Gene therapy and genome editing for lipoprotein disorders

Graphical Abstract

Graphical Abstract.

Cumulative lifelong exposure to elevated blood low-density lipoprotein cholesterol (LDL-C is a primary cause of atherosclerotic cardiovascular disease (ASCVD)), yet despite the availability of multiple lipid-lowering therapies, many patients fail to achieve or maintain target LDL-C levels due to limited access, poor adherence, or treatment discontinuation. CRISPR base editing is a new treatment approach designed to permanently inactivate cholesterol raising genes in the liver after a single intravenous infusion. The treatment consists of a lipid nanoparticle (LNP) containing mRNA encoding adenine base editing (ABE) and a guide RNA targeting the PCSK9 gene, conjugated with GalNAc for hepatocyte-specific delivery via asialoglycoprotein receptor (ASGPR) or LDL receptor (LDLR)-mediated endocytosis. Once inside the hepatocyte, the ABE is translated, binds to the guide RNA, and introduces a single base pair change (A-to-G) in the PCSK9 gene, without introducing a double-strand break in the DNA. This change disrupts a splice site leading to a stop codon in the mature PCSK9 transcript that effectively inactivates the gene. The intended result is sustained reduction of circulating PCSK9, increased LDLR expression, and lifelong lowering of LDL-C. This approach may address key unmet needs in ASCVD prevention by overcoming the limitations of chronic lipid-lowering therapy and reducing cumulative LDL-C exposure over the lifespan.

Abstract

Genetic factors play a critical role in the development of lipoprotein disorders, which significantly contribute to atherosclerotic cardiovascular disease (ASCVD). Traditional management of these conditions has relied on lipid-lowering therapies, which require lifelong adherence. Recent advancements in gene addition and editing technologies offer novel and potentially transformative approaches for treating lipoprotein disorders by targeting the relevant genetic pathways for each disease. This review revisits major monogenic and polygenic disorders of lipoprotein metabolism, including familial hypercholesterolemia, elevated lipoprotein(a), and familial chylomicronemia syndrome, and discusses the genetic-based therapies for management. RNA-based, gene addition and gene editing therapies, including Clustered Regularly Interspaced Short Palindromic Repeats, base editing and interventions whereby, are highlighted for their potential to provide durable treatments which overcome the adherence challenge. Integration of machine learning for risk prediction and the use of polygenic risk scores to enhance risk stratification further demonstrate the promise of personalized approaches, and overall potential for gene-based treatments to revolutionize ASCVD prevention and management.

Introduction

Advancements in gene editing technology hold the promise of revolutionizing the management of cardiovascular and metabolic disorders by targeting the genetically regulated pathways underlying these diseases.¹ Many lipoprotein disorders involving monogenic traits require lifelong therapies to reduce the risk of atherosclerotic cardiovascular disease (ASCVD) and other target-organ disease.^2–4 Even among patients receptive to initiating lipid-lowering therapies, lack of persistence is accompanied by higher rates of cardiovascular morbidity and mortality along with associated healthcare costs.² The challenges of treating asymptomatic conditions for life may be addressed with more durable treatment approaches, including RNA-based therapies and gene editing. This review was based on search of PubMed as a data source from inception to the present. In addition, we accessed databases of ongoing trials including ClinicalTrials.gov (http://clinicaltrials.gov/), the EU Clinical Trials register (https://www.clinicaltrialsregister.eu/) and the World Health Organization (WHO) International Clinical Trials Registry Platform Search Portal (http://apps.who.int/trialsearch/).

Major monogenic disorders requiring lifelong or decades of therapy

Homozygous familial hypercholesterolemia (HoFH) is a rare genetic disorder characterized by severely elevated low-density lipoprotein cholesterol (LDL-C) levels of above 10.3 mmol/L (400 mg/dL), leading to cutaneous and tendon manifestations, such as xanthelasmas and xanthomas, as well as premature ASCVD that often begins in childhood.⁵ Current treatments for HoFH include high-dose statins, ezetimibe, proprotein convertase subtilisin/kexin type 9 (PCSK9) inhibitors, the microsomal transfer protein inhibitor lomitapide, angiopoietin-like protein 3 (ANGPTL3)-directed therapy, and lipoprotein apheresis. However, low adherence as well as limited LDL-C efficacy in HoFH patients with markedly reduced LDL receptor (LDLR) activity,⁶ results in a low LDL-C goal achievement in most patients. A recent analysis of a global registry of HoFH patients indicated that the LDL-C goal was achieved in only 21% of patients in high-income regions and even fewer—3%—in non-high-income regions.⁷

Heterozygous familial hypercholesterolemia (HeFH), a co-dominant and highly penetrant monogenic disorder, affects a larger population, with an estimated worldwide prevalence of 1:200 to 1:300.⁸ HeFH is characterized by elevated LDL-C concentration from birth and, if untreated, leads to premature ASCVD that may begin as early as 20 to 25 years of age in men and 40 years of age in women.^2,4 Genes causative for HeFH include: LDLR, which codes for the LDL-R; APOB, which codes for apolipoprotein (apo) B, the major protein component of LDL particles; PCSK9 (gain-of-function mutation), which codes for the PCSK9 protein that terminates the lifecycle of the LDL-R; and far less commonly, APOE (an in-frame three base-pair deletion at position 167 in exon 4) and LDLRAP1.⁹ Although statins and other lipid-lowering therapies may be effective in managing HeFH, only ≈48% of the patients receive statins, and the majority of patients do not achieve guideline recommended LDL-C goals.²

Elevated lipoprotein(a) [Lp(a)] levels are an independent risk factor for ASCVD, peripheral arterial disease, and calcific aortic valve stenosis.¹⁰ Despite optimal control of modifiable risk factors through the use of LDL-C-lowering therapies, patients with elevated Lp(a) have higher rates of major adverse cardiovascular events when compared with those who have normal Lp(a) concentrations.¹⁰ Genome-wide association studies have established a correlation between single nucleotide polymorphisms in the LPA gene and elevated Lp(a) levels, significantly increasing the risk of coronary heart disease (CHD). Plasma Lp(a) concentrations are predominantly influenced by genetic factors, with more than 90% of the variance attributable to the LPA gene.^10,11 While Lp(a) was previously considered a monogenic disorder, its diverse genetic determinants suggest that a more complex inheritance pattern than a simple monogenic defect.^12,13 Consequently, individuals with elevated Lp(a) plasma levels, which remains the most significant determinant of Lp(a) derived atherogenicity, exhibit a heightened lifetime risk of cardiovascular events despite optimal LDL-C management. This highlights the critical importance of targeting Lp(a) for reducing ASCVD risk.^10,11

Familial chylomicronemia syndrome (FCS) is a rare autosomal recessive disorder caused by genetic variants affecting the lipoprotein lipase (LPL) pathway, characterized by severe hypertriglyceridaemia with triglyceride levels typically of 10 mmol/L (880 mg/dL) or greater and recurrent episodes of acute pancreatitis.^3,14 Current treatment options for FCS focus on strict restrictions in dietary fat and complete avoidance of alcohol, but these approaches may not be sufficient for many patients, highlighting the need for innovative therapies such as durable RNA-based therapies and gene editing.^15,16

Figure 1 summarizes the targetable genes in lipoprotein disorders and their different roles.

Figure 1.

Targetable genes in lipoprotein disorders. This illustration highlights the major targetable genes in lipoprotein disorders and their roles in lipoprotein metabolism. APOB encodes the structural protein Apolipoprotein B (ApoB), critical for assembling and secreting low-density lipoproteins (LDL), chylomicrons and very low-density lipoproteins (VLDL), facilitated by Microsomal Triglyceride Transfer Protein (MTP). Therapeutically, MTP inhibitors can reduce the production of ApoB-containing lipoproteins. LDLR encodes the LDL-receptor protein, which promotes LDL clearance, with therapies such as statins enhancing its hepatic expression to lower LDL-C. PCSK9 gene encodes the proprotein convertase subtilisin/kexin type 9 protein, which promotes LDL receptor degradation, and its inhibition by monoclonal antibodies or by reducing its translation with small interfering RNA (siRNA) prevents this process, to increase receptor recycling and reduce LDL-C. ANGPTL3 encodes the angiopoietin-like protein 3, which inhibits lipoprotein lipase (LPL) activity, reducing triglycerides breakdown from lipoproteins and thus increasing levels of LDL, VLDL and chylomicrons. By inhibiting and endothelial lipase (EL) it also reduces phospholipids hydrolysis, leading to increased levels of high-density lipoproteins (HDL) cholesterol. ANGPTL4 encodes the protein by the same name, which inhibits LPL as well, but is tissue-specific, primarily modulating lipid uptake in adipose tissue and muscles. APOC3 encodes apolipoprotein C-III, which inhibits LPL and hepatic uptake of triglyceride-rich particles, while antisense therapies target Apo-C3 to lower triglycerides. LPA gene is involved in the synthesis of lipoprotein (a), a particle composed of ApoB-100 covalently linked to apolipoprotein(a) via a disulphide bond and enriched with oxidized phospholipids (OxPL) that contribute to its pro-atherogenic and pro-inflammatory properties. siRNA therapies designed to reduce Lp(a) levels are currently being studied in phase 3 clinical trials. The diagram uses arrows to show regulatory pathways and interactions (solid arrows represents direct effect, dashed arrow represents indirect effect) genes (in boxes) from enzymes/transporter proteins (in hexagons)

Contribution of polygenic risk to monogenic disorders

In recent years, the interplay between traditional risk factors, polygenic predisposition, and monogenic disorders has become increasingly evident.¹⁷ Many monogenic disorders, traditionally attributed to single gene mutations, are now understood to be influenced by a myriad of common genetic variants that collectively contribute to disease susceptibility.¹⁸ These polygenic risk factors can modify the phenotypic expression of monogenic disorders, affecting disease severity, onset, and progression.¹⁸

Additionally, individuals with polygenic hypercholesterolemia have been shown to have higher cardiovascular risk, similar to monogenic hypercholesterolemia, when compared with individuals with matching LDL-C levels but no apparent genetic predisposition.¹⁹ The importance of polygenic risk scores (PRS) has been studied in other lipoprotein disorders as well. In a recent prospective study in 703 patients who underwent coronary angiography, after adjustment for cardiac risk factors and CAD, the odds of calcified aortic valve disease increased with increasing LPA genetic risk score (OR 1.054 per 10-unit increase, 95% CI: 1.024–1.086; P < .001).²⁰ The recognition of the polygenic nature of lipoprotein disorders has led to new terminology to describe disease phenotypes. For example, multifactorial chylomicronemia syndrome (MCS) and familial combined hyperlipidaemia (FCH) are now used to describe polygenic severe hypertriglyceridaemia and mixed hyperlipidaemia, respectively.^21,22

The complexity of genetic contributions to disease presents an opportunity for personalized therapeutic strategies that consider both monogenic and polygenic influences. Integrating PRS with traditional monogenic diagnoses could enhance risk prediction.¹⁸ Moreover, PRS may assist in refining cardiovascular risk prediction by integrating genetic data into standard clinical risk assessments. One example is the SCORE2 assessment. By multiplying the absolute risk estimated by SCORE2 with the odds ratio derived from PRS, one can estimate the total genetic risk and generate a more accurate prediction of patient susceptibility to cardiovascular events.²³ In the UK Biobank study, this multiplicative model reclassified 9.55% of individuals from the intermediate-risk group (n = 145 337) to the high-risk category, increasing the population in this high-risk group by 56.6%. Among those reclassified to high risk by the PRS factor, 8.08% experienced incident CVD, nearly double the 4.08% incidence observed in individuals who remained at intermediate-risk.²³ Indeed, a recent European Society of Cardiology (ESC) position paper indicates that inclusion of PRS data may have the strongest implications on decision-making in persons with an intermediate clinical risk who consider medical treatment.²⁴ Studies have shown that LDL-C-lowering lifestyle modifications and pharmacotherapy tend to be more effective in lowering ASCVD events among individuals with a higher PRS.^25–27 In a meta-analysis of studies including the West of Scotland Coronary Prevention Study (WOSCOPS), Anglo-Scandinavian Cardiac Outcomes Trial (ASCOT), and Justification for the Use of Statins in Prevention: an Intervention Trial Evaluating Rosuvastatin (JUPITER) primary prevention trials, the relative risk reduction for those at high genetic risk was 46%, compared with 26% in all other participants (P for heterogeneity = 0.05). Across all three studies, the absolute risk reduction with statin therapy was 3.6% (95% CI: 2.0–5.1) for the high genetic risk group, compared with 1.3% (95% CI: 0.6–1.9) for all others.²⁸

However, the efficacy of PRS as a risk stratifying tool remains controversial. In a secondary analysis of the PRS Catalogue, incorporating both conventional risk factors and PRS was shown to prevent 974 events compared with 957 events prevented by a conventional risk factor model alone. This gain of 17 patients means the number needed to genotype to prevent one additional event is 5882.²⁵ Of note, risk prediction using PRS is most reliable at the extremes, effectively identifying individuals at very high or very low genetic risk for cardiovascular disease.²⁵ This enhanced risk prediction can guide clinical decision-making help identify patients who may benefit from novel therapies, and potentially improve patient outcomes in cardiovascular and metabolic health.

Role of machine learning in predicting coronary artery disease risk and major adverse outcomes

The advent of artificial intelligence (AI) and machine learning (ML) techniques has revolutionized the predictive modelling landscape for coronary artery disease (CAD) and major adverse outcomes. ML algorithms excel at analyzing large datasets to identify complex patterns and interactions among numerous risk factors, providing a more accurate individualized cardiovascular risk assessments compared with traditional statistical methods.²⁹ These techniques can integrate diverse data types, including genetic information, clinical features, imaging data, and lifestyle factors, to enhance predictive power. For example, ML models such as random forests, support vector machines, and neural networks have been successfully applied to predict CAD risk, demonstrating superior performance in identifying high-risk individuals and anticipating adverse events like myocardial infarction (MI) and stroke.^29–31 Additionally, ML can continuously learn and adapt from new data, improving its predictive accuracy over time.³¹ Recently, in a meta-analysis using an ML-based marker for CAD, researchers identified rare and ultrarare coding variants in 17 genes from diverse biobank samples. This analysis revealed novel genetic mechanisms affecting CAD beyond traditional risk factors, including processes like oxysterol transportation and mitotic spindle assembly.³² This approach, utilising continuous in-silico markers, which are computed from digital data rather than physical experiments, can potentially improve the statistical power and uncover additional genetic insights into CAD, suggesting that ML-based digital markers can contribute to discovery of rare genetic variants, which might serve as drug targets for genetic therapies.³³

The integration of ML into clinical practice promises advanced personalized medicine, enabling clinicians to tailor interventions and management strategies more effectively based on precise risk stratification.

Unmet need for better management of cardiovascular disease

Despite advancements in cardiovascular-related therapy, a significant unmet need remains in ASCVD prevention and management, primarily due to issues with treatment adherence. A considerable proportion of patients are not able to adhere to the medication regimens as recommended, which is associated with higher rates of hospitalisations and adverse cardiovascular events.³⁴ The European Society of Cardiology (ESC)/European Atherosclerosis Society (EAS) 2019 guidelines for dyslipidemias and the ESC 2021 guidelines for cardiovascular prevention recommend targets of ≤1.4 mmol/L (55 mg/dL) for patients with ‘very high cardiovascular risk’, such as patients with prior cardiovascular events or HeFH patients with known ASCVD or a major risk factor for ASCVD.^35,36 For patients at ‘high risk’, including HeFH patients without ASCVD or a major risk factor, a target of LDL-C ≤ 1.8 mmol/L (70 mg/dL) is recommended.^35,36 Although the calculated percentage of patients who could achieve LDL-C targets with a combination of high-dose statin, ezetimibe, and a PCSK9 inhibitor is up to 86%, registry data from very-high-risk patients with HeFH indicates that only ∼2% achieve these targets within current clinical practice.³⁷ Furthermore, among very-high-risk HeFH patients treated in lipid clinics and prescribed triple lipid-lowering therapy with a high-dose statin, ezetimibe, and PCSK9 inhibitor, only 45% reached the LDL-C goal of ≤1.4 mmol/L (55 mg/dL).³⁸

Maintaining adherence to medication for asymptomatic conditions like elevated LDL-C poses a challenge for both patients and physicians. Non-adherence stems from a variety of reasons and associated factors, which can be broadly categorized into patient-related, physician-related, and health system-related causes, with patient-related factors often playing the most significant role.^39,40 Among patient-related factors are younger age, non-Caucasian ethnicity, and low awareness of the necessity for treatment.⁴¹ Other factors contributing to poor adherence include complex medication schedules, polypharmacy, adverse drug reactions, forgetfulness, poor communication with healthcare providers, and lack of patient education and support.^34,42 In the case of statins, for example, it has been shown that although patients acknowledge the importance of adhering to therapy, they have stronger concerns regarding side effects, including short and long-term effects. Moreover, the public perception of medication safety plays a significant role in statins adherence.⁴⁰ Low persistence with cholesterol lowering therapies highlights the need for interventions aimed at improving adherence, such as patient-centered care approaches, simplified treatment regimens, and digital health technologies to monitor and encourage compliance.³⁴ Behavioural studies have previously demonstrated that the preference for lipid-lowering agent among patients diagnosed with hypercholesterolaemia is based on treatment regimen convenience (lower dosage and less frequent dosing), effectiveness, side effects, health provider’s recommendations, drug interactions, and out-of-pocket costs.⁴² In a recent survey of 484 patients with hypercholesterolaemia and/or ASCVD, 46% of patients preferred a once-daily oral pill and 35% indicated a preference for one-time gene editing treatment. Subcutaneous injections were less preferred (14% favoured twice-annual injection and 4% preferred a twice-monthly injection).⁴³ Addressing these preferences is crucial for improving the management of CVD and reducing the burden of cardiovascular events on patients and the healthcare system. Implementing gene editing therapies, especially those that only require a single administration in a person’s lifetime, has the potential to overcome the adherence obstacle in cardiovascular disease prevention.

Gene addition for lipid disorders

Introducing gene addition

Gene therapies are a diverse class of treatment. Gene addition or gene replacement describes therapies where a functional copy of a gene is delivered to cells for a therapeutic benefit. Delivery of gene addition therapies may be ex vivo or in vivo depending on the target cell type. Ex vivo autologous gene addition approaches require cell collection from the patient. Commonly, haematopoietic stem cells or T cells are the target. Patient cells are transduced with a viral vector in a laboratory setting (typically retrovirus or lentivirus) with the result being a set of gene-modified cells carrying copies of the functional gene that can be readministered to the patient. In vivo gene addition therapies involve direct administration of the viral or non-viral vector to the patient. Different vectors are engineered to have tropism for select tissue types that dictate where the gene copies are added. Viral vectors, specifically adeno-associated virus (AAV), are the predominant delivery system for in vivo gene addition.^44–46

AAV vectors for genetic therapies

AAV vectors have revolutionized genetic therapies by providing a highly efficient and relatively safe method for delivering therapeutic genes to target cells.⁴⁶ AAV vectors are derived from the non-pathogenic parvovirus, AAV, which has the advantage of minimally integrating into the host genome, thereby reducing the risk of insertional mutagenesis.⁴⁶ The mechanism of action involves the binding of AAV vectors to specific receptors on the target cell surface, followed by entry through endocytosis. After entering the cell, AAV vectors escape from the endosome into the cytoplasm and are transported to the nucleus. Here, the AAV capsid uncoats and releases the therapeutic DNA, which exists as an episome (a stable, extrachromosomal element) in the nucleus. This DNA is then transcribed and translated into the therapeutic protein, correcting the genetic defect.⁴⁶ AAV vectors are engineered to carry a therapeutic gene flanked by AAV inverted terminal repeats (ITRs), which are essential for the replication and packaging of the viral genome during vector production. The ability to target specific tissues and sustain long-term expression of the therapeutic gene makes AAV vectors highly effective for treating chronic conditions such as metabolic diseases.

Experience with gene addition therapy for lipoprotein disorders

First attempts to replace genes in the field of lipoprotein disorders were initiated as early as the early 1990s, when the University of Pennsylvania initiated a gene therapy program for HoFH. Initially, Grossman et al.⁴⁷ conducted the first gene therapy trial for HoFH, using an ex vivo approach to transfer LDLR-expressing retroviruses into hepatocytes, which were then re-infused into patients. Despite some LDL-C reductions (6%–25% in three subjects), the results were largely disappointing, with minimal transduction efficiency and transient metabolic effects.⁴⁷ Thereafter, the program focused on delivering a functional LDLR gene to liver cells using first-generation adenoviral vectors. These vectors were modified to prevent replication but retained the ability to elicit strong immune responses. Early trials in the late 1990s revealed significant safety concerns, particularly with high doses required for therapeutic effects. Adenoviral vectors were associated with severe inflammation, hepatotoxicity, and immune activation, which could lead to systemic complications, including multi-organ failure. The tragic outcome of the 1999 Jesse Gelsinger case, in which an adenoviral vector was used to treat ornithine transcarbamylase deficiency and resulted in a rapid onset of fever, clotting abnormalities, and fatal liver failure, highlighted the risks of this technology.⁴⁸ This incident led to heightened regulatory scrutiny and the eventual discontinuation of LDLR gene therapy programs employing these vectors, as the safety profile was deemed unacceptable for clinical use.^49,50

Despite these early setbacks, advancements in gene therapy have continued. Novel AAV vectors have shown promise in preclinical models for delivering functional LDLR genes. On the past decade the U-Penn program was reinitiated and a progression has been made in LDLR replacement using recombinant AAV vectors. Recombinant AAV vectors, with most native sequences removed except for the essential ITRs, maximize packaging capacity while minimising immunogenicity and cytotoxicity.⁵¹ In preclinical studies on humanized HoFH mice models, a single intravenous injection of AAV8.TBG.hLDLR was found as efficient and safe, with an increase in LDLR protein, total cholesterol and non-HDL cholesterol decrease from 808 to 353 mg/dL and from 675 to 255 mg/dL, respectively, by day 35 after treatment. Moreover, in HeFH humanized mice there was a statistically significant reduction in total cholesterol from 325 to 200 mg/dL.^52,53 Based on this preliminary safety and efficacy data, a first-in-human phase 1/2 human clinical trial with the recombinant AAV8 vector AAV8.TBG.hLDLR commenced in 2016 (ClinicalTrials.gov Identifier: NCT02651675). A whereby human LDLR cDNA was packaged into rAAV8 and used to treat patients with HoFH. The trial concluded in November 2020, but its results have not yet been published, and a 2020 press release announced that the sponsor, Regenxbio (Rockville, Maryland, USA), was ceasing internal clinical development of the product.⁵¹

Overview of successful gene addition therapies

The first European Medicines Agency (EMA)-approved AAV gene replacement therapy, alipogene tiparvovec (Glybera®, UniQure, formerly known as Amsterdam Molecular Therapeutics (AMT), Amsterdam, Netherlands), was approved in 2012 in Europe for patients with lipoprotein lipase deficiency (LPLD).⁴⁶ This rare condition results from monogenic autosomal recessive mutations in the LPL gene that lead to a complete absence of the catalytically active LPL enzyme. Due to chylomicron accumulation, LPLD is characterized by severe hypertriglyceridaemia and related complications, including eruptive xanthomas, hepatosplenomegaly, and acute pancreatitis. Alipogene tiparvovec utilizes an AAV1 capsid to deliver a human gain-of-function LPL S447X (hLPLS447X) transgene driven by a cytomegalovirus promoter. By replacing the absent protein, alipogene tiparvovec led to significant reductions in plasma triglycerides of 60% as well as of acute pancreatitis episodes.⁵⁴ Nevertheless, mainly due to high costs as well as suboptimal vector design and formulation and manufacturing processes, alipogene tiparvovec was withdrawn from the European market in 2017 and the plans for global expansion were abandoned.⁵⁴ Currently, novel AAV-hLPLS447X vectors are being studied for LPLD. Specifically, AAV8 pVR59 was recently identified and demonstrated significantly higher efficacy of up to 10- to 100-fold lower doses needed, compared with an AAV1 vector based on Glybera, when delivered intramuscularly or intravenously, respectively, in mice with LPLD.⁵⁴

Gene addition therapies continue to grow in fields beyond cardiovascular disease and metabolism. In 2016, Strimvelis® (Orchard Therapeutics, London, United Kingdom), a retroviral ex vivo gene addition therapy, was EMA-approved for severe combined immune deficiency. Other hematological conditions were next to follow, with US Food and Drug Administration (FDA) approval in 2017 of ex vivo chimeric antigen receptor T-cell (CAR-T) therapies for the treatment of acute lymphoblastic leukaemia and large B cell lymphoma.⁴⁶ Subsequently, the FDA approved gene addition therapies for transfusion-dependent β-thalassaemia and for sickle cell.⁵⁵ In the in vivo setting, an AAV gene addition therapy was approved by the FDA for the treatment of spinal muscular atrophy, onasemnogene abeparvovec (Zolgensma®, Novartis, Basel, Switzerland) in 2019.⁴⁶ More recently, multiple AAV gene addition therapies have been approved in the U.S. and Europe for the treatment of haemophilia A and B.

Challenges and limitations with gene addition therapy

Despite the success of in vivo gene addition approaches, several key challenges remain, including immune responses to AAV vectors, the need for precise vector targeting, and the high cost of therapy. The host immune response is a major obstacle for AAV vectors and can prevent effective long-term therapeutic gene expression. The host immune response comprises a cytotoxic T-lymphocyte response to the AAV capsid as well as to the therapeutic protein itself. Additionally, it includes antibodies, both neutralising antibodies (NAbs) to AAV virions and alloantibodies to therapeutic proteins. Moreover, an innate immune response to AAV transduction is formed over time.⁴⁶ The immune response to AAV vectors means that patients are limited to a single course of treatment and generally cannot be re-dosed if there is insufficient efficacy from a first dose.

Long-term durability of AAV vector-induced gene expression and additional potential off-target effects are areas of ongoing research and development. Addressing these challenges requires a multidisciplinary approach, incorporating advancements in vector design, immunomodulation, and manufacturing processes to ensure broad and sustained therapeutic efficacy.

Gene editing for cardiovascular and metabolic diseases

Existing or emerging therapies targeting genetic pathways

Gene editing is a different approach to genetic medicine that holds great promise for the treatment of cardiovascular and metabolic diseases. Unlike gene addition, gene editing approaches do not add copies of genes to cells but instead use nucleases or other engineered proteins to make targeted changes to host cell DNA. In the cardiometabolic space, targets for genetic therapies include PCSK9, ANGPTL3, ANGPTL4, APOC3 (apolipoprotein C-III, or apoC-III), and LPA. As noted above, the protein products of these genes play crucial roles in lipoprotein metabolism and cardiovascular risk, and some of them are targets of established treatments for hyperlipoproteinemias [Table 1].

Table 1.

Genetic targeted therapies for lipoprotein disorders

Pathway	Agent	Mode of action	Frequency of administration	Main efficacy endpoints	Cardiovascular outcome data
PCSK9	Inclisiran	siRNA	Biannually	LDL-C reduction up to 50%	Phase 3 CVOT are ongoing (ORION-4, VICTORION 2-PREVENT)56
ANGPTL3	Vupanorsen	ASO	Monthly	Triglycerides reduction up to 53%	Phase 2 completed (TRANSLATE-TIMI 70),57 program discontinued.
	Zodasiran	siRNA	Monthly	Triglycerides reduction up to 74%, LDL-C reductions up to 48%	Phase 2 in patients with mixed hyperlipidaemia completed (ARCHES-2)58,59 Phase 2 in HoFH interim data presented (GATEWAY).57 Phase 3 in HoFH was launched.60
	Solbinsiran	siRNA	Monthly	Triglycerides reduction up to 86%	Phase 1 completed.61 Phase 2b study (PROLONG-ANG3, NCT05256654) completed but not published yet.
ANGPTL4	Lipisense	ASO	Weekly	Triglycerides reduction up to 48% total cholesterol reductions up to 56%, reduced atherosclerotic lesion size (−86%)	Preclinical studies in mice were completed, phase 1 completed.62 Phase 2a in patients with severe HTG and T2DM is underway.
APOC3	Volanesorsen	ASO	Weekly	Triglycerides reduction up to 77% (APPROACH) in FCS and 71% in MCS (COMPASS)	Phase 3 laboratory trials completed63 [approved in Europe for FCS], CVOT not planned.
	Olezarsen	ASO	Monthly	Triglycerides reduction up to 53% in moderate HTG with high cardiovascular risk or severe HTG, and 43% in FCS	Phase 3 FCS study completed.64 FDA has accepted for Priority Review the NDA for FCS. Phase 2b in moderate HTG and high CV risk completed.65 Phase 3 is ongoing.66
	Plozasiran	siRNA	Monthly	Triglycerides reduction of up to 62% in mixed hyperlipidaemia, 57% in severe HTG and 80% in chylomicronemia	Phase 2 trials completed;67,68 Phase 3 in FCS completed;69 Phase 3 in mixed hyperlipidaemia and severe HTG enrolling (MUIR, SASHTA, CAPITAN).70–72
LPA	Pelacarsen	ASO	Monthly	Lp(a) reduction up to 92%	Phase 3 CVOT (HORIZON-Lp(a)) and phase 3 aortic valve stenosis (CAVS) are ongoing.73,74
	Olpasiran	siRNA	Quarterly	Lp(a) reduction up to 101.1% (placebo-adjusted)	Phase 3 CVOT is ongoing (OCEAN(a)).75
	Lepodisiran	siRNA	Biannually or annually	Lp(a) reduction up to 100.5% (placebo-adjusted)	Phase 3 CVOT is ongoing (ACCLAIM).76
	Zerlasiran	siRNA	Every 16 or 24 weeks	Lp(a) reduction up to 98% (APOLLO)	Phase 2 is completed (ALPACAR-360).77

ANGPTL3, angiopoietin-Like 3; ANGPTL4, angiopoietin-Like 4; APOC3, apolipoprotein C-III; ASO, antisense oligonucleotide; CV, cardiovascular; CVOT, cardiovascular outcomes trial; FCS, familial chylomicronemia syndrome; FDA, U.S. Food and Drug Administration; HTG, hypertriglyceridaemia; LDL-C, low-density lipoprotein cholesterol; Lp(a), lipoprotein(a); MACE, major adverse cardiovascular events; MCS, mixed chylomicronemia syndrome; NDA, new drug application; PCSK9, proprotein convertase subtilisin/kexin type 9; siRNA, small interfering RNA; T2DM, type 2 diabetes mellitus.

For example, PCSK9 inhibitors have demonstrated effectiveness in lowering LDL-C levels and cardiovascular morbidity. PCSK9 monoclonal antibodies (mAb) have become an important treatment for patients with hypercholesterolaemia for both primary and secondary prevention of cardiovascular disease (see Supplementary data online, Table S1).^78–80 Their mechanism of action is to prevent the degradation of LDL receptors (LDLR) in the hepatocytes. Inhibition of the PCSK9 protein promotes LDLR recycling to the hepatocyte surface, thereby reducing plasma levels of LDL-C. More recently, a small interfering RNA (siRNA) therapy targeting PCSK9, inclisiran, was approved. Inclisiran reduces LDL-C by 48% (placebo adjusted) in patients with HeFH and 50%–55% in patients with established ASCVD.^81,82 With a twice-yearly frequency of dosing, administered by a healthcare provider, this agent serves as a solution for many patients who have been non-adherent to oral therapy or are incapable of self-injections.

Another important drug target for cardiovascular disease prevention is ANGPTL3. This protein, which is mainly expressed in the liver, is an inhibitor of LPL, endothelial lipase, and hepatic lipase. Enhanced by ANGPTL8, which is primarily expressed in the liver and adipocytes post-prandially, ANGPTL3 inhibits LPL efficiently in various tissues, including the heart and skeletal muscle. Genetic studies indicate that loss of function variants in the ANGPTL3 gene are linked to lower levels of LDL-C, VLDL-C, and HDL-C, along with a reduced risk of developing CAD.^83–85 Importantly, the LDL-C lowering achievable with ANGPTL3 inhibition occurs via a mechanism of action that—unlike most currently approved lipid-lowering therapies such as statins, ezetimibe, and proprotein PCSK9 inhibitors—is independent of the LDLR. Treatment with ANGPTL3 mAb in HoFH patients lowers LDL-C by 47% with equal efficacy in patients with and without residual LDLR activity (see Supplementary Data online, Table S1).⁸⁶ Similarly, siRNA inhibition of ANGPTL3 is currently under investigation in HoFH patients. The AROANG3-2003 GATEWAY study (NCT05217667) is a Phase 2 open-label trial evaluating the efficacy, safety, and tolerability of zodasiran, a siRNA targeting ANGPTL3, in 18 patients with HoFH, with a primary endpoint of LDL-C percent change from baseline. Preliminary data presented in May 2023 at the European Atherosclerosis Society (EAS) scientific session showed mean reductions at week 20 of 48.1% and 44.0% in LDL-C, 39.2% and 34.5% in apoB, and 82.7% and 80.1% in ANGPTL3 for 200 and 300 mg doses, respectively.⁵⁸

SiRNA inhibition of ANGPTL3 was also shown to be effective among patients with mixed hyperlipidaemia. In the recent phase 2b ARCHES-2 trial, zodasiran significantly reduced triglycerides, remnant cholesterol, and apoB levels in patients with mixed hyperlipidaemia, with reductions up to 63%, 56%, and 20%, respectively, compared with placebo.⁵⁹ Solbinsiran, another ANGPTL3 siRNA agent, reported to produce dose-dependent reductions in ANGPTL3, triglycerides, non-HDL-C, and apoB, by up to 86%, 73%, 46% and 36%, respectively in patients with mixed dyslipidemia. Results from the recently completed phase 2b study are yet to be published.⁶¹

ANGPTL4 is another protein which is currently being studied for its role in lipoprotein metabolism, in the arena of cardiovascular and diabetes prevention. ANGPTL4 is primarily expressed in adipose tissue, liver, muscle, macrophages and intestine, and by inhibiting LPL it increases levels of circulating glucose and triglycerides under fasting conditions. Since ANGPTL4 has been associated as highly correlated with CHD, ANGPTL4 inhibition is currently studied as another target for novel therapies for cardiovascular prevention.⁸⁷ Human genetic studies suggested that targeting ANGPTL4 has the potential to lower the risk of CAD, as well as provide added benefits including improved glycemic balance and triglycerides plasma levels in patients with type 2 diabetes mellitus.^87,88 Recently, liver-targeted ANGPTL4 silencing with antisense oligonucleotide (ASO) was reported to potently reduce plasma triglycerides (−48%) and total cholesterol (−56%) as well as reduced atherosclerotic lesion size (−86%) and improved lesion stability in APOE*3-Leiden.CETP mice, and liver-targeted ANGPTL4 silencing was reported to be well-tolerated in non-human primates.⁸⁹ Phase 1 in 24 healthy participants who received four repeated doses of either Lipisense® or a placebo, resulted in up to 29% reduction in plasma levels of ANGPTL4 with Lipisense® vs. placebo.⁶² Of note, other strategies of ANGPTL4 inhibition as monoclonal antibodies were previously discontinued due to concerns regarding development of mesenteric lymphadenopathy secondary to granulomatous lipid accumulation in treated mice and monkeys.⁹⁰ However, MAR001, a monoclonal antibody targeting ANGPTL4, demonstrated recently significant reductions in remnant cholesterol and triglycerides in a Phase 2a trial involving 55 participants with hypertriglyceridaemia. The treatment was well tolerated with no serious adverse events, and the sponsor plans to present full results at an upcoming medical meeting while preparing for a Phase 2b trial in early 2025.⁹¹

ApoC-III has recently gained attention as a promising pharmacological target for the treatment of hypertriglyceridaemia. Evidence from Mendelian randomisation studies reveal that individuals with APOC3 null alleles tend to have lower triglycerides levels and a decreased risk of cardiovascular disease over their lifetime.⁹² A recent phase 3 randomized controlled study in FCS patients demonstrated a 43.5% reduction in triglycerides with the highest dose of olezarsen, an ASO RNA inhibitor of APOC3. Acute pancreatitis risk was reduced significantly (mean rate ratio of olezarsen groups vs. placebo was 0.12; 95% CI: 0.02 to 0.66).⁶⁴ Moreover, in another phase 3 trial among patients with moderate hypertriglyceridaemia and increased cardiovascular risk, olezarsen significantly reduced levels of triglycerides, apoB, and non-HDL cholesterol.⁶⁵ A recently completed phase 3 trial in patients with clinically or genetically confirmed FCS with plozasiran, an siRNA inhibitor of APOC3, demonstrated up to an 80% median reduction in triglycerides levels at month 10, as well as a statistically significant 83% reduction in the incidence of acute pancreatitis compared with placebo.⁶⁹ The triglyceride lowering and prevention of acute pancreatitis were similar in patients with genotypically-confirmed FCS and severe hypertriglyceridaemia without biallelic loss of function mutations in LPL.⁶⁹

Currently, multiple cardiovascular outcomes trials are evaluating whether Lp(a)-lowering RNA-targeted therapies reduce the risk of ASCVD events. Pelacarsen, a second-generation ASO, has shown promising results in reducing Lp(a) levels in phase 1/2a and 2b trials, with reductions up to 92%.¹⁰ siRNA therapies including olpasiran, zerlasiran, and lepodisiran have demonstrated significant Lp(a) reductions, with olpasiran achieving up to 101.1% placebo-adjusted reduction in the phase 2 OCEAN(a) DOSE trial, lepodisiran up to 97% reduction with the maximal dose in the phase 1 study,⁹³ and zerlasiran showing a 96.4% reduction in the recently published phase 2 trial.⁷⁷ A small-molecule strategy targeting Lp(a) with muvalaplin is currently under investigation (see Supplementary Data online, Table S1).⁹⁴ Phase 3 cardiovascular outcomes trials with Lp(a) inhibitors are currently ongoing.¹⁰

Introducing CRISPR nuclease editing, base editing, and reverse transcriptase editing techniques

Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) nuclease editing, and base editing are two revolutionary techniques in gene editing. CRISPR nuclease editing uses the Cas9 enzyme to create double-strand breaks at specific DNA locations, which are then repaired by the cell’s machinery via an error-prone process, predominantly leading to small insertions or deletions (indels) in the genetic code that can disrupt gene function [Figure 2]. Base editing, however, involves the direct chemical alteration of DNA bases without the requirement for double-strand breaks, offering the potential for precise and specific single base pair changes [Figure 3]. Both techniques can inactivate genes implicated in cardiovascular and metabolic diseases, with base editing providing a potential advantage in reducing off-target effects and enhancing safety profiles. The recently emerging reverse transcriptase (RT-) editors are gene editing tools that can introduce all types of point mutations, small insertions, and deletions without double-strand breaks or DNA donor templates. They consist of a Cas9 nickase fused with an RT, and guide RNAs (pegRNAs) that both target the site and encode the desired edit.^96,97

Figure 2.

CRISPR/Cas9 nuclease editing. The protospacer-adjacent motif (PAM) in the DNA and spacer sequence in the guide RNA direct CRISPR/Cas9 to a specific genomic site. There, it generates a double-strand break (indicated by arrows pointing to DNA strands) that is repaired by one of two repair mechanisms: non-homologous end joining (NHEJ) or homology-directed repair (HDR). In NHEJ, free DNA ends are ligated together, which can restore the original DNA sequence or introduce insertions or deletions. This strategy is suitable for disrupting genes or non-coding elements in the genome. HDR enables more precise changes but requires the addition of a template, which reduces its efficiency. Adapted with permission from the Journal of Clinical Investigation.⁹⁵

Figure 3.

Base editing. Tethering of an editing domain (cytosine deaminase) to catalytically impaired Cas9 (either dCas9 or a nickase form of Cas9) can result in site-specific alteration of cytosine-guanine base pairs to thymine-adenine base pairs without the need for double-strand breaks. Adapted with permission from the Journal of the American College of Cardiology.¹

In the past year gene editing therapies have started to score regulatory approvals. Exagamglogene autotemcel (exa-cel [Casgevy, Vertex Pharmaceuticals/CRISPR Therapeutics]) is the first FDA- and EMA-approved gene editing therapy, indicated for treating sickle cell disease and transfusion-dependent beta thalassaemia in patients aged 12 and older. This ex vivo CRISPR-Cas9-based therapy modifies haematopoietic stem cells so that they produce red blood cells with fetal haemoglobin (HbF).⁵⁵ In patients with sickle cell disease, increased levels of HbF prevent the sickling of red blood cells and thereby reduce vaso-occlusive events.⁹⁷ In a recently published phase 3 clinical study, this agent eliminated vasoocclusive crises in 97% of patients with sickle cell disease for a period of 12 months or more.⁹⁸ An additional phase 3 study with exa-cel demonstrated transfusion independence achieved in 32 of 35 treated patients with β-thalassaemia.⁹⁷ Other promising investigational CRISPR-Cas9 based gene-editing medicines are currently being evaluated in Phase 3 pivotal studies for transthyretin amyloidosis associated cardiomyopathy and neuropathy (NTLA-2001) and hereditary angioedema (NTLA-2002). These agents have shown decreases in serum TTR protein and reduced angioedema attacks in phase 1/2 studies in patients with TTR amyloidosis and hereditary angioedema, respectively.^99–101 These advancements represent a major step forward in gene editing applications, providing new targeted treatment options for patients with these challenging genetic conditions.

Efficacy of gene editing for lipids disorders in animal models

Numerous studies have demonstrated the efficacy of gene editing in animal models for cardiovascular and metabolic diseases, including their ability to reduce blood lipid levels in animal models. For instance, CRISPR-Cas9-mediated knockout of PCSK9 by introducing non-sense or splice-site mutations in mice and in non-human primates has resulted in significantly lower cholesterol levels, providing a model for potential human therapies.⁹⁶ Two studies conducted in cynomolgus macaques used adenine base editors (ABE) to inactivate the PCSK9 gene, resulting in significant reductions in serum PCSK9 and LDL-C. The ABEmax editor achieved 30% DNA editing, 40% reduction in PCSK9, and 20% reduction in LDL-C,¹⁰² while the ABE8.8 editor achieved 66% DNA editing, as well as 90% and 60% reduction in PCSK9 and LDL-C, respectively,¹⁰³ with minimal off-target effects observed in both studies.⁹⁶

Similarly, editing the ANGPTL3 or APOC3 gene in animal models can generate favourable changes in lipid profiles and reduce atherosclerosis. In homozygous LDLR mutant mice, it was shown in vivo that AAV-CRISPR/Cas9-mediated gene editing to correct the LDLR mutation resulted in decreased atherosclerosis and macrophage infiltration compared with placebo, as well as significant reductions in serum total cholesterol, LDL-C, and TG.¹⁰⁴

Introduction of specific agents under development

Among the promising agents under development, Verve Therapeutics is developing in vivo base editing medicines with the goal of inactivating PCSK9 or ANGPTL3. The therapies consist of an mRNA encoding an ABE and guide RNA (gRNA) targeting the respective gene packaged in a lipid nanoparticle (LNP) for delivery to the liver. In hepatocytes, the ABE inactivates the target gene by introducing a precise single base pair change (A-to-G) that disrupts the translation of the gene.¹⁰⁵ This approach has the potential to provide a durable solution for patients with HeFH, as well as polygenic hypercholesterolaemia, and might be useful for patients with low adherence to therapy and for patients in whom statins are deferred due to adverse side effects. Studies conducted with VERVE-101 in non-human primates showed 83% and 69% reductions in PCSK9 and LDL-C levels, respectively, which were durable 476 days after dosing.¹⁰⁶

Initial proof-of-concept data for this approach in humans was provided in the Heart-1 trial, Phase 1b study of VERVE-101 (NCT05398029). This study has enrolled 13 patients with a history of both HeFH and pre-existing ASCVD with administered doses ranging from 0.1 to 0.6 mg/kg.^107,108 Interim results reported to date allow for three conclusions. First, dose-dependent reductions from baseline of up to 73% for LDL-C and up to 84% for PCSK9 were observed in individual patients, consistent with saturating on-target editing of both chromosomes in nearly all hepatocytes of these individuals. Second, observed reductions in LDL-C and PCSK9 were durable up to two years after dosing, with additional follow-up ongoing, consistent with a potential permanent treatment effect resulting from DNA modification.¹⁰⁹ Third, transient acute laboratory safety events were observed—including grade 3 drug-induced events of ALT increase and thrombocytopenia in one patient. The laboratory abnormalities were suspected to be due to the LNP delivery system and prompted a pause in Heart-1 study enrolment. Serious cardiovascular adverse events occurred in two patients, including a fatal cardiac arrest in one patient (0.3 mg/kg) ∼5 weeks after infusion that was determined to be unrelated to study treatment by the investigators and independent Data and Safety Monitoring Board (DSMB) and a myocardial infarction after infusion of a dose of 0.45 mg/kg that was considered potentially related given proximity to dosing. The DSMB concluded that the events were consistent with the severe, advanced ASCVD patient population enrolled and recommended continuing dosing.

However, Verve is currently prioritising development of VERVE-102, a second investigational medicine also designed to inactivate PCSK9 using the same ABE and gRNA that demonstrated efficacy in studies of VERVE-101, but with a different proprietary LNP delivery system that includes two notable differences [Graphical Abstract]. First, the LNP formulation for VERVE-102 employs a different ionisable lipid component that has been well-tolerated to date in third-party clinical trials.¹¹⁰ Second, N-acetylgalactosamine (GalNAc) has been incorporated into the LNP delivery system. GalNAc enables uptake through the asialoglycoprotein receptor (ASGPR) that is abundantly expressed on hepatocytes and provides an alternative delivery path to standard LDLR-mediated LNP endocytosis. VERVE-102 is being studied in the ongoing Heart-2 Phase 1b trial in patients with HeFH or a history of premature coronary artery disease who are unable to achieve adequate LDL-C lowering with currently available oral lipid-lowering therapies such as statins or ezetimibe (NCT06164730). Dosing of the first patient in Heart-2 was announced in May 2024. Initial data from 14 participants showed that VERVE-102 was well-tolerated with no clinically significant laboratory abnormalities or cardiovascular events. Dose-dependent reductions in blood PCSK9 and LDL-C were observed, with a mean LDL-C reduction of 53% in the 0.6 mg/kg dosing group. Dose escalation in the trial is ongoing.¹⁰⁹

VERVE-201 is a third investigational in vivo base editing medicine that uses the same GalNAc LNP delivery system as VERVE-102, but has a guide RNA designed to inactivate the ANGPTL3 gene instead of PCSK9.¹¹¹ Non-clinical data reported to date have provided evidence for potent and highly specific on-target editing of the ANGPTL3 gene in primary human hepatocytes. Species-specific surrogates of VERVE-201 have achieved mean reductions in circulating ANGPTL3 of ∼90% in both murine and non-human primate models, even within the context of severe LDLR deficiency. These surrogates have been well-tolerated in non-clinical studies to date, with no detectable evidence of long-term liver toxicity or increase in liver steatosis. In Ldlr^−/− mice on a Western diet, a 97% reduction in blood ANGPTL3 concentration was observed, leading to a mean 47% reduction in LDL-C and 72% reduction in triglyceride concentrations. Similarly, in a non-human primate model of severe LDLR deficiency, mean LDL-C decreased from 11.8 to 6.4 mmol/L (458 to 247 mg/dL), corresponding to a 46% decrease from baseline. These observations of a 40% to 50% reduction in LDL-C concentrations in models of LDLR deficiency are well-aligned with the results of clinical trials of the evinacumab and zodasiran ANGPTL3 targeted therapies in patients with HoFH described above.^58,86 VERVE-201 is being evaluated in a recently announced Phase 1b study of high-risk patients with hypercholesterolaemia refractory to approved standard of care therapies (NCT06451770).

Multiple additional companies are developing products, including CRISPR-based therapies and other gene editing techniques, with multiple candidates in the pipeline having recently entered clinical trials. In 2023, CRISPR Therapeutics began two phase 1 trials targeting blood lipids. The first one, CTX310, targets the ANGPTL3 gene, whereas the second, CTX320, targets the LPA gene. These therapies also use in vivo LNP delivery to target the liver but use a Cas9 nuclease to disrupt the target genes by imparting a double-stranded break in the DNA rather than editing a specific nucleotide base using a base editor. The phase 1 trials were based on preclinical studies that were presented at the 2023 American Heart Association Scientific Sessions.^112,113 A single dose of CTX310 in non-human primates resulted in 70% mean editing of ANGPTL3 in the liver, an over 85% reduction in plasma ANGPTL3 protein, and a 60% reduction in TG, with effects lasting beyond a year and only transient liver enzyme elevation.¹¹² Similarly, CTX320 led to a 95% reduction in plasma Lp(a) levels in NHPs, with durable effects and a well-tolerated safety profile, suggesting potential use in reducing plasma Lp(a) levels in humans.¹¹³ The advancements in these technologies and their translation from bench to bedside hold the promise of transforming the treatment landscape for cardiovascular and metabolic diseases.

Challenges and limitations with gene editing therapy

Gene editing therapies for lipoprotein disorders face several limitations which might affect their widespread adoption. One concern is the potential for unpredictable off-target effects, which may cause unintended genetic modifications and lead to unpredictable adverse outcomes. Off-target mutagenesis generates de novo mutations at undesired genetic loci, including in the DNA and RNA level.⁹⁶ However, a comprehensive set of non-clinical studies of VERVE-201 that have assessed potential off-target editing have indicated a high degree of specificity for ANGPTL3 editing, and a low risk of off-target genomic modifications that could be expected to have an associated clinical adverse effect.¹¹¹ Additionally, the permanent nature of DNA editing raises safety concerns, particularly if targeting genes with pleiotropic effects or those that lack pharmacologic or human genetic validation. Furthermore, delivery systems, such as viral vectors may not efficiently reach target tissues or may provoke immune responses. While nanoparticles are a potential effective delivery option, they have limited biodistribution, with tendency to accumulate primarily in the liver, which makes delivery to extrahepatic potential organs challenging. Variability in individual genetic and epigenetic landscapes can influence the efficacy and durability of therapeutic effects. Finally, currently approved gene therapies are extremely costly, which has limited global adoption. These challenges highlight the need for ongoing optimization and clinical studies, to ensure the safety and applicability of gene editing in clinical practice.^96,97

Epigenetic silencing for lipoprotein disorders

Programmable epigenetic silencing also known as ‘Hit-and-Run’ approach is another emerging technology in the field of genomic medicine therapeutics. Epigenome editing using CRISPR off demonstrates durable knockdown of PCSK9 expression via targeted promoter methylation, offering potential for long-term cardiovascular risk reduction. However, transient effects observed in ANGPTL3 and AGT knockdowns highlight variability in methylation durability across target genes.¹¹⁴ More recently, the use of zinc-finger protein-based editors for epigenetic silencing showed a potential as a durable gene therapy for lipoprotein disorders. In mice, a single lipid nanoparticle delivery of editor mRNAs targeting PCSK9 achieved nearly 50% reduction in circulating PCSK9 levels for a year, with silencing maintained after liver regeneration. This approach promises high specificity and efficacy without DNA breaks, paving the way for potential human studies based on epigenetic modulation.¹¹⁵

Conclusion

Advancements in genetic therapies, particularly AAV-mediated gene addition therapy and CRISPR-based editing, offer transformative potential in the management of cardiovascular and metabolic diseases. These novel approaches hold promise for addressing the genetic bases of lipoprotein disorders, providing more effective and durable treatments compared with traditional therapies. The integration of gene editing into clinical practice could significantly improve outcomes for patients with monogenic disorders such as HoFH, HeFH, elevated Lp(a), and FCS as well as those with polygenic hyperlipidaemia. Additionally, the utilisation of ML for risk prediction and the development of innovative agents like VERVE-102 and VERVE-201 further enhance the prospects for personalized and precise therapeutic strategies. However, especially in the case of strategies dependent on viral vectors for delivery, challenges such as immune responses, precise vector targeting, and the high cost of therapy need to be addressed. There is a growing understanding of the necessity of the involvement of advocacy groups in assessing patients’ preferences and values concerning the new genetic therapies. Continued research and collaboration across disciplines concomitantly with involving patients’ representation in every step of the process, are essential to overcome these barriers and fully realize the potential of gene editing in revolutionizing cardiovascular and metabolic disease management.

Supplementary data

Supplementary data are available at European Heart Journal online.

Declarations

Disclosure of Interest

C.G. received consultant fees from Sanofi, and non-promotional honoraria from Novartis, Madison, Sanofi, Boehringer Ingelheim and AstraZeneca. A.B. receives research funding from Alexion, Amgen, Arrowhead, Eli Lilly, Ionis, Kaneka Medical, NewAmsterdam Pharma, Novartis, and Regeneron; and honoraria from Ionis, Kaneka Medical, Medscape, Novartis, and Regeneron. A.V.K. is an employee of Verve Therapeutics; has served as a scientific advisor to Marea Therapeutics and Foresite Labs; and holds equity in Verve Therapeutics, Marea Therapeutics, Color Health. R.D. is supported by the National Institute of General Medical Sciences of the NIH (R35-GM124836), consultant and equity holder for Pensieve Health and consultant for Variant Bio. H.S. received honoraria from Amarin, Amgen, Astra-Zeneca, Bayer Vital GmbH, Boehtinger Inelheim, Bristol-Myers Swuibb, Daiihi Sankyo, Medtronic, MSD Sharp & Dohme, Novartis, Pharmacosmos, Sanofi Aventis, Servier, Synlab. K.M. is an advisor to and holds equity in Verve Therapeutics and Variant Bio, is an advisor to LEXEO Therapeutics and Capstan Therapeutics, and receives research funding from Nava Therapeutics and Beam Therapeutics. R.S.R. reports research funding to his institution from Amgen, Arrowhead, Eli Lilly, Merck, NIH, Novartis, Novo Nordisk, and 89Bio, consulting fees from Amgen, Arrowhead, CRISPER Therapeutics, Editas Medicine, Eli Lilly, Intercept Pharmaceuticals, Life Extension, Lipigon, New Amsterdam, Novartis, Regeneron, Rona Therapeutics, and Verve Therapeutics, non-promotional honoraria from Kowa American Corporation, TD Cowen and Viatris, royalties from Wolters Kluwer (UpToDate), and stock holding in MediMergent, LLC. He reports patent applications on: Methods and systems for biocellular marker detection and diagnosis using a microfluidic profiling device. EFS ID: 32278349. Application No. (PCT/US2019/026364) (provisional); Compositions and methods relating to the identification and treatment of immunothrombotic conditions. New International Application No. (PCT/US2021/63104926).

Data Availability

No data were generated or analysed for or in support of this paper.

Funding

All authors declare no funding for this contribution.