A practical approach to curate clonal hematopoiesis of indeterminate potential in human genetic data sets.

Clonal hematopoiesis of indeterminate potential (CHIP) is a common form of age-related somatic mosaicism that is associated with significant morbidity and mortality. CHIP mutations can be identified in peripheral blood samples that are sequenced using approaches that cover the whole genome, the whole exome, or targeted genetic regions; however, differentiating true CHIP mutations from sequencing artifacts and germ line variants is a considerable bioinformatic challenge. We present a stepwise method that combines filtering based on sequencing metrics, variant annotation, and population-based associations to increase the accuracy of CHIP calls. We apply this approach to ascertain CHIP in ∼550 000 individuals in the UK Biobank complete whole exome cohort and the All of Us Research Program initial whole genome release cohort. CHIP ascertainment on this scale unmasks recurrent artifactual variants and highlights the importance of specialized filtering approaches for several genes, including TET2 and ASXL1. We show how small changes in filtering parameters can considerably increase CHIP misclassification and reduce the effect size of epidemiological associations. Our high-fidelity call set refines previous population-based associations of CHIP with incident outcomes. For example, the annualized incidence of myeloid malignancy in individuals with small CHIP clones is 0.03% per year, which increases to 0.5% per year among individuals with very large CHIP clones. We also find a significantly lower prevalence of CHIP in individuals of self-reported Latino or Hispanic ethnicity in All of Us, highlighting the importance of including diverse populations. The standardization of CHIP calling will increase the fidelity of CHIP epidemiological work and is required for clinical CHIP diagnostic assays.

COVID-19 and the Genetics of Inflammation.

OBJECTIVE: Interindividual variability in the clinical progression of COVID-19 may be explained by host genetics. Emerging literature supports a potential inherited predisposition to severe forms of COVID-19. Demographic and inflammatory characteristics of COVID-19 suggest that acquired hematologic mutations leading to clonal hematopoiesis (CH) may further increase vulnerability to adverse sequelae. This review summarizes the available literature examining genetic predispositions to severe COVID-19 and describes how these findings could eventually be used to improve its clinical management. DATA SOURCES: A PubMed literature search was performed. STUDY SELECTION: Studies examining the significance of inherited genetic variation or acquired CH mutations in severe COVID-19 were selected for inclusion. DATA EXTRACTION: Relevant genetic association data and aspects of study design were qualitatively assessed and narratively synthesized. DATA SYNTHESIS: Genetic variants affecting inflammatory responses may increase susceptibility to severe COVID-19. Genome-wide association studies and candidate gene approaches have identified a list of inherited mutations, which likely alter cytokine and interferon secretion, and lung-specific mechanisms of immunity in COVID-19. The potential role of CH in COVID-19 is more uncertain at present; however, the available evidence suggests that the various types of acquired mutations and their differential influence on immune cell function must be carefully considered. CONCLUSIONS: The current literature supports the hypothesis that host genetic factors affect vulnerability to severe COVID-19. Further research is required to confirm the full scope of relevant variants and the causal mechanisms underlying these associations. Clinical approaches, which consider the genetic basis of interindividual variability in COVID-19 and potentially other causes of critical illness, could optimize hospital resource allocation, predict responsiveness to treatment, identify more efficacious drug targets, and ultimately improve outcomes.

A Lower Frequency of Spliceosome Mutations Distinguishes Clonal Cytopenias of Undetermined Significance From Low-Risk Myelodysplastic Syndromes, Despite Inherent Similarities in Genomic, Laboratory, and Clinical Features.

Clonal cytopenias of undetermined significance (CCUS) are associated with an increased risk of developing a myelodysplastic syndrome (MDS); however, the mechanism and factors associated with evolution remain unclear. We propose that next-generation sequencing (NGS) of cytopenic cases with equivocal morphologic dysplasia will improve patient clinical care and that serial sequencing of such equivocal cases could identify the factors that predict evolution to MDS. We performed targeted NGS of samples from 193 individuals with confirmed or suspected MDS or MDS/myeloproliferative neoplasm, including sequential investigation for 28 individuals at the time of diagnosis and during follow-up. NGS facilitated the diagnosis of all suspicious cases as myeloid neoplasm (21%), CCUS (34%), or idiopathic cytopenias of undetermined significance (45%) when no variants were detected. We found that there was no significant difference in most measured clinical features or clonal phenotypes, such as cell counts, number of variants, variant allele frequencies, and overall survival, between CCUS and International Prognostic Scoring System-Revised-defined low-risk MDS. However, there was a significant difference in the types of variants between CCUS and low-risk MDS, with a significantly lower number of splicing factor mutations in CCUS cases (P < .001). Moreover, we observed an increased probability of evolution to MDS of individuals with CCUS compared with that in those with idiopathic cytopenias of undetermined significance over the first 5 years (P = .045). Our analyses revealed no conclusive pattern associating clonal expansion or the number of variants with the evolution of CCUS to MDS, perhaps further supporting the similarity of these diseases and the clinical importance of recognizing and formally defining CCUS as a category of precursor myeloid disease state in the next revision of the World Health Organization guidelines.

Clonal Hematopoiesis of Indeterminate Potential and Kidney Function Decline in the General Population.

RATIONALE & OBJECTIVE: Clonal hematopoiesis of indeterminate potential (CHIP), defined by the age-related ontogenesis of expanded leukemogenic variants indicative of a genetically distinct clonal leukocyte population, is associated with risk of hematologic malignancy and cardiovascular disease. In experimental models, recapitulation of CHIP promotes kidney interstitial fibrosis with direct tissue infiltration of donor macrophages. We tested the hypothesis that CHIP is associated with kidney function decline in the general population. STUDY DESIGN: Cohort study. SETTING & PARTICIPANTS: 12,004 individuals from 3 community-based cohorts in the TOPMed Consortium. EXPOSURE: CHIP status from whole-genome sequences obtained from DNA extracted from peripheral blood. OUTCOME: Risk of 30% decline in estimated glomerular filtration rate (eGFR) and percent eGFR decline per year during the follow-up period. ANALYTICAL APPROACH: Cox proportional hazards models for 30% eGFR decline end point and generalized estimating equations for annualized relative change in eGFR with meta-analysis. Study-specific estimates were combined using fixed-effect meta-analysis. RESULTS: The median baseline eGFR was 84mL/min/1.73m2. The prevalence of CHIP was 6.6%, 9.0%, and 12.2% in persons aged 50-60, 60-70, and>70 years, respectively. Over a median follow-up period of 8 years, for the 30% eGFR outcome 205 events occurred among 1,002 CHIP carriers (2.1 events per 100 person-years) and 2,041 events in persons without CHIP (1.7 events per 100 person-years). In meta-analysis, CHIP was associated with greater risk of a 30% eGFR decline (17% [95% CI, 1%-36%] higher; P=0.04). Differences were not observed between those with baseline eGFR above or below 60mL/min/1.73m2, of age above or below 60 years, or with or without diabetes. LIMITATIONS: Small number of participants with moderate-to-advanced kidney disease and restricted set of CHIP driver variants. CONCLUSIONS: We report an association between CHIP and eGFR decline in 3 general population cohorts without known kidney disease. Further studies are needed to investigate this novel condition and its potential impact among individuals with overt kidney disease.

Association of Clonal Hematopoiesis of Indeterminate Potential with Worse Kidney Function and Anemia in Two Cohorts of Patients with Advanced Chronic Kidney Disease.

BACKGROUND: Clonal hematopoiesis of indeterminate potential (CHIP) is an inflammatory premalignant disorder resulting from acquired genetic mutations in hematopoietic stem cells. This condition is common in aging populations and associated with cardiovascular morbidity and overall mortality, but its role in CKD is unknown. METHODS: We performed targeted sequencing to detect CHIP mutations in two independent cohorts of 87 and 85 adults with an eGFR<60 ml/min per 1.73m2. We also assessed kidney function, hematologic, and mineral bone disease parameters cross-sectionally at baseline, and collected creatinine measurements over the following 5-year period. RESULTS: At baseline, CHIP was detected in 18 of 87 (21%) and 25 of 85 (29%) cohort participants. Participants with CHIP were at higher risk of kidney failure, as predicted by the Kidney Failure Risk Equation (KFRE), compared with those without CHIP. Individuals with CHIP manifested a 2.2-fold increased risk of a 50% decline in eGFR or ESKD over 5 years of follow-up (hazard ratio 2.2; 95% confidence interval, 1.2 to 3.8) in a Cox proportional hazard model adjusted for age, sex, and baseline eGFR. The addition of CHIP to 2-year and 5-year calibrated KFRE risk models improved ESKD predictions. Those with CHIP also had lower hemoglobin, higher ferritin, and higher red blood cell mean corpuscular volume versus those without CHIP. CONCLUSIONS: In this exploratory analysis of individuals with preexisting CKD, CHIP was associated with higher baseline KFRE scores, greater progression of CKD, and anemia. Further research is needed to define the nature of the relationship between CHIP and kidney disease progression.

Isolated Arthroscopic Partial Meniscectomy Is More Effective at Improving Meniscal Symptoms in Comparison With Mechanical Symptoms in Patients With Concomitant Untreated Chondral Lesions.

PURPOSE: To rank Knee Injury and Osteoarthritis Outcome Score (KOOS) questions from most to least improvement after arthroscopic partial meniscectomy (APM) and compare improvement of meniscal versus mechanical symptoms. METHODS: A secondary analysis of the Chondral Lesions and Meniscus Procedures (ChAMP) Trial was performed. Inclusion criteria were age 30 years or older with degenerative meniscal tear failing nonoperative management, with or without associated unstable chondral lesions. No chondral debridement was performed. Responses to the 42 KOOS questions ranged from 0 (extreme problems) to 4 (no problems), and were answered preoperatively and at 1 year after isolated APM. The 1-year mean change, or delta (Δ), was calculated for each KOOS question and the Δ for meniscal and mechanical symptoms were statistically compared. RESULTS: Greatest improvement in 135 eligible patients was observed for questions about (1) awareness of knee problems (Δ = 1.93, standard deviation [SD] = 1.38), (2) frequency of knee pain (Δ = 1.93, SD = 1.29), (3) degree of difficulty while twisting/pivoting on the injured knee (Δ = 1.88, SD = 1.13), (4) degree of difficulty while running (Δ = 1.67, SD = 1.30), and (5) being troubled by lack of confidence in the knee (Δ = 21.67, SD = 1.11). Least improvement was observed for questions about: (1) degree of difficulty while getting on/off the toilet (Δ = 0.94, SD = 0.96), (2) feel grinding or hear clicking when the knee moves (Δ= 0.90, SD = 1.25), 3) degree of difficulty while getting in/out of the bath (Δ= 0.88, SD = 1.00), (4) knee catches/hangs up during movement (Δ= 0.80, SD = 1.09), and (5) the ability to straighten the knee fully (Δ= 0.54, 1.44). There was greater improvement for the KOOS questions pertaining to meniscal versus mechanical symptoms (P < .00001). CONCLUSIONS: KOOS symptoms as reported by subjects' responses to the questions pertaining to the frequency of knee pain, twisting/pivoting, running, squatting, and jumping showed the most improvement 1 year after isolated APM, whereas those relating to mechanical symptoms improved the least. Focusing on meniscal rather than mechanical symptoms may help surgeons better identify patients expected to benefit from APM. LEVEL OF EVIDENCE: IV, retrospective analysis of prospectively collected data.

Better Outcomes but No Difference in Joint Space Narrowing at Five Years Among Patients Without Unstable Chondral Lesions Versus Those With Unstable Chondral Lesions (Left In Situ) at the Time of Arthroscopic Partial Meniscectomy.

PURPOSE: To compare 5-year outcomes among patients with and without unstable chondral lesions undergoing arthroscopic partial meniscectomy (APM). METHODS: Using data from the Chondral Lesions And Meniscal Procedures (ChAMP) Trial, we compared outcomes for patients with unstable chondral lesions found at the time of APM and left in situ (CL-noDeb, N = 71) versus patients without unstable chondral lesions (NoCL, N = 47) at 5 years after APM. Outcomes included the Western Ontario and McMaster Universities Arthritis Index (WOMAC), Knee Injury and Osteoarthritis Outcome Score (KOOS), visual analog pain scale, Short-form Health Survey (SF-36), physical knee measurements, progressive joint space narrowing on radiographs, and the rate of additional knee surgery. Multivariate linear regression was used to obtain mean differences (MDs) with corresponding 95% confidence intervals (CIs) adjusted for age, body mass index, and preoperative score (for postoperative scores). RESULTS: Compared with CL-noDeb, NoCL subjects had significantly greater improvement at 5 years in the KOOS score for function in sport and recreation (MD = 9.9 [95% CI, 0.7-19.1]), SF-36 pain (MD = 13.9 [95% CI, 5.5-22.3]), knee extension (MD = 0.8 [95% CI, 0.1-1.5]), and decreased quadriceps circumference at the mid-portion of the patella (MD = -1.5 [95% CI, -2.7 to -0.3). A greater proportion of patients in the NoCL group achieved the MCID for all outcome scores except for the WOMAC pain score (89% CL-NoDeb vs 87% NoCL) and SF-36 general (29% CL-NoDeb vs 23% NoCL). There were no significant group differences in measures of progressive radiographic joint space narrowing in any compartments of the operative knee and no significant difference in the rate of additional knee surgery within 5 years of the initial APM. CONCLUSIONS: Patients undergoing APM without unstable chondral lesions had statistically significantly better outcomes than patients with unstable chondral lesions at 5 years after surgery; however, there were no group differences in progressive radiographic joint space narrowing. LEVEL OF EVIDENCE: Level II, prospective comparative study.

Novel Mutations and Decreased Expression of the Epigenetic Regulator TET2 in Pulmonary Arterial Hypertension.

BACKGROUND: Pulmonary arterial hypertension (PAH) is a lethal vasculopathy. Hereditary cases are associated with germline mutations in BMPR2 and 16 other genes; however, these mutations occur in <25% of patients with idiopathic PAH and are rare in PAH associated with connective tissue diseases. Preclinical studies suggest epigenetic dysregulation, including altered DNA methylation, promotes PAH. Somatic mutations of Tet-methylcytosine-dioxygenase-2 (TET2), a key enzyme in DNA demethylation, occur in cardiovascular disease and are associated with clonal hematopoiesis, inflammation, and adverse vascular remodeling. The role of TET2 in PAH is unknown. METHODS: To test for a role of TET2, we used a cohort of 2572 cases from the PAH Biobank. Within this cohort, gene-specific rare variant association tests were performed using 1832 unrelated European patients with PAH and 7509 non-Finnish European subjects from the Genome Aggregation Database (gnomAD) as control subjects. In an independent cohort of 140 patients, we quantified TET2 expression in peripheral blood mononuclear cells. To assess causality, we investigated hemodynamic and histological evidence of PAH in hematopoietic Tet2-knockout mice. RESULTS: We observed an increased burden of rare, predicted deleterious germline variants in TET2 in PAH patients of European ancestry (9/1832) compared with control subjects (6/7509; relative risk=6; P=0.00067). Assessing the whole cohort, 0.39% of patients (10/2572) had 12 TET2 mutations (75% predicted germline and 25% somatic). These patients had no mutations in other PAH-related genes. Patients with TET2 mutations were older (71±7 years versus 48±19 years; P<0.0001), were more unresponsive to vasodilator challenge (0/7 versus 140/1055 [13.2%]), had lower pulmonary vascular resistance (5.2±3.1 versus 10.5±7.0 Wood units; P=0.02), and had increased inflammation (including elevation of interleukin-1ß). Circulating TET2 expression did not correlate with age and was decreased in >86% of PAH patients. Tet2-knockout mice spontaneously developed PAH, adverse pulmonary vascular remodeling, and inflammation, with elevated levels of cytokines, including interleukin-1ß. Long-term therapy with an antibody targeting interleukin-1ß blockade resulted in regression of PAH. CONCLUSIONS: PAH is the first human disease related to potential TET2 germline mutations. Inherited and acquired abnormalities of TET2 occur in 0.39% of PAH cases. Decreased TET2 expression is ubiquitous and has potential as a PAH biomarker.

Immune Dysregulation and Recurring Mutations in Myelodysplastic Syndromes Pathogenesis.

Myelodysplastic syndromes (MDS) are clonal stem cell malignancies characterized by ineffective hematopoiesis leading to peripheral cytopenias and variable risk of progression to acute myeloid leukemia. Inflammation is associated with MDS pathogenesis. Several cytokines, reactive species of oxygen/nitrogen and growth factors are directly or indirectly involved in dysfunction of the MDS bone marrow (BM) microenvironment. Mutations in genes mainly regulating RNA splicing, DNA methylation and chromatin accessibility, transcription factors, signal transduction and the response to DNA damage contribute to ineffective hematopoiesis, genomic instability and MDS development. The inflammation-associated DNA damage in hematopoietic stem cells may also contribute to MDS development and progression with aggressive clinical characteristics. Many studies have aimed at clarifying mechanisms involved in the activity of immature myeloid cells as powerful modulators of the immune response and their correlation with aging, autoimmunity, and development of cancer. In this review, we explore recent advances and accumulating evidence uniting immune dysregulation, inflammaging and recurring mutations in the pathogenesis of MDS.

Age-Associated TET2 Mutations: Common Drivers of Myeloid Dysfunction, Cancer and Cardiovascular Disease.

Acquired, inactivating mutations in Tet methylcytosine dioxygenase 2 (TET2) are detected in peripheral blood cells of a remarkable 5%-10% of adults greater than 65 years of age. They impart a hematopoietic stem cell advantage and resultant clonal hematopoiesis of indeterminate potential (CHIP) with skewed myelomonocytic differentiation. CHIP is associated with an overall increased risk of transformation to a hematological malignancy, especially myeloproliferative and myelodysplastic neoplasms (MPN, MDS) and acute myeloid leukemia (AML), of approximately 0.5% to 1% per year. However, it is becoming increasingly possible to identify individuals at greatest risk, based on CHIP mutational characteristics. CHIP, and particularly TET2-mutant CHIP, is also a novel, significant risk factor for cardiovascular diseases, related in part to hyper-inflammatory, progeny macrophages carrying TET2 mutations. Therefore, somatic TET2 mutations contribute to myeloid expansion and innate immune dysregulation with age and contribute to prevalent diseases in the developed world-cancer and cardiovascular disease. Herein, we describe the impact of detecting TET2 mutations in the clinical setting. We also present the rationale and promise for targeting TET2-mutant and other CHIP clones, and their inflammatory environment, as potential means of lessening risk of myeloid cancer development and dampening CHIP-comorbid inflammatory diseases.

Longitudinal sequencing of RUNX1 familial platelet disorder: new insights into genetic mechanisms of transformation to myeloid malignancies.

The mechanisms by which patients with RUNX1 familial platelet disorder with propensity to myeloid malignancies (FPDMM) develop myeloid malignancies (MM) are not fully understood. We report the results of targeted next-generation sequencing on three patients with RUNX1 FPDMM who developed acute myeloid leukaemia or myelodysplastic syndromes (AML/MDS). DNA samples were collected from bone marrow, peripheral blood and buccal swabs at different time points. One patient had clonal haematopoiesis, represented by an SRSF2 p.P95R variant, prior to his AML diagnosis, when he developed an additional NRAS p.G12D variant. His sister presented to us with MDS, with a TET2 p.S471fs and identical NRAS p.G12D variant. The third patient, from another family, had an additional RUNX1 p.R204X and an NFE2 p.Q139fs variant at AML diagnosis. This constitutes the first report of NFE2 variants in AML without extramedullary disease and NRAS variants in AML/MDS in the setting of FPDMM. A systematic review of the literature including our findings distinguishes two genetic landscapes at AML transformation from FPDMM characterized by either the presence or absence of somatic abnormalities in RUNX1 with or without variants in genes usually associated with MM. Whether clonal haematopoiesis precedes transformation only in patients without somatic abnormalities in RUNX1 needs further confirmation.

Implementation of an NGS-based sequencing and gene fusion panel for clinical screening of patients with suspected hematologic malignancies.

OBJECTIVES: The diagnosis of hematologic malignancies integrates multiple diagnostic and clinical disciplines. Historically, targeted (single-analyte) genetic testing has been used as reflex to initial prescreening by other diagnostic modalities including flow cytometry, anatomic pathology, and clinical cytogenetics. Given the wide range of mutations associated with hematologic malignancies a DNA/RNA-based NGS panel can provide a more effective and economical approach to comprehensive testing of patients as an initial, tier-1 screen. METHODS: Using a cohort of 380 patients, we performed clinical validation of a gene panel designed to assess 40 genes (DNA), and 29 fusion driver genes with over 600 gene fusion partners (RNA), including sample exchange data across three clinical laboratories, and correlation with cytogenetic testing results. RESULTS: The clinical validation of this technology demonstrated that its accuracy, sensitivity, and specificity are comparable to the majority of targeted single-gene approaches, while assessment of the initial patient cohort data demonstrated a high diagnostic yield of 50.5%. CONCLUSIONS: Implementation of a tier-1 NGS-based protocol for gene panel screening provides a comprehensive alternative to targeted molecular testing in patients with suspected hematologic malignancies, with increased diagnostic yield, scalability, reproducibility, and cost effectiveness, making it ideally suited for implementation in clinical laboratories.

The Effect of Body Mass Index on Clinical Outcomes in Patients Without Radiographic Evidence of Degenerative Joint Disease After Arthroscopic Partial Meniscectomy.

PURPOSE: To examine the effect of obesity on clinical outcomes at 1 year after arthroscopic partial meniscectomy. METHODS: We conducted a secondary analysis of the ChAMP (Chondral Lesions and Meniscus Procedures) randomized controlled trial (N = 256). The visual analog scale for pain, Western Ontario and McMaster Universities Osteoarthritis Index (WOMAC), Knee Injury and Osteoarthritis Outcome Score (KOOS), range of motion, and presence of effusion were assessed preoperatively and at 1 year after arthroscopic partial meniscectomy. Body mass index was categorized as normal weight, 24.99 or less; overweight, 25 to 29.99; or obese, 30 or greater. Analysis of variance or the Cochran-Mantel-Haenszel test was used to examine differences in clinical outcomes between body mass index categories, and mean ± standard deviation or number (percentage) is reported. RESULTS: Preoperatively, obese patients had worse WOMAC pain (56.2 ± 17.2 vs 61.3 ± 17.2, P = .02), WOMAC physical function (55.8 ± 17.1 vs 62.8 ± 17.1, P = .004), pain visual analog scale (4.9 ± 2.1 vs 4.2 ± 1.9, P = .01), KOOS pain (49.5 ± 14.9 vs 54.0 ± 15.1, P = .02), and KOOS quality-of-life (27.9 ± 18.3 vs 36.9 ± 17.0, P = .001) scores, as well as decreased flexion (121.8° ± 22.6° vs 132.3° ± 16.5°, P = .003), compared with normal-weight patients. Overweight patients (n = 51 [51.5%], P = .03) and obese patients (n = 56 [52.8%], P = .002) were more likely to have knee effusion before surgery than normal-weight patients (n = 17 [34%]). At 1 year after surgery, overweight (130.2° ± 7.7°, P = .03) and obese (128.1° ± 7.1°, P = .003) patients had decreased flexion compared with normal-weight patients (134.5° ± 8.3°). CONCLUSIONS: Obese patients had worse pain, physical functioning, and quality-of-life scores, as well as decreased flexion, compared with normal-weight patients before arthroscopic partial meniscectomy. At 1 year after arthroscopic partial meniscectomy, there were no statistically significant differences in clinical outcomes but obesity was associated with decreased knee flexion. LEVEL OF EVIDENCE: Level II, prospective comparative trial.

Clonal haematopoiesis, ageing and kidney disease.

Clonal haematopoiesis of indeterminate potential (CHIP) is a preclinical condition wherein a sizeable proportion of an individual's circulating blood cells are derived from a single mutated haematopoietic stem cell. CHIP occurs frequently with ageing - more than 10% of individuals over 65 years of age are affected - and is associated with an increased risk of disease across several organ systems and premature death. Emerging evidence suggests that CHIP has a role in kidney health, including associations with predisposition to acute kidney injury, impaired recovery from acute kidney injury and kidney function decline, both in the general population and among those with chronic kidney disease. Beyond its direct effect on the kidney, CHIP elevates the susceptibility of individuals to various conditions that can detrimentally affect the kidneys, including cardiovascular disease, obesity and insulin resistance, liver disease, gout, osteoporosis and certain autoimmune diseases. Aberrant pro-inflammatory signalling, telomere attrition and epigenetic ageing are potential causal pathophysiological pathways and mediators that underlie CHIP-related disease risk. Experimental animal models have shown that inhibition of inflammatory cytokine signalling can ameliorate many of the pathological effects of CHIP, and assessment of the efficacy and safety of this class of medications for human CHIP-associated pathology is ongoing.

Clonal hematopoiesis of indeterminate potential-associated non-small cell lung cancer risk is potentiated by small particulate matter air pollution among non-smokers: a novel somatic variant-environment interaction.

Small particulate matter air pollution (PM 2.5 ) is a recognized driver of non-small cell lung cancer (NSCLC) among non-smoking individuals. Inhaled PM 2.5 recruits pro-inflammatory macrophages to the air-lung interface, which promotes malignant lung epithelial cell growth and progression to overt cancer. We sought to determine whether clonal hematopoiesis of indeterminate potential (CHIP), a common age-related condition characterized by hyperinflammatory macrophages, exacerbates PM 2.5 -associated NSCLC in non-smokers using genetic, environmental, and phenotypic data from 413,901 individuals in the UK Biobank. Among non-smokers, PM 2.5 is not associated with NSCLC and not associated with prevalence of CHIP, but CHIP is associated with a doubling of NSCLC risk (hazard ratio (HR) 2.01, 95% confidence interval (CI): 1.34-3.00). Moreover, CHIP-associated NSCLC risk is exacerbated in the setting of above-median PM 2.5 levels (HR 2.70, 95% CI: 1.60-4.55). PM 2.5 × CHIP is also associated with significantly greater markers of systemic inflammation (CRP, IL-6, and IL-1ß) than expected. Altogether, these results suggest CHIP and PM 2.5 form a novel gene × environment interaction promoting NSCLC tumorigenesis in non-smokers.

Chronic TNF in the aging microenvironment exacerbates Tet2-loss-of-function myeloid expansion.

Somatic mutations in the TET2 gene occur more frequently with age, imparting an intrinsic hematopoietic stem cell (HSC) advantage and contributing to a phenomenon termed clonal hematopoiesis of indeterminate potential (CHIP). Individuals with TET2-mutant CHIP have a higher risk of developing myeloid neoplasms and other aging-related conditions. Despite its role in unhealthy aging, the extrinsic mechanisms driving TET2-mutant CHIP clonal expansion remain unclear. We previously showed an environment containing TNF favours TET2-mutant HSC expansion in vitro. We therefore postulated that age-related increases in TNF also provide an advantage to HSCs with TET2-mutations in vivo. To test this hypothesis, we generated mixed bone marrow chimeric mice of old wild-type (WT) and TNF-/- genotypes reconstituted with WT CD45.1+ and Tet2-/-CD45.2+ HSCs. We show that age-associated increases in TNF dramatically increased the expansion of Tet2-/-cells in old WT recipient mice, with strong skewing towards the myeloid lineage. This aberrant myelomonocytic advantage was mitigated in old TNF-/- recipient mice, suggesting that TNF signalling is essential for the expansion Tet2-mutant myeloid clones. Examination of human rheumatoid arthritis patients with clonal hematopoiesis revealed that hematopoietic cells carrying certain mutations, including in TET2, may be sensitive to reduced TNF bioactivity following blockade with adalimumab. This suggests that targeting TNF may reduce the burden of some forms of CHIP. To our knowledge, this is the first evidence to demonstrate that TNF has a causal role in driving TET2-mutant CHIP in vivo. These findings highlight TNF as a candidate therapeutic target to control TET2-mutant CHIP.

Neutrophil-mediated innate immune resistance to bacterial pneumonia is dependent on Tet2 function.

Individuals with clonal hematopoiesis of indeterminate potential (CHIP) are at increased risk of aging related health conditions and all-cause mortality, but whether CHIP affects risk of infection is much less clear. Using UK Biobank data, we revealed a positive association between CHIP and incident pneumonia in 438,421 individuals. We show that inflammation enhanced pneumonia risk, as CHIP carriers with a hypomorphic IL6 receptor polymorphism were protected. To better characterize the pathways of susceptibility, we challenged hematopoietic Tet Methylcytosine Dioxygenase 2-knockout (Tet2-/-) and floxed control mice (Tet2fl/fl) with Streptococcus pneumoniae. As with human CHIP carriers, Tet2-/- mice had hematopoietic abnormalities resulting in the expansion of inflammatory monocytes and neutrophils in peripheral blood. Yet, these cells were insufficient in defending against S. pneumoniae and resulted in increased pathology, impaired bacterial clearance, and higher mortality in Tet2-/- mice. We delineated the transcriptional landscape of Tet2-/- neutrophils and found that, while inflammation-related pathways were upregulated in Tet2-/- neutrophils, migration and motility pathways were compromised. Using live-imaging techniques, we demonstrated impairments in motility, pathogen uptake, and neutrophil extracellular trap (NET) formation by Tet2-/- neutrophils. Collectively, we show that CHIP is a risk factor for bacterial pneumonia related to innate immune impairments.