This editorial refers to ‘The chromosome 9p21 risk locus is associated with angiographic severity and progression of coronary artery disease’†, by R.S. Patel et al., on page 3017
Coronary artery disease (CAD) is well known to be, to some degree, heritable. A family history of premature CAD is an established risk factor, and studies in monozygotic twins indicate that this effect is genetic rather than from a shared environment.1 These well established truths have been the driving force behind genetic studies of patients with CAD. An initial wave of studies ∼15 years ago examining informative mutations in candidate genes produced a large collection of underpowered studies that were characterized by lack of reproducibility when tested in larger cohorts, with the exception of variants in the apo ε gene.2 The next wave of genetic studies consisted of family-based, linkage studies usually of prematurely affected individuals. These repeated the experience of lack of reproducibility between studies and introduced a new problem, that of identified areas of the genome thought to be causative having no identifiable gene to which a function could be ascribed. Linkage studies did, however, indicate that the task of identifying genetic markers/causes of CAD was going to be tough, with many genes having small additive or interactive effects. The lack of power in the linkage studies, and their vulnerability to disease misclassification, prompted a return to association-based structures but now with herculean numbers of patients and unbiased screening of the whole genome using increasingly dense maps of identified genetic variants [single nucleotide polymorphisms (SNPs)] scattered across the human genome. These have led to the genome-wide association studies (GWAS), which have led to new and potentially very exciting data.
Why so much excitement? First, there has now been replication between studies and, secondly, a handful of markers associated with CAD have eye-wateringly large P-values of significance.3 An example of this is the association of a marker (rs10757278 ) on the short arm of chromosome 9 at position 21.3—so-called Ch9p21.3—with CAD or myocardial infarction.4–7 The excitement, perhaps driven in part by relief that effort has at last been rewarded, has again come with the problem that there is not an obvious gene at the site of the peak of SNP association; therefore, no function can be assigned to this. Carriage of the disease-associated SNP is common (∼50%) in the population and, as a result, the odds ratio for an individual is low, but of course at a population level it is important. In other words, this is going to be more important as a new way of understanding pathogenic mechanisms of CAD rather than as a risk test for an one individual.
Scanning the area of the genome adjacent to the peak on Ch9p21.3 shows two genes involved with cell proliferation (CDKN2a and CDKN2b), cyclin-dependent kinases involved in cell cycle regulation, and an overlapping non-coding RNA, ANRIL, which may interfere with the adjacent CDKN2 genes.8 The finding that deletions in this region are associated with some cancers, and that deletion of the orthologous region in mice produces smooth muscle cells that proliferate at a higher level than normal,9 drives an idea that carriage of this allele is pro-proliferative and, therefore, accelerates atheroma development in susceptible individuals (see Figure 1).
In this edition of the journal, Patel et al. suggest that carriage of the risk allele is associated with a greater burden of atherosclerosis on coronary angiography, and in a small subset of these patients (13%) who had two angiograms for clinical reasons over a mean period of 4.5 years there was an association with progression of disease.10 These data are presented as supporting the idea that the Ch9p21.3 allele is causally linked to exaggerated atheroma development because of increased proliferation.
Patel et al. have shown in their cohort of 2334 angiograms that carriage of the SNP rs10757278 is associated with coronary burden of atheroma. They have used two different angiographic scores as well as a categoric single, two vessel, and three vessel classification of the population, and all are associated with the risk SNP and the effect is additive—the effect is greater in homozygotes than heterozygotes. The angiographic assessment scores used were different from the Duke score previously used by Anderson et al.11 which had not shown an association with extent of coronary disease. There was a just significant association with a history of prior myocardial infarction. In order to examine the proliferative potential of carriage of this SNP, progression of atheroma was examined in the subgroup who had two angiograms for analysis. In this group, carriage of the disease-associated SNP predicted progression of the atheroma as assessed by an increase in angiogram score.
To interpret these data, and to feel comfortable with the conclusions, it is necessary to examine the significance of the burden of coronary atherosclerosis, what angiographic progression means in a small subgroup, and to be aware of the potential biology of this genetic variant.
An important consideration is that any population undergoing coronary angiography represents patients who have presented to the medical profession, in the main because of chest pain. Within this population, the extent of disease may be linked to outcomes,12 but the link is weak and overwhelmed by the effect of left ventricular function,13,14 which has significant prognostic implications. In the general population, histopathological data suggest an imprecise relationship between burden of coronary atheroma and myocardial infarction15—a fact realized by many cardiologists who, when looking after young patients with myocardial infarction, will be aware that the plaque presenting may be very minor in angiographic terms. It may well be that the extent of atheroma is a marker of age, which is the major risk factor for presentation of disease, and/or left ventricular function. So we know very little of what it means to have an angiographic score that is high or low in biological terms and, as a result, the relevance of the association with an SNP is difficult to assess in biological terms.
Progression between angiograms has been much studied in the past, and conclusions have been difficult. Progression is probably driven by traditional risk factors,16 and unstable lesions progress more than stable lesions.17 What might allow for angiographic progression? An angiographic score can progress because either pre-existing lesions become more severe or there are new lesions. Biologically this may be driven by smooth muscle cell division, thrombus accumulation, and plaque inflammation—all of which are interlinked. Carriage of a risk SNP at 9p21 by virtue of data we have on the juxtaposed cdk genes, and the experimental evidence from mice, suggest that the first of these may be the pathogenic link, but one has to be aware that these are somatic mutations and thus affect all cells in the plaque. Furthermore, CDKs have functions in inflammatory cells. So, it may be easy to overinterpret this observation.
What do the progression data tell us about the patients? First this is a very small subset of the whole. Secondly, progressors and non-progressors have, at least within the group, all presented in such a way as to have a second angiogram, i.e. they have survived to get to the medical profession who feel that the situation warrants re-investigation rather than continued therapy. If we assume all of these were driven by new incident cases of chest pain (i.e. their clinical phenotype is homogeneous) then some have no obvious angiographic reason why their chest pain is worse, a fact well known from previous studies.17 These patients have a lower carriage rate of the risk SNP. Those with evidence of progression of disease have a higher carriage rate. The possible explanations are numerous—increased smooth muscle cell proliferation as suggested, impairment of collateral development in those non-progressors not carrying the disease SNP, increased inflammation driving new plaque instability, etc. It is also worth remembering that some have speculated that increased smooth muscle cell proliferation resulting in plaque expansion stabilizes the plaque, whereas dysregulated proliferation culminating in apoptosis (cell death) causes plaque instability. Which of these possibilities would be identified in an angiographic subset is difficult to imagine. An alternative interpretation of this patient group is that it is heterogeneous in terms of the reason why they have had a second angiogram, in which case there are multiple phenotypes and the interpretation is even less easy.
What do we know? Certainly the carriage of the risk allele at Ch9p21 is strongly associated with CAD—it is just too consistent across all studies and these data are another piece of fuel on this fire. The region has effects on vascular cells, but little is known of the function in other compartments of the atherogenic apparatus. The story will be unpicked by detailed cell biology of these components and the examination of highly characterized patient groups with tightly described phenotypes. In this way, contemporary genetic studies of CAD will tell us about mechanisms. It is a long way off from prognostic testing in individuals.
Conflict of interest: D.C.C. is supported by the National Institute for Health Research Cardiovascular Biomedical Research award to Sheffield Teaching Hospitals NHS Foundation Trust. He is Director of this Unit.