Clonal hematopoiesis of indeterminate potential and cardiovascular diseases: A review

Abstract

Cardiovascular disease (CVD) is a major driver of mortality and declining health worldwide. Cardiovascular diseases (CVD) is the most common cause of morbidity and mortality globally. Although dyslipidemia, smoking, diabetes, hypertension and obesity are some well-known causes of CVD, the overlapping genetic pathways between other diseases and those affecting cardiovascular health have been overlooked.

In the past decade, mutations in TET2, DNMT3A, ASXL1, and JAK2 are found to cause clonal hematopoiesis of intermediate potential (CHIP), a disease associated with age-related haematological malignancies without the presence of cytopenias or dysplasia. Coronary artery disease, heart failure, aortic stenosis, and arrhythmias have been shown to be associated with the presence of CHIP mutations. Addressing the association between CHIP could significantly reduce residual risk patients with CVD. The link between CHIP and CVD can potentially be addressed through inhibitors of inflammasomes, antagonists in the interleukin pathway, or direct antagonists of CHIP mutations.

1. Introduction

1.1. Clonal hematopoiesis of indeterminate potential (CHIP)

Clonal hematopoiesis of indeterminate potential (CHIP) is described as the presence of somatic mutations with a variant allele frequency of ≥2 % in peripheral blood, resulting in clonal expansion without causing cytopenias or dysplastic hematopoiesis. CHIP usually is benign; They do not progress normally; rather, they can be considered a precursor to haematological cancers such as monoclonal gammopathy of unknown significance (MGUS).¹ Some individuals suffer from idiopathic cytopenia of uncertain significance (ICUS), which is the presence of persistent unexplained cytopenia that lacks morphologic or cytogenetic evidence of MDS^2,3 (or other underlying illnesses). Another condition related to clonal hematopoiesis, called as clonal cytopenia of undetermined significance (CCUS) is defined as by the presence of unexplained cytopenia, does not meet the diagnostic criteria for MDS, presence of one or more MDS-related mutations, no or only mild (<10 %) dysplasia and blast cells <5 %.^4,5 Age remains an important non-modifiable risk factor for Atherosclerotic Cardiovascular Disease (ASCVD). The pathogenesis of ASCVD revolves around inflammatory cells derived from erythroid, myeloid, lymphoid and platelet series. The hematopoietic stem cells (HSC) is the precursor of such cell series. Due to high-turnover, various environmental and genetic factors, HSCs get acquired mutations. Exonic mutations may arise at the frequency of 1 mutation per decade of life in HSCs.⁶ Some of these mutations affect the tendency to identify, prevent or repair the damages in DNA leading to selective expansion of such mutated cells ending up in a selective clonal expansion of those mutated cells. Other than age, smoking and few germline polymorphisms have been shown to be involved in CHIP. Prior chemotherapy and radiation therapy may augment the formation of CHIP mutations.

People with CHIP have been shown to have a 13-fold increased risk of hematologic malignancies and a 1.4-fold higher risk of death,^1,7, 8, 9 although the annual risk of developing hematologic malignancies is only 0.5–1%.¹ Most individuals with CHIP mutations remain asymptomatic, similar to those with MGUS, but carriers have a 40 % increased risk of all-cause mortality.¹⁰ Prior chemotherapy and radiotherapy are linked to CHIP formation, often involving DNA repair genes like TP53, potentially leading to therapy-related leukemia or myelodysplastic syndrome. CHIP in bone marrow transplant patients is also associated with poorer outcomes and higher mortality.¹¹

1.2. Global burden of cardiovascular diseases (CVD)

CVD remain the most important cause of morbidity and mortality globally. Amongst CVD, Coronary artery diseases (CAD) remain the leading contributor for the same. Being so, it contributes to significant loss of health and health-related economic expenditure for the society.¹² Such burden of diseases is assessed by various societies. However, the most important one amongst them is Global burden of diseases (GBD).¹² The GBD study involves more than 204 countries and territories collaborating with >8000 people across the globe. They produce several epidemiological measures including disability-adjusted life years (DALYs).

1.3. CAD epidemic in India: A matter of serious concern

The India State-level disease burden study which is part of the GBD study group reported an alarming 2.3-fold increase in the prevalence of CVD from 1990 to 2016.¹³ Nearly 1/4th of all deaths and 1/7th of DALY in India was attributed to CVD. Amongst the CVD, CAD remains the leading contributor of death responsible for 16 % of total deaths.¹⁴

1.4. Risk factors of CAD

The INTERHEART study¹⁵ of over 15,000 patients across 52 countries found that dyslipidemia, smoking, diabetes, hypertension, obesity, poor diet, and lack of exercise are major global risk factors for myocardial infarction, regardless of region. The GBD study¹² identified additional risk factors for CAD, including lead exposure, pollution, non-optimal temperatures, second-hand smoke, and kidney dysfunction. The recent COVID-19 pandemic also led to a rise in acute myocardial infarctions.¹⁶ Despite controlling conventional risk factors like blood pressure, LDL, and HbA1c, a certain percentage of patients experience recurrent CVD events, known as residual risk. The FOURIER trial¹⁷ found a 9.8 % recurrent event rate at 3 years, even with PCSK9 inhibitor use, similar to findings from the ODYSSEY study.¹⁸ Residual risk is categorized into inflammatory, thrombotic, and metabolic types.¹⁹ Reducing metabolic risk may involve targeting Lipoprotein(a) with new treatments, lowering triglycerides with icosapent ethyl, and using GLP-1 analogues or SGLT2 inhibitors. The COMPASS trial¹⁹ showed that low-dose rivaroxaban reduces residual thrombotic risk. Addressing residual inflammation is still developing, with CHIP being one factor implicated in this pathway.

1.5. Genes involved in CHIP

Xie et al.²⁰ observed that the prevalence of CHIP mutations increased with age, rising from 0.9 % in the 4th decade to 6.1 % in the 8th decade. Genovese et al.⁷ found that 1 in 150 individuals under 50 had detectable clonal hematopoiesis, while the prevalence increased to 1 in 17 in those over 65. Though more than 70 genes have been described to have CHIP mutations, studies have shown that such somatic mutations predominantly happened in few locations related to hematologic malignancies.^1,7, 8, 9 Most of the genes involved in CHIP have important roles in the methylation and demethylation proteins that affect the structural properties of DNA.²¹ The most commonly mutated genes linked to hematologic malignancy were DNMT3A, TET2, ASXL1, TP53, JAK2, and SF3B1 (Table 1).⁸ DNMT3A is a DNA methyltransferase that epigenetically regulates hematopoiesis and functions as a tumour suppressor gene. Loss of function in DNMT3A can lead to leukemogenesis.²¹ Decrease in function of TET2 was shown to impact both the renewal and differentiation of HSCs into myeloid lineages. It is very uncommon to have two mutations simultaneously in an individual patient.⁸ Furthermore, the clonal proliferation of these mutant cells may hinder the generation of inflammatory cells like macrophages, and endothelial cells that are necessary for the vascular repair process. Mutations in JAK2 is much less commonly associated with CHIP than DNMT3A or TET2.

Table 1.

Function of most commonly reported clonal hematopoiesis of indeterminate potential (CHIP) genes.

Gene	Chromosome Location	Function
DNMT3A	2p23.3	•Essential for genome-wide de novo methylation and establishment of DNA methylation patterns during development. •Promotes vascular smooth muscle cell proliferation and fibrosis, contributing to plaque formation, vascular remodeling, and stiffness. •Coordinates DNA and histone methylation patterns in vascular cells, influencing gene expression related to atherosclerosis, inflammation, and endothelial dysfunction.
TET2	4q24	•Regulates DNA demethylation by converting 5-methylcytosine (5 mC) to 5-hydroxymethylcytosine (5hmC), impacting gene expression related to vascular homeostasis, inflammation, and endothelial function. •Inhibits vascular smooth muscle cell (VSMC) proliferation and migration, affecting atherosclerotic plaque formation and stability while also playing a critical role in immune cell regulation, especially in macrophage polarization, essential for atherosclerosis development. •Loss or mutation of TET2 is associated with chronic inflammation, worsening CVD and contributing to atherogenesis and vascular dysfunction.
ASXL1	20q11.21	•Probable Polycomb group (PcG) protein involved in transcriptional regulation through chromatin remodeling and affecting pathways involved in vascular biology and inflammation. •ASXL1 plays a role in the adipogenic differentiation of precursor cells into mature adipocytes by regulating key transcription factors involved in the process.
TP53	17p13.1	•Induces cell cycle arrest through the transcription of p21 and other cyclin-dependent kinase inhibitors, preventing the proliferation of damaged or dysfunctional endothelial and smooth muscle cells. •Promotes programmed cell death in response to cellular stress, such as oxidative stress and hypoxia, by upregulating pro-apoptotic genes thereby eliminating damaged cells from the vasculature, maintaining vascular integrity.
JAK2	9p24.1	•Involved in various processes such as cell growth, development, differentiation or histone modifications. •Mediates the signaling of pro-inflammatory cytokines (e.g., IL-6, TNF-α), contributing to the inflammatory response in the vasculature. •Plays pivotal role for signal transduction from hematopoietic cytokine receptors, mediating the activation of STAT proteins, required in definitive erythropoiesis.
SF3B1	2q33.1	•SF3B1 is a critical component of the spliceosome, a complex responsible for the splicing of pre-mRNA into mature mRNA. By influencing the alternative splicing of pre-mRNAs, SF3B1 plays a role in the production of various protein isoforms.

1.6. CHIP and associated cardiovascular disease

Various CVDs are shown to be associated with the presence of CHIP mutations as described below (Fig. 1).

Fig. 1.

Central illustration illustrating role of clonal hematopoiesis of indeterminate potential (CHIP) genes in various cardiovascular diseases.

1.7. CHIP and coronary artery disease (CAD)

An emerging association has been identified between CHIP and both coronary artery calcification (CAC) and early-onset myocardial infarction (MI). In a nested case–control study by Jaiswal et al.,²² patients with CAD exhibited a higher prevalence of CHIP compared to matched controls (17 % vs. 7 %), with a median age of 70 years. The study also identified mutations in DNMT3A, TET2, ASXL1, and JAK2 as individually linked to CAD. Notably, patients with CHIP had a threefold increased risk of having a high CAC score (>615 Agatston units). An elevated variant allele frequency (VAF) > 10 % correlated with an increased incidence of CAC and ASCVD. Additionally, younger patients showed a lower prevalence of CHIP mutations compared to older individuals. However, among younger patients, those with CHIP mutations demonstrated a fourfold increased risk of early MI. These findings underscore the importance of identifying CHIP mutations and elucidating their potential etio-pathogenic role in acute coronary syndromes. In addition to macrovascular disease, Akhiyat et al.,²³ recently found that CHIP was associated with a worse coronary flow reserve. In addition, they found that there existed association between CHIP and major adverse cardiac events in patients with microvascular diseases.

1.8. CHIP and heart failure (HF)

Few studies have also indicated that patients with advanced heart failure are more prone to developing cancer. This implies that there could be an association among CHIP, heart failure and tumor growth.^24,25 Abplanalp et al.,²⁶ studied patients with heart failure; it was found that those with CHIP mutations exhibited higher expression of proinflammatory cytokines, such as IL-1β, as well as their receptors, including the interleukin-6 receptor and the cellular receptor CD163. An increased expression of the Nod-like receptor protein3 (NLRP3) inflammasome complex was also observed in heart failure patients with CHIP. The researchers proposed that this heightened inflammatory state could potentially contribute to the increased mortality and morbidity in these patients if infected with the COVID-19. Additionally, peripheral blood cells from patients with HF harbouring DNMT3A mutations were found to exhibit increasingly inflamed transcriptomes, particularly with elevated levels of inflammatory cytokines such as IL-1β, IL-6, and IL-8.²⁷ Assmus et al.,²⁸ conducted a study involving 419 patients with heart failure to investigate the presence of CHIP mutations in DNMT3A and TET2. They found that more than half of the patients (56.2 %) were carriers of a CHIP mutation with a variant allele frequency (VAF) > 0.5 %. Additionally, 59 patients (14 %) harbored mutations in both genes. The five-year mortality rates were 18 %, 29 %, and 42 % in patients without any mutation, those with a single CHIP mutation, and those with mutations in both genes, respectively. Furthermore, recent studies have identified somatic mutations with lower frequency in non-classical driver genes such as CBL, CEBPA, PHF6, SMC1A, and SRSF2, which were also associated with increased mortality in heart failure patients, independent of the classic CHIP driver genes like DNMT3A or TET2. These mutations were notably not uncommon in younger heart failure patients.²⁹ Additionally, TET2 CHIP has recently been identified as a risk factor for heart failure with preserved ejection fraction (HFpEF).³⁰

1.9. CHIP and aortic stenosis

Aortic stenosis has traditionally been regarded as an age-related degeneration of the aortic valve. However, coronary artery disease and degenerative aortic valve disease share common risk factors, including age, hypertension, dyslipidemia, smoking, metabolic syndrome, chronic kidney disease, and diabetes, as well as similar pathogenic mechanisms. Mas-Peiro et al.³¹ studied 279 patients with aortic stenosis undergoing transcatheter aortic valve implantation (TAVI) to investigate the presence of two commonly observed CHIP mutations, namely DNMT3A and TET2. They found that 33 % of patients with severe aortic stenosis had a CHIP mutation. Those with CHIP mutations exhibited higher levels of pro-inflammatory cytokines and decreased survival rates after TAVI.³¹ Furthermore, an association was observed between clonal size and mortality; patients with a VAF ≥10 % experienced higher event rates compared to those with a VAF <10 %.

1.10. CHIP and transplantation

The role of CHIP and poor outcomes following cardiac transplantation is an intriguing issue.³² Scolari et al,³³ found that individuals with CHIP after transplant had higher chance to die and also had an increased risk to have cardiac allograft arteriopathy. It has been proposed that inflammation may mediate this response.

1.11. CHIP and arrhythmias

Lin et al found a modest association between AF and CHIP, suggesting altered calcium handling as a potential mechanism in a mouse model, potentially mediated through the NLRP3 inflammasome activation.³⁴ Recently, Schuermans et al identified a significant association between CHIP and supraventricular arrhythmias, bradyarrhythmias, and ventricular arrhythmias.³⁵ This association was found to be independent of coronary artery disease (CAD) and heart failure.

1.12. CHIP and miscellaneous CV diseases

Patients with CHIP have also been reported to have a higher risk of haemorrhagic and small vessel ischemic strokes.³⁶ An increased prevalence of CHIP was observed in patients with systemic lupus erythematosus (SLE) concerning age; however, no correlation with the incidence of SLE itself has been identified.³⁷ Studies examining the DNA methylation patterns associated with DNMT3A and TET2 CHIP have revealed distinct and directionally opposing patterns consistent with their regulatory roles. Uddin et al performed Mendelian randomization analyses, indicating a subset of DNA changes associated with DNMT3A and TET2 CHIP genes that may promote the risk for CAD.³⁸ Additionally, it has been noted that patients with HIV exhibit a two-fold increase in CHIP prevalence, suggesting that CHIP may contribute to the heightened cardiovascular risk among people living with human immunodeficiency virus³⁹

1.13. Mechanisms linking CHIP and CVD

Inflammation is a key factor in the pathophysiology of atherothrombotic CAD. Somatic mutations in blood cells can influence the risk of ASCVD in two ways: by increasing the likelihood of thrombosis or by exacerbating underlying atherosclerosis. In order to prove the causal relation of CHIP and atherosclerosis, various murine based research models have been done. Fuster et al,⁴⁰ studied the role of TET2 deficiency in mice by developing chimeric Ldlr-deficient mice implanted with 10 % TET2 deficient bone marrow and 90 % TET2 wild-type bone marrow. After approximately three months, they observed that 69 % of hematopoietic stem cells (HSCs) were TET2-deficient, indicating clonal expansion. They also found increased atherosclerosis in the aortic root and a higher presence of macrophages in the plaque compared to wild-type TET2 mice. Moreover, the mutant cells preferentially differentiated into macrophages within the atherosclerotic vascular wall. Additionally, increased levels of interleukin-1β (IL-1β) were noted in TET2-deficient mice when stimulated by oxidized LDL, tumor necrosis factor, and interferon gamma. Similar finding of increased aortic plaque burden was observed in heterozygous and homozygous mice with TET2 deficient allele.⁴¹ Activation of macrophages through various cytokines and chemokines have been proposed to cause monocyte adhesion and recruitment, proliferation of vascular smooth muscle cells and plaque destabilization.^40,41 Such response was reduced by NLRP3 inhibition. Inhibition of NLRP3, not only reduced IL-1β but also reduced aortic plaque size.^40,42 Hence, identification of such somatic mutations may lead to pattern shift in the management of patients with CVD. CHIP carriers also had increased serum levels of hsCRP, IL-6 and tumour necrosis factor alpha.⁴³

Another common CHIP mutation in JAK2 is V617F, where valine is substituted by phenylalanine at the 617th position. Granulocytes carrying this JAK2 mutation have been implicated in a pro-inflammatory, pro-plaque forming, and pro-thrombotic state. This mutation has been shown to activate neutrophils, leading to the formation of extracellular traps that facilitate neutrophil adhesion via β1 and β2 integrins.^44,45 Mutated macrophages with the JAK2 mutation have been associated with defective efferocytosis—the impaired clearance of dead leukocytes from atheroma—resulting in increased expression of various chemokines and cytokines. Additionally, erythrocytes harbouring the JAK2 mutation have been found to be more susceptible to erythrophagocytosis due to decreased expression of CD47. In atherosclerosis-prone mice, the JAK2 mutation was shown to promote increased neutrophil infiltration and activation, enhanced erythrophagocytosis, and defective efferocytosis. These processes collectively contribute to increased plaque size and heightened vulnerability to rupture.⁴⁶

1.14. CHIP genes and screening

Genetic association studies have consistently shown that CHIP has a polygenic and hereditary risk.⁴⁷ Currently, detecting and monitoring CHIP mutations present clinical challenges for both patients and physicians. CHIP can be identified through elective whole-genome sequencing during clinical trials, genome sequencing to assess conventional cardiovascular risks, genome sequencing in organ donors, or during the diagnosis of hematologic or solid malignancies.⁴⁸ In the future, cost-effective strategies for CHIP detection will need to be evaluated both systematically and strategically.⁴⁹ A survey by Sella et al revealed that young survivors of haematological malignancies and heart disease generally favoured CHIP screening.⁵⁰

1.15. Linking CHIP, inflammation and CVD - therapeutic potential

In the inflammatory process, it is well-established that macrophages infiltrate the arterial media, engulf lipids, and form foam cells. The evolving understanding of inflammation's role in the pathogenesis of CAD and aortic stenosis highlights its dual function—not only making the plaque more vulnerable but also contributing to its formation.⁵¹ Consequently, CAD, particularly acute coronary syndrome (ACS), is increasingly recognized as an inflammatory condition. This paradigm shift in understanding the immune system's involvement in CAD and ACS paves the way for new treatment strategies in atherosclerosis. A new frontier in the treatment of patients with acute coronary syndrome was postulated recently in CANTOS trial (Canakinumab Anti-inflammatory Thrombosis Outcome Study) which studied the effect of canakinumab, a therapeutic monoclonal antibody targeting interleukin-1β in patients with prior MI with high CRP.⁵² A sub-group analysis from CANTOS trial⁵³ found the prevalence of CHIP + to be 8.8 % with mean age of 66 years. In their exploratory analyses, it was observed that placebo-treated patients with a somatic mutation in either TET2 or DNMT3A (n = 58) had a greater scale of risk for MACE (HR = 1.76, p = 0.037). In contrast, an improved response to canakinumab was observed in CHIP (+) patients with TET2 mutant (HR = 0.36, p = 0.034). This led to a hypothesis that patients with CHIP (+) with mutations in TET2 and DNMT3A may do better with reduced MACE when treated with canakinumab.

Since CHIP ultimately leads to inflammasome activation, the cardiovascular effects related to CHIP can be mitigated by inhibiting the inflammasome. For instance, NLRP3 inhibition can be achieved by various agents acting at different levels.^54,55 NLRP3 requires a specific stimulus, referred to as a "priming" or "licensing" signal, to achieve full activation. NLRP3 priming can be classified into two categories: transcriptional priming, which is not required for NLRP3 activation, and post-translational modification (PTM) mediated priming, which is essential for the assembly of the NLRP3 inflammasome.^56,57 Key influencing factors include binding partners like NIMA related kinase 7 (NEK7),⁵⁸ PYD-only protein (POPs),⁵⁹ and CARD-only protein (COPs).⁶⁰ However, numerous questions persist about how different NLRP3 PTMs, such as phosphorylation and ubiquitination, are spatially and temporally coordinated during the assembly of the inflammasome. Newer molecules like MCC950 (CP-456,773) directly inhibits NLRP3 formation and function.⁶¹ Drugs like colchicine, glyburide, thiolutin and selenofast are known to inhibit inflammasome synthesis.^50,51

Downstream mediators of the inflammasome can also be targeted; for example, anakinra (an anti-IL-1 receptor antibody), tocilizumab (an anti-IL-6 receptor antibody), or ziltivekimab (an IL-6 inhibitor).⁶² Additionally, CHIP-related gene mutations can be targeted directly with anti-cancer drugs like azacitidine and decitabine, which inhibit TET2, or ruxolitinib, which inhibits JAK.⁶³ Colchicine is proven to reduce MACE in patients with CAD^64,65

Currently, various studies are being conducted to know the role of anti-inflammatory therapies in patients with CAD. ARTEMIS - A Research Study to Look at How Ziltivekimab Works Compared to Placebo in People With a Heart Attack [ClinicalTrials.gov number NCT06118281]⁶⁶ is done in patients with myocardial infarction, It's a global, multicentric, randomized control trial involving more than 600 sites to assess the efficacy of Ziltivekimab on major adverse cardiac events. It has started recruiting patients. The result of the same is expected in 2027.

2. Conclusion

CHIP represents a novel risk factor for CVD, and potentially the most significant contributor to residual inflammatory risk. By driving inflammation through the activation of the inflammasome, CHIP plays a pivotal role in the pathogenesis of CVD. Ongoing research is exploring therapeutic interventions targeting the inflammasome at various levels, its downstream cytokines, and CHIP-related mutations may offer promising avenues for reducing the inflammatory burden associated with CHIP and ultimately improving cardiovascular outcomes (Fig. 2).

Fig. 2.

Illustration of the connection between CHIP genes, inflammasomes and cardiovascular diseases. It also depicts the potential therapeutic targets to mitigate the inflammatory risk.

Declaration of competing interest

The authors declare the following financial interests/personal relationships which may be considered as potential competing interests:Dr. Nagendra Boopathy Senguttuvan reports financial support was provided by Indian Council of Medical Research. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.