Clonal haematopoiesis and risk of chronic liver disease.

Chronic liver disease is a major public health burden worldwide1. Although different aetiologies and mechanisms of liver injury exist, progression of chronic liver disease follows a common pathway of liver inflammation, injury and fibrosis2. Here we examined the association between clonal haematopoiesis of indeterminate potential (CHIP) and chronic liver disease in 214,563 individuals from 4 independent cohorts with whole-exome sequencing data (Framingham Heart Study, Atherosclerosis Risk in Communities Study, UK Biobank and Mass General Brigham Biobank). CHIP was associated with an increased risk of prevalent and incident chronic liver disease (odds ratio = 2.01, 95% confidence interval (95% CI) [1.46, 2.79]; P < 0.001). Individuals with CHIP were more likely to demonstrate liver inflammation and fibrosis detectable by magnetic resonance imaging compared to those without CHIP (odds ratio = 1.74, 95% CI [1.16, 2.60]; P = 0.007). To assess potential causality, Mendelian randomization analyses showed that genetic predisposition to CHIP was associated with a greater risk of chronic liver disease (odds ratio = 2.37, 95% CI [1.57, 3.6]; P < 0.001). In a dietary model of non-alcoholic steatohepatitis, mice transplanted with Tet2-deficient haematopoietic cells demonstrated more severe liver inflammation and fibrosis. These effects were mediated by the NLRP3 inflammasome and increased levels of expression of downstream inflammatory cytokines in Tet2-deficient macrophages. In summary, clonal haematopoiesis is associated with an elevated risk of liver inflammation and chronic liver disease progression through an aberrant inflammatory response.

Inherited causes of clonal haematopoiesis in 97,691 whole genomes.

Age is the dominant risk factor for most chronic human diseases, but the mechanisms through which ageing confers this risk are largely unknown1. The age-related acquisition of somatic mutations that lead to clonal expansion in regenerating haematopoietic stem cell populations has recently been associated with both haematological cancer2-4 and coronary heart disease5-this phenomenon is termed clonal haematopoiesis of indeterminate potential (CHIP)6. Simultaneous analyses of germline and somatic whole-genome sequences provide the opportunity to identify root causes of CHIP. Here we analyse high-coverage whole-genome sequences from 97,691 participants of diverse ancestries in the National Heart, Lung, and Blood Institute Trans-omics for Precision Medicine (TOPMed) programme, and identify 4,229 individuals with CHIP. We identify associations with blood cell, lipid and inflammatory traits that are specific to different CHIP driver genes. Association of a genome-wide set of germline genetic variants enabled the identification of three genetic loci associated with CHIP status, including one locus at TET2 that was specific to individuals of African ancestry. In silico-informed in vitro evaluation of the TET2 germline locus enabled the identification of a causal variant that disrupts a TET2 distal enhancer, resulting in increased self-renewal of haematopoietic stem cells. Overall, we observe that germline genetic variation shapes haematopoietic stem cell function, leading to CHIP through mechanisms that are specific to clonal haematopoiesis as well as shared mechanisms that lead to somatic mutations across tissues.

PPM1D modulates hematopoietic cell fitness and response to DNA damage and is a therapeutic target in myeloid malignancy.

PPM1D encodes a phosphatase that is recurrently activated across cancer, most notably in therapy-related myeloid neoplasms. However, the function of PPM1D in hematopoiesis and its contribution to tumor cell growth remain incompletely understood. Using conditional mouse models, we uncover a central role for Ppm1d in hematopoiesis and validate its potential as a therapeutic target. We find that Ppm1d regulates the competitive fitness and self-renewal of hematopoietic stem cells (HSCs) with and without exogenous genotoxic stresses. We also show that although Ppm1d activation confers cellular resistance to cytotoxic therapy, it does so to a lesser degree than p53 loss, informing the clonal competition phenotypes often observed in human studies. Notably, loss of Ppm1d sensitizes leukemias to cytotoxic therapies in vitro and in vivo, even in the absence of a Ppm1d mutation. Vulnerability to PPM1D inhibition is observed across many cancer types and dependent on p53 activity. Importantly, organism-wide loss of Ppm1d in adult mice is well tolerated, supporting the tolerability of pharmacologically targeting PPM1D. Our data link PPM1D gain-of-function mutations to the clonal expansion of HSCs, inform human genetic observations, and support the therapeutic targeting of PPM1D in cancer.

TET2-mutant clonal hematopoiesis and risk of gout.

Gout is a common inflammatory arthritis caused by precipitation of monosodium urate (MSU) crystals in individuals with hyperuricemia. Acute flares are accompanied by secretion of proinflammatory cytokines, including interleukin-1ß (IL-1ß). Clonal hematopoiesis of indeterminate potential (CHIP) is an age-related condition predisposing to hematologic cancers and cardiovascular disease. CHIP is associated with elevated IL-1ß, thus we investigated CHIP as a risk factor for gout. To test the clinical association between CHIP and gout, we analyzed whole exome sequencing data from 177 824 individuals in the MGB Biobank (MGBB) and UK Biobank (UKB). In both cohorts, the frequency of gout was higher among individuals with CHIP than without CHIP (MGBB, CHIP with variant allele fraction [VAF] ≥2%: odds ratio [OR], 1.69; 95% CI, 1.09-2.61; P = .0189; UKB, CHIP with VAF ≥10%: OR, 1.25; 95% CI, 1.05-1.50; P = .0133). Moreover, individuals with CHIP and a VAF ≥10% had an increased risk of incident gout (UKB: hazard ratio [HR], 1.28; 95% CI, 1.06-1.55; P = .0107). In murine models of gout pathogenesis, animals with Tet2 knockout hematopoietic cells had exaggerated IL-1ß secretion and paw edema upon administration of MSU crystals. Tet2 knockout macrophages elaborated higher levels of IL-1ß in response to MSU crystals in vitro, which was ameliorated through genetic and pharmacologic Nlrp3 inflammasome inhibition. These studies show that TET2-mutant CHIP is associated with an increased risk of gout in humans and that MSU crystals lead to elevated IL-1ß levels in Tet2 knockout murine models. We identify CHIP as an amplifier of NLRP3-dependent inflammatory responses to MSU crystals in patients with gout.

Impact of diagnostic genetics on remission MRD and transplantation outcomes in older patients with AML.

Older patients with acute myeloid leukemia (AML) have high relapse risk and poor survival after allogeneic hematopoietic cell transplantation (HCT). Younger patients may receive myeloablative conditioning to mitigate relapse risk associated with high-risk genetics or measurable residual disease (MRD), but older adults typically receive reduced-intensity conditioning (RIC) to limit toxicity. To identify factors that drive HCT outcomes in older patients, we performed targeted mutational analysis (variant allele fraction ≥2%) on diagnostic samples from 295 patients with AML aged ≥60 years who underwent HCT in first complete remission, 91% of whom received RIC, and targeted duplex sequencing at remission in a subset comprising 192 patients. In a multivariable model for leukemia-free survival (LFS) including baseline genetic and clinical variables, we defined patients with low (3-year LFS, 85%), intermediate (55%), high (35%), and very high (7%) risk. Before HCT, 79.7% of patients had persistent baseline mutations, including 18.3% with only DNMT3A or TET2 (DT) mutations and 61.4% with other mutations (MRD positive). In univariable analysis, MRD positivity was associated with increased relapse and inferior LFS, compared with DT and MRD-negative mutations. However, in a multivariable model accounting for baseline risk, MRD positivity had no independent impact on LFS, most likely because of its significant association with diagnostic genetic characteristics, including MDS-associated gene mutations, TP53 mutations, and high-risk karyotype. In summary, molecular associations with MRD positivity and transplant outcomes in older patients with AML are driven primarily by baseline genetics, not by mutations present in remission. In this group of patients, where high-intensity conditioning carries substantial risk of toxicity, alternative approaches to mitigating MRD-associated relapse risk are needed.

Association of clonal hematopoiesis with chronic obstructive pulmonary disease.

Chronic obstructive pulmonary disease (COPD) is associated with age and smoking, but other determinants of the disease are incompletely understood. Clonal hematopoiesis of indeterminate potential (CHIP) is a common, age-related state in which somatic mutations in clonal blood populations induce aberrant inflammatory responses. Patients with CHIP have an elevated risk for cardiovascular disease, but the association of CHIP with COPD remains unclear. We analyzed whole-genome sequencing and whole-exome sequencing data to detect CHIP in 48 835 patients, of whom 8444 had moderate to very severe COPD, from four separate cohorts with COPD phenotyping and smoking history. We measured emphysema in murine models in which Tet2 was deleted in hematopoietic cells. In the COPDGene cohort, individuals with CHIP had risks of moderate-to-severe, severe, or very severe COPD that were 1.6 (adjusted 95% confidence interval [CI], 1.1-2.2) and 2.2 (adjusted 95% CI, 1.5-3.2) times greater than those for noncarriers. These findings were consistently observed in three additional cohorts and meta-analyses of all patients. CHIP was also associated with decreased FEV1% predicted in the COPDGene cohort (mean between-group differences, -5.7%; adjusted 95% CI, -8.8% to -2.6%), a finding replicated in additional cohorts. Smoke exposure was associated with a small but significant increased risk of having CHIP (odds ratio, 1.03 per 10 pack-years; 95% CI, 1.01-1.05 per 10 pack-years) in the meta-analysis of all patients. Inactivation of Tet2 in mouse hematopoietic cells exacerbated the development of emphysema and inflammation in models of cigarette smoke exposure. Somatic mutations in blood cells are associated with the development and severity of COPD, independent of age and cumulative smoke exposure.

The clinical and functional effects of TERT variants in myelodysplastic syndrome.

Germline pathogenic TERT variants are associated with short telomeres and an increased risk of developing myelodysplastic syndrome (MDS) among patients with a telomere biology disorder. We identified TERT rare variants in 41 of 1514 MDS patients (2.7%) without a clinical diagnosis of a telomere biology disorder who underwent allogeneic transplantation. Patients with a TERT rare variant had shorter telomere length (P < .001) and younger age at MDS diagnosis (52 vs 59 years, P = .03) than patients without a TERT rare variant. In multivariable models, TERT rare variants were associated with inferior overall survival (P = .034) driven by an increased incidence of nonrelapse mortality (NRM; P = .015). Death from a noninfectious pulmonary cause was more frequent among patients with a TERT rare variant. Most variants were missense substitutions and classified as variants of unknown significance. Therefore, we cloned all rare missense variants and quantified their impact on telomere elongation in a cell-based assay. We found that 90% of TERT rare variants had severe or intermediate impairment in their capacity to elongate telomeres. Using a homology model of human TERT bound to the shelterin protein TPP1, we inferred that TERT rare variants disrupt domain-specific functions, including catalysis, protein-RNA interactions, and recruitment to telomeres. Our results indicate that the contribution of TERT rare variants to MDS pathogenesis and NRM risk is underrecognized. Routine screening for TERT rare variants in MDS patients regardless of age or clinical suspicion may identify clinically inapparent telomere biology disorders and improve transplant outcomes through risk-adapted approaches.

Contribution of clonal hematopoiesis to adult-onset hemophagocytic lymphohistiocytosis.

Adult-onset hemophagocytic lymphohistiocytosis (HLH) is a rare, life-threatening disease of immune hyperactivation. Unlike pediatric HLH, adult HLH is rarely driven by germline genetic variants. Although numerous precipitating etiologies have been identified, the reason that HLH occurs in only a subset of individuals and how other factors contribute to the disease remains unknown. We hypothesized that clonal hematopoiesis (CH), a state in which somatic mutations in blood cells cause an expanded population of mutant hematopoietic cells and drive an aberrant inflammatory state, could contribute to adult-onset HLH. In a highly annotated cohort of older adults with HLH we found that CH was more prevalent than in control cohorts. Using the adult-onset HLH mouse model in which repeated treatments of the TLR9 agonist, ODN1826, was delivered to the mouse, we observed that macrophages carrying mutations in Tet2, one of the most commonly mutated genes in CH, have an enhanced inflammatory response to TLR9 agonism. Finally, mice carrying Tet2 mutations in the hematopoietic compartment (a common model for CH) displayed an exaggerated response to TLR9 agonism, including worse splenomegaly and anemia. Our data suggest that CH is more common in individuals with adult-onset HLH and can contribute to the pathophysiology of this disease.

Short telomere length predicts nonrelapse mortality after stem cell transplantation for myelodysplastic syndrome.

Allogeneic hematopoietic stem cell transplantation is the only potentially curative treatment for patients with myelodysplastic syndrome (MDS), but long-term survival is limited by the risk of transplant-related complications. Short telomere length, mediated by inherited or acquired factors, impairs cellular response to genotoxic and replicative stress and could identify patients at higher risk for toxicity after transplantation. We measured relative telomere length in pretransplant recipient blood samples in 1514 MDS patients and evaluated the association of telomere length with MDS disease characteristics and transplantation outcomes. Shorter telomere length was significantly associated with older age, male sex, somatic mutations that impair the DNA damage response, and more severe pretransplant cytopenias, but not with bone marrow blast count, MDS treatment history, or history of prior cancer therapy. Among 1267 patients ≥40 years old, telomere length in the shortest quartile was associated with inferior survival (P < .001) because of a high risk of nonrelapse mortality (NRM; P = .001) after adjusting for significant clinical and genetic variables. The adverse impact of shorter telomeres on NRM was independent of recipient comorbidities and was observed selectively among patients receiving more intensive conditioning, including myeloablative regimens and higher dose melphalan-based reduced-intensity regimens. The effect of shorter telomeres on NRM was prominent among patients who developed severe acute graft-versus-host disease, suggesting that short telomere length may limit regenerative potential of mucosal tissues after acute injury. MDS patients with shorter telomere length, who have inferior survival driven by excess toxicity, could be considered for strategies focused on minimizing toxic effects of transplantation.

Clonal Hematopoiesis and Risk of Atherosclerotic Cardiovascular Disease.

BACKGROUND: Clonal hematopoiesis of indeterminate potential (CHIP), which is defined as the presence of an expanded somatic blood-cell clone in persons without other hematologic abnormalities, is common among older persons and is associated with an increased risk of hematologic cancer. We previously found preliminary evidence for an association between CHIP and atherosclerotic cardiovascular disease, but the nature of this association was unclear. METHODS: We used whole-exome sequencing to detect the presence of CHIP in peripheral-blood cells and associated such presence with coronary heart disease using samples from four case-control studies that together enrolled 4726 participants with coronary heart disease and 3529 controls. To assess causality, we perturbed the function of Tet2, the second most commonly mutated gene linked to clonal hematopoiesis, in the hematopoietic cells of atherosclerosis-prone mice. RESULTS: In nested case-control analyses from two prospective cohorts, carriers of CHIP had a risk of coronary heart disease that was 1.9 times as great as in noncarriers (95% confidence interval [CI], 1.4 to 2.7). In two retrospective case-control cohorts for the evaluation of early-onset myocardial infarction, participants with CHIP had a risk of myocardial infarction that was 4.0 times as great as in noncarriers (95% CI, 2.4 to 6.7). Mutations in DNMT3A, TET2, ASXL1, and JAK2 were each individually associated with coronary heart disease. CHIP carriers with these mutations also had increased coronary-artery calcification, a marker of coronary atherosclerosis burden. Hypercholesterolemia-prone mice that were engrafted with bone marrow obtained from homozygous or heterozygous Tet2 knockout mice had larger atherosclerotic lesions in the aortic root and aorta than did mice that had received control bone marrow. Analyses of macrophages from Tet2 knockout mice showed elevated expression of several chemokine and cytokine genes that contribute to atherosclerosis. CONCLUSIONS: The presence of CHIP in peripheral-blood cells was associated with nearly a doubling in the risk of coronary heart disease in humans and with accelerated atherosclerosis in mice. (Funded by the National Institutes of Health and others.).

High NPM1-mutant allele burden at diagnosis predicts unfavorable outcomes in de novo AML.

Acute myeloid leukemia (AML) with mutated NPM1 is a newly recognized separate entity in the revised 2016 World Health Organization classification and is associated with a favorable prognosis. Although previous studies have evaluated NPM1 in a binary fashion, little is known about the significance of its mutant allele burden at diagnosis, nor has the effect of comutations (other than FLT3) been extensively evaluated. We retrospectively used targeted sequencing data from 109 patients with de novo AML with mutated NPM1 to evaluate the potential significance of NPM1 variant allele frequency (VAF), comutations, and clinical parameters with regard to patient outcomes. We observed that high NPM1 VAF (uppermost quartile) correlated with shortened overall survival (median, 12.1 months vs not reached; P < .0001) as well as event-free survival (median, 7.5 vs 65.44 months; P < .0001) compared with the other NPM1-mutated cases. In both univariate and multivariable analyses, high NPM1 VAF had a particularly adverse prognostic effect in the subset of patients treated with stem-cell transplantation in first remission (P = .0004) and in patients with mutated DNMT3A (P < .0001). Our findings indicate that the prognostic effect of NPM1 mutation in de novo AML may be influenced by the relative abundance of the mutated allele.

Control of spatially rotating structures in diffractive Kerr cavities.

Turing patterns in self-focussing nonlinear optical cavities pumped by beams carrying orbital angular momentum (OAM) m are shown to rotate with an angular velocity ω=2m/R 2 on rings of radii R. We verify this prediction in 1D models on a ring and for 2D Laguerre-Gaussian and top-hat pumps with OAM. Full control over the angular velocity of the pattern in the range -2m/R 2≤ω≤2m/R 2 is obtained by using cylindrical vector beam pumps that consist of orthogonally polarized eigenmodes with equal and opposite OAM. Using Poincaré beams that consist of orthogonally polarized eigenmodes with different magnitudes of OAM, the resultant angular velocity is ω=(m L+m R)/R 2, where m L,m R are the OAMs of the eigenmodes, assuming good overlap between the eigenmodes. If there is no, or very little, overlap between the modes then concentric Turing pattern rings, each with angular velocity ω=2m L,R /R 2 will result. This can lead to, for example, concentric, counter-rotating Turing patterns creating an optical peppermill-type structure. Full control over the speeds of multiple rings has potential applications in particle manipulation and stretching, atom trapping, and circular transport of cold atoms and BEC wavepackets.

Association of mutations with morphological dysplasia in de novo acute myeloid leukemia without 2016 WHO Classification-defined cytogenetic abnormalities.

Despite improvements in our understanding of the molecular basis of acute myeloid leukemia (AML), the association between genetic mutations with morphological dysplasia remains unclear. In this study, we evaluated and scored dysplasia in bone marrow (BM) specimens from 168 patients with de novo AML; none of these patients had cytogenetic abnormalities according to the 2016 World Health Organization Classification. We then performed targeted sequencing of diagnostic BM aspirates for recurrent mutations associated with myeloid malignancies. We found that cohesin pathway mutations [q (FDR-adjusted P)=0.046] were associated with a higher degree of megakaryocytic dysplasia and STAG2 mutations were marginally associated with greater myeloid lineage dysplasia (q=0.052). Frequent megakaryocytes with separated nuclear lobes were more commonly seen among cases with cohesin pathway mutations (q=0.010) and specifically in those with STAG2 mutations (q=0.010), as well as NPM1 mutations (q=0.022 when considering the presence of any vs no megakaryocytes with separated nuclear lobes). RAS pathway mutations (q=0.006) and FLT3-ITD (q=0.006) were significantly more frequent in cases without evaluable erythroid cells. In univariate analysis of the 153 patients treated with induction chemotherapy, NPM1 mutations were associated with longer event-free survival (EFS) (P=0.042), while RUNX1 (P=0.042), NF1 (P=0.040), frequent micromegakaryocytes (P=0.018) and presence of a subclone (P=0.002) were associated with shorter EFS. In multivariable modeling, NPM1 was associated with longer EFS, while presence of a subclone and frequent micromegakaryocytes remained significantly associated with shorter EFS.

Optical Rogue Waves in Vortex Turbulence.

We present a spatiotemporal mechanism for producing 2D optical rogue waves in the presence of a turbulent state with creation, interaction, and annihilation of optical vortices. Spatially periodic structures with bound phase lose stability to phase unbound turbulent states in complex Ginzburg-Landau and Swift-Hohenberg models with external driving. When the pumping is high and the external driving is low, synchronized oscillations are unstable and lead to spatiotemporal vortex-mediated turbulence with high excursions in amplitude. Nonlinear amplification leads to rogue waves close to turbulent optical vortices, where the amplitude tends to zero, and to probability density functions (PDFs) with long tails typical of extreme optical events.

Reevaluation of the pretransplant assessment of mortality score after allogeneic hematopoietic transplantation.

The Pretransplant Assessment of Mortality (PAM) score was developed in 2006 to predict risk of mortality after allogeneic hematopoietic cell transplantation (HCT). Transplant practices have evolved during the past decade, suggesting the need to reevaluate the performance of the PAM score. We used statistical modeling to analyze and recalibrate mortality based on overall PAM scores, its components, and conditioning regimen in a retrospective cohort of 1549 patients who had HCT from 2003 through 2009. PAM scores correlated with mortality, but the effect size was smaller in the current study than in previous studies. PAM scores also demonstrated a stronger association with mortality in patients who received myeloablative conditioning than in those who received reduced-intensity conditioning. In contrast to the original study, carbon monoxide diffusing capacity, serum alanine aminotransferase, and serum creatinine concentrations were no longer significantly associated with 2-year mortality, whereas patient and donor cytomegalovirus serology was associated with mortality in the current cohort. Based on our findings, we developed and tested a revised PAM score for clinicians to estimate survival after allogeneic HCT with myeloablative conditioning regimens for patients with hematologic malignancy. Prognostic models such as the PAM score should be updated and recalibrated periodically to accommodate changes in clinical practice.