Background
The National Emphysema Treatment Trial, a multicenter randomized trial of lung volume reduction surgery (LVRS) versus medical management for emphysema, found that on average, LVRS led to improved functional status, but not increased survival in patients with emphysema and severe chronic airflow obstruction [
1]. However, substantial variability in response to LVRS was observed. Based on pulmonary function testing and emphysema distribution on chest computed tomography (CT), a patient population with a high risk of death was identified [
2]. Among non-high risk patients, baseline exercise capacity and emphysema distribution on chest CT scans were used to define subgroups with greater or lesser chances of improvement post-LVRS. Yet these clinical subgroups did not fully account for the variable response to LVRS among NETT participants.
We hypothesized that genetic differences may explain some of this variability in response to LVRS. To test this hypothesis, we studied participants in the NETT Genetics Ancillary Study. We examined the association between LVRS outcomes and variants in five genes previously shown to be associated with chronic obstructive pulmonary disease (COPD) susceptibility, exercise capacity, or emphysema distribution on chest CT [
3‐
7]: glutathione S-transferase pi (
GSTP1), microsomal epoxide hydrolase (
EPHX1), transforming growth factor beta-1 (
TGFB1), serpin peptidase inhibitor E2 (
SERPINE2) and surfactant, pulmonary-associated protein B (
SFTPB). Though not a "pharmacogenetic" study in the classic sense of the term – since the intervention studied is a surgical procedure and not a pharmacological agent – the present study is the first to examine genetic associations for response to a specific therapy for COPD.
Discussion
In participants from the NETT Genetics Ancillary Study, we tested associations between variants in five candidate genes and four measures of response to LVRS, finding significant associations for SNPs in two genes, GSTP1 and EPHX1. The effects of a SNP upstream from GSTP1 and a coding SNP in EPHX1 were strongest in the clinically defined subgroup of patients with non-upper lobe predominant emphysema and low baseline exercise tolerance. Additional SNPs in these two genes, including a promoter SNP in EPHX1, appeared to have stronger effects in patients with upper lobe predominant emphysema and low baseline exercise tolerance.
Analysis of the NETT data has demonstrated that non-high risk patients in the upper lobe predominant, low baseline exercise capacity subgroup are most likely to benefit from LVRS, with a survival advantage compared to medical therapy [
1]. Based on these results and previous studies of LVRS [
21], LVRS is widely accepted for patients with severe airflow obstruction due to upper lobe predominant emphysema. Our findings in the upper lobe predominant, low exercise capacity subgroup may distinguish a subset of these patients most likely to respond to surgery. However, the role of LVRS for non-upper lobe predominant emphysema is much less clear [
16]. NETT found no survival improvement from LVRS in the non-upper lobe predominant, low baseline exercise capacity subgroup, but did show the potential for symptomatic benefit in these patients [
1]. The genetic associations in this subgroup may possibly identify patients with non-upper lobe predominant emphysema who have the potential to benefit from LVRS. However, the number of patients included in this subgroup was small.
In contrast to traditional pharmacogenetic studies of drugs and their metabolizing enzymes, the potential effect of SNPs in
GSTP1 and
EPHX1, two genes encoding xenobiotic metabolizing enzymes, on the response to LVRS is not obvious. Variants in these genes may influence an individual's response to the inflammation produced by surgery or to the oxidative stress resulting from single lung ventilation during lung resection [
22]. Alternatively, these genetic variants may be identifying patients with different subtypes of emphysema, beyond the subgroups defined by radiographic distribution and baseline exercise capacity. The fact that we could not replicate these associations in patients randomized to medical therapy demonstrates that the effects of these SNPs are not explained by genetic influences on the natural history of emphysema with severe airflow obstruction. The effects of variants in
EPHX1 may be at least partially mediated through effects on the post-operative course, including complications, evidenced by the change in effect estimates in the analyses excluding patients with post-LVRS hospital stays greater than thirty days. It is unlikely that the associated SNPs are exerting their effects through comorbid illnesses, since the number of major comorbidities in NETT subjects was low due to the study exclusion criteria [
23].
One must also consider the potential effects of the specific SNPs that we have determined to be significantly associated with LVRS outcome. A coding variant in
GSTP1 (Ile105Val) has been associated with COPD and related traits in several studies [
24,
25], but the results have not been consistently replicated [
5,
26]. The SNP with the strongest association in our study, rs612020, is located upstream from the transcription start site of the
GSTP1 gene. The functional effect of this particular SNP is not clear, yet it is in complete LD (in European-Americans from the HapMap project) with another upstream SNP, rs7927381 (which was not genotyped in our study), which may alter a putative CCAAT/enhancer-binding protein (CEBP) site. The transcription factor CEBP-γ may be an important regulator of
GSTP1 expression in human bronchial epithelial cells [
27].
In
EPHX1, rs3753658 is in the promoter region, 290 bp upstream from the transcription start site. The SNP is in complete LD with another promoter SNP (rs3753660, not genotyped in our study) [
28], which may affect a binding site for peroxisome proliferator-activated receptor-γ, a modulator of airway inflammation in COPD [
29]. SNP rs2234922 is located in exon 4 and leads to an amino acid change (His139Arg). Enzymes carrying this variation may have increased activity [
30]; this variant has been termed the "fast" allele. Several studies have reported association between another coding variant (Tyr113His, "slow" allele) and COPD [
31,
32]. As with
GSTP1, this finding has not been consistently replicated. We have previously reported a protective effect of the His139Arg variant on COPD risk, comparing patients from NETT with control subjects [
5]; however, this association was not found in a family-based study of COPD.
The published studies of
GSTP1 and
EPHX1 above have largely examined associations with COPD susceptibility. The present study is the first association analysis examining genetic influences on the response to a specific therapy for COPD or emphysema. In a study of outcomes from thoracic surgery, Shaw and colleagues genotyped six polymorphisms in five genes, finding associations for SNPs in tumor necrosis factor (TNF) and interleukin-6 (IL6) with the risk of complications in 155 patients undergoing lung resections for cancer [
33]. On average, their patients had relatively preserved baseline pulmonary function. In addition, multiple studies have examined genetic and genomic factors influencing outcomes from cardiac surgery [
34].
Our study has several limitations. In NETT, DNA samples were collected at various times following enrollment, and not prior to randomization. Because subjects were recruited into the NETT Genetics Ancillary Study after enrollment into NETT, we could not examine whether genetic variants influenced survival post-LVRS, since patients who died soon after enrollment (e.g. peri-operative deaths) would not be included in the study.
In our analyses of four phenotypes and five genes, including multiple SNPs in those genes, it is possible that the positive results represent spurious associations due to the multiple tests performed. Using a more stringent p-value of 0.01, only three of the genotype-phenotype associations in our study (two SNPs in
GSTP1 and one in
EPHX1) remained significant. In the complex trait genetics literature, there is no clear consensus regarding the optimal statistical methodology to control for multiple testing [
35]. Increasingly, replication of the findings in an independent population has emerged as the standard for confirming a true genetic association [
36]. A limitation of our study is the lack of a suitable replication population. Other clinical trials of LVRS [
21] would likely be underpowered for an adequate replication study, even if DNA were collected on all subjects in these studies. For example, in the combined analysis of the Canadian Lung Volume Reduction Study and the Overholt-Blue Cross Emphysema Surgery Trial, one of the largest LVRS trials outside of NETT, only 58 patients were randomized to surgery [
37]. For a replication study, ideally one targets a sample size at least as large as in the original study [
38].
Acknowledgements
The authors thank Barbara Klanderman, Jody Sylvia, Ankur Patel, Lisa Catalano, and Dawn Ciulla for their assistance with genotyping and sample management. Co-investigators in the NETT Genetics Ancillary Study include Joshua Benditt, Gerard Criner, Malcolm DeCamp, Philip Diaz, Mark Ginsburg, Larry Kaiser, Marcia Katz, Mark Krasna, Neil MacIntyre, Barry Make, Rob McKenna, Fernando Martinez, Zab Mosenifar, Andrew Ries, Paul Scanlon, Frank Sciurba, and James Utz.
This work was supported by National Institutes of Health grants HL080242, HL71393, HL075478, U01HL065899, P01HL083069, a grant from the Alpha-1 Foundation, and an American Lung Association Career Investigator Award. The National Emphysema Treatment Trial was supported by contracts with the National Heart, Lung, and Blood Institute (N01HR76101-N01HR76116, N01HR76118, N01HR76119), the Centers for Medicare and Medicaid Services, and the Agency for Healthcare Research and Quality.
The study sponsors of the NETT Genetics Ancillary Study had no role in study design, data collection, analysis and interpretation, manuscript preparation and submission for publication.
Competing interests
Dr. Silverman has received honoraria, consultant fees, and research grants from GlaxoSmithKline for COPD genetics studies and honoraria from Wyeth, Bayer, and Astra-Zeneca for lectures on COPD genetics. None of the other authors report any relevant competing interests.
Authors' contributions
CPH designed the analysis, participated in data collection, performed the analyses and interpretation of results, and drafted the manuscript. DLD participated in the conceptualization of the analysis, data collection, and revision of the manuscript. JJR participated in the conceptualization of the analysis, subject recruitment, and revision of the manuscript. EKS participated in the design of the analysis, subject recruitment, data collection, interpretation of the results, and revision of the manuscript. All authors have read and approved the final manuscript.