysis on a prospective cohort of patients with myeloid malignancies on whom plasma vitamin C levels were measured serially at diagnosis and during treatment. Baseline characteristics including hematological indices, cytogenetics, and molecular mutations are described in this cohort. Among 64 patients included in our study, 11 patients (17%) had low vitamin C levels. We noted a younger age at diagnosis for patients with myeloid malignancies who had low plasma vitamin C levels. Patients with low plasma vitamin C levels were more likely to have acute myeloid leukemia compared to other myeloid malignancies. Low vitamin C levels were associated with ASXL1 mutations. Our study calls for further multi-institutional studies to understand the relevance of low plasma vitamin C level in myeloid neoplasms, the role of vitamin C deficiency in leukemogenesis, and the potential benefit of vitamin C supplementation.

Disabling Uncompetitive Inhibition of Oncogenic IDH Mutations Drives Acquired Resistance.

Mutations in IDH genes occur frequently in acute myeloid leukemia (AML) and other human cancers to generate the oncometabolite R-2HG. Allosteric inhibition of mutant IDH suppresses R-2HG production in a subset of patients with AML; however, acquired resistance emerges as a new challenge, and the underlying mechanisms remain incompletely understood. Here we establish isogenic leukemia cells containing common IDH oncogenic mutations by CRISPR base editing. By mutational scanning of IDH single amino acid variants in base-edited cells, we describe a repertoire of IDH second-site mutations responsible for therapy resistance through disabling uncompetitive enzyme inhibition. Recurrent mutations at NADPH binding sites within IDH heterodimers act in cis or trans to prevent the formation of stable enzyme-inhibitor complexes, restore R-2HG production in the presence of inhibitors, and drive therapy resistance in IDH-mutant AML cells and patients. We therefore uncover a new class of pathogenic mutations and mechanisms for acquired resistance to targeted cancer therapies. SIGNIFICANCE: Comprehensive scanning of IDH single amino acid variants in base-edited leukemia cells uncovers recurrent mutations conferring resistance to IDH inhibition through disabling NADPH-dependent uncompetitive inhibition. Together with targeted sequencing, structural, and functional studies, we identify a new class of pathogenic mutations and mechanisms for acquired resistance to IDH-targeting cancer therapies. This article is highlighted in the In This Issue feature, p. 1.

Epo-IGF1R cross talk expands stress-specific progenitors in regenerative erythropoiesis and myeloproliferative neoplasm.

We found that in regenerative erythropoiesis, the erythroid progenitor landscape is reshaped, and a previously undescribed progenitor population with colony-forming unit-erythroid (CFU-E) activity (stress CFU-E [sCFU-E]) is expanded markedly to restore the erythron. sCFU-E cells are targets of erythropoietin (Epo), and sCFU-E expansion requires signaling from the Epo receptor (EpoR) cytoplasmic tyrosines. Molecularly, Epo promotes sCFU-E expansion via JAK2- and STAT5-dependent expression of IRS2, thus engaging the progrowth signaling from the IGF1 receptor (IGF1R). Inhibition of IGF1R and IRS2 signaling impairs sCFU-E cell growth, whereas exogenous IRS2 expression rescues cell growth in sCFU-E expressing truncated EpoR-lacking cytoplasmic tyrosines. This sCFU-E pathway is the major pathway involved in erythrocytosis driven by the oncogenic JAK2 mutant JAK2(V617F) in myeloproliferative neoplasm. Inability to expand sCFU-E cells by truncated EpoR protects against JAK2(V617F)-driven erythrocytosis. In samples from patients with myeloproliferative neoplasm, the number of sCFU-E-like cells increases, and inhibition of IGR1R and IRS2 signaling blocks Epo-hypersensitive erythroid cell colony formation. In summary, we identified a new stress-specific erythroid progenitor cell population that links regenerative erythropoiesis to pathogenic erythrocytosis.

Outcomes of liver transplant recipients with high MELD scores: an experience from a Canadian centre.

BACKGROUND: The frequency with which patients with high Model for End-Stage Liver Disease (MELD) scores undergo liver transplantation has been increasing. Canadian literature regarding the outcomes of liver transplantation in recipients with high MELD scores is limited. The primary objective of this study was to assess patient and graft survival among recipients with high (> 35) and low (≤ 35) MELD scores. Secondary objectives were to potentially identify independent predictors of graft failure and patient mortality. METHODS: We conducted a retrospective chart review of patients undergoing liver transplantation at a single Canadian centre from 2012 to 2017. RESULTS: A total of 332 patients were included in the study: 280 patients had a MELD score of 35 or lower, and 52 had a MELD score above 35. Patients with high MELD scores had higher rates of pretransplant acute kidney injury and dialysis (p < 0.001), admission to the intensive care unit (ICU) or intubation (p < 0.001), intraoperative blood product transfusions (p < 0.001) and post-transplantation acute kidney injury and dialysis (p < 0.001), as well as longer ICU (p < 0.001) and hospital stays (p = 0.002). One- and 3-year patient survival in recipients with MELD scores of 35 or lower was 93.1% and 84.9% versus 85.0% and 80.0% in recipients with MELD scores above 35 (p = 0.37). One- and 3-year graft survival in recipients with MELD scores of 35 or lower was 91.7% and 90.9% versus 77.2% and 72.8% in recipients with MELD scores above 35 (p < 0.001). Prior liver transplant was an independent predictor of patient mortality, and no independent predictors of graft failure were identified. When MELD was replaced with D-MELD (donor age × recipient MELD), it predicted graft failure but not patient survival. CONCLUSION: No difference in patient mortality was found between MELD groups. Graft survival was significantly lower in recipients with MELD scores above 35. D-MELD may potentially be used as an adjunct in determining risk of graft failure in recipients with high MELD scores.

Chromatographic analysis of dithiocarbamate residues and their metabolites in foods employed in dietary exposure studies-a review.

Dithiocarbamates (DTCs) belong to a group of compounds used as fungicides in food production and can be divided into three major groups. Since DTCs easily oxidise and hydrolyse in alkaline and acidic medium respectively, precautions have to be implemented during preparation/homogenisation and extraction of samples. As such, test samples are commonly prepared individually by cutting into small pieces just before the digestion of DTCs with a hot acid to give carbon disulphide (CS2) and the results are expressed as CS2 without any differentiation of individual DTCs. However, individual DTCs have different toxicological potencies whilst their metabolites are more toxic than the parent compound. Apart from the hot digestion method, chromatographic separation of three major groups of DTCs has been developed by a number of different researchers. This review provides a comprehensive examination of sample preparation, extraction, clean-up and chromatographic methods for the determination of individual DTCs and their more toxic metabolites in foodstuffs. Moreover, this review also studies on how dietary exposure of DTCs can be efficiently and effectively estimated using different methods of analysis.

Pancreas tail adenocarcinoma tumor size is highly predictive of positive margins after a nonradical resection.

BACKGROUND: Radical resection (RAMPS) of left sided pancreatic ductal adenocarcinoma (PDAC) is effective in achieving R0 margins; however, not universally accepted due to lack of improved survival. We hypothesized that only larger tumors lead to R1 in non-RAMPS procedures. METHODS: A retrospective review of charts between 2008 and 2020 was performed. The primary outcome was evaluating R0 resection based on left-sided tumors' size and location, and secondary outcomes were OS and DFS. RESULTS: Sixty-eight percent had R0 resection. R1 groups' tumors were larger (5.5 cm vs. 3.8 cm, p = 0.004) and had higher LVI involvement (p = 0.003). OS and DFS did not differ on multivariate analysis. Tumor size above 4 cm in the tail was associated with R1 (p = 0.01). CONCLUSIONS: Larger tumors in the tail, but not body were associated with R1, but not worse survival. Perhaps larger tumors in the tail are a surrogate marker of poor disease biology.

The impact of race and ethnicity on outcomes of patients with myelodysplastic syndromes: a population-based analysis.

Race and ethnic backgrounds affect the disease characteristics and clinical outcomes in many cancers, including acute myeloid leukemia; however, the association of race/ethnicity on myelodysplastic syndrome (MDS) is still controversial. Therefore, we aimed to study the impact of race/ethnicity on the disease characteristics and survival outcomes in patients with MDS. Adult patients with MDS diagnosed in 2004-2016 were selected using the SEER database. Race/ethnicity was categorized as non-Hispanic White (NHW), non-Hispanic Black (NHB), and Hispanic. Hispanic and NHB patients had significantly lower incidence rate ratio (IRR) in age group ≥01 years (p < .001) compared to NHW; however, in the age group <50 years, NHB patients had significantly higher IRR with an increased incidence rate of 49%. NHB patients had better overall survival than Hispanic and NHW patients (p < .001), even after adjusting for confounding variables. MDS have significant differences in age at diagnosis, disease risk, and survival outcomes based on racial/ethnic backgrounds.

The association between bacterobilia and the risk of postoperative complications following pancreaticoduodenectomy.

BACKGROUND: Pre-operative biliary bacterial colonisation (bacterobilia) is considered a risk factor for infectious complications after pancreaticoduodenectomy (PD). This study aimed to investigate the role of the PD biliary microbiome grown in the development of post-PD complications. METHODS: In a retrospective study of 162 consecutive patients undergoing PD (2008-2018), intra-operative bile cultures were analyzed and sensitivities compared to pre-anesthetic antibiotics and thirty-day post-surgery complications. RESULTS: Bacterobilia was present in 136 patients (84%). Most bile cultures grew bacteria resistant to pre-operative antibiotics (n = 112, 82%). Patients with bacterobilia had significantly higher rates of major complication than patients without (P = 0.017), as well as higher rates of surgical-site infections (SSI) (P = 0.010). Patients with negative bile cultures (n = 26) had significantly lower rates of major complication and SSI than those growing sensitive (n = 24) or non-sensitive (n = 112) bacteria (major complication P = 0.029 and SSI P = 0.011). CONCLUSION: Positive bile cultures were associated with a higher incidence of major complications and SSI. Patients with sterile bile cultures had the lowest risk of post-operative complications and efforts to reduce rates of bacterobilia, such as limitation of biliary instrumentation, should be considered. Sensitivity to antibiotics had no effect upon the rate of post-operative complications, but this may reflect low cohort numbers.

Convergence of oncogenic cooperation at single-cell and single-gene levels drives leukemic transformation.

Cancers develop from the accumulation of somatic mutations, yet it remains unclear how oncogenic lesions cooperate to drive cancer progression. Using a mouse model harboring NRasG12D and EZH2 mutations that recapitulates leukemic progression, we employ single-cell transcriptomic profiling to map cellular composition and gene expression alterations in healthy or diseased bone marrows during leukemogenesis. At cellular level, NRasG12D induces myeloid lineage-biased differentiation and EZH2-deficiency impairs myeloid cell maturation, whereas they cooperate to promote myeloid neoplasms with dysregulated transcriptional programs. At gene level, NRasG12D and EZH2-deficiency independently and synergistically deregulate gene expression. We integrate results from histopathology, leukemia repopulation, and leukemia-initiating cell assays to validate transcriptome-based cellular profiles. We use this resource to relate developmental hierarchies to leukemia phenotypes, evaluate oncogenic cooperation at single-cell and single-gene levels, and identify GEM as a regulator of leukemia-initiating cells. Our studies establish an integrative approach to deconvolute cancer evolution at single-cell resolution in vivo.

End stage liver disease etiology & transplantation referral outcomes of major ethnic groups in British Columbia, Canada: A cohort study.

ABSTRACT: Liver disease etiology and transplantation outcomes may vary by ethnicity. We aimed to determine if disparities exist in our province.We reviewed the provincial database for liver transplant referrals. We stratified cohorts by ethnicity and analyzed disease etiology and outcomes.Four thousand nine hundred sixteen referrals included 220 South Asians, 413 Asians, 235 First Nations (Indigenous), and 2725 Caucasians. Predominant etiologies by ethnicity included alcohol (27.4%) and primary sclerosing cholangitis (PSC) (8.8%) in South Asians, hepatitis B (45.5%) and malignancy (13.9%) in Asians, primary biliary cholangitis (PBC) (33.2%) and autoimmune hepatitis (AIH) (10.8%) in First Nations, and hepatitis C (35.9%) in Caucasians. First Nations had lowest rate of transplantation (30.6%, P = .01) and highest rate of waitlist death (10.6%, P = .03). Median time from referral to transplantation (268 days) did not differ between ethnicities (P = .47). Likelihood of transplantation increased with lower body mass index (BMI) (hazard ratio [HR] 0.99, P = .03), higher model for end stage liver disease (MELD) (HR 1.02, P < .01), or fulminant liver failure (HR 9.47, P < .01). Median time from referral to ineligibility status was 170 days, and shorter time was associated with increased MELD (HR 1.01, P < .01), increased age (HR 1.01, P < .01), fulminant liver failure (HR 2.56, P < .01) or South Asian ethnicity (HR 2.54, P < .01). Competing risks analysis revealed no differences in time to transplant (P = .66) or time to ineligibility (P = .91) but confirmed increased waitlist death for First Nations (P = .04).We have noted emerging trends such as alcohol related liver disease and PSC in South Asians. First Nations have increased autoimmune liver disease, lower transplantation rates and higher waitlist deaths. These data have significance for designing ethnicity specific interventions.