The Effects of Rare SERPINA1 Variants on Lung Function and Emphysema in SPIROMICS

Abstract

Rationale: The role of PI (protease inhibitor) type Z heterozygotes and additional rare variant genotypes in the gene encoding alpha-1 antitrypsin, SERPINA1 (serpin peptidase inhibitor, clade A, member 1), in determining chronic obstructive pulmonary disease risk and severity is controversial.Objectives: To comprehensively evaluate the effects of rare SERPINA1 variants on lung function and emphysema phenotypes in subjects with significant tobacco smoke exposure using deep gene resequencing and alpha-1 antitrypsin concentrations.Methods: DNA samples from 1,693 non-Hispanic white individuals, 385 African Americans, and 90 Hispanics with ≥20 pack-years smoking were resequenced for the identification of rare variants (allele frequency < 0.05) in 16.9 kB of SERPINA1.Measurements and Main Results: White PI Z heterozygotes confirmed by sequencing (MZ; n = 74) had lower post-bronchodilator FEV₁ (P = 0.007), FEV₁/FVC (P = 0.003), and greater computed tomography-based emphysema (P = 0.02) compared with 1,411 white individuals without PI Z, S, or additional rare variants denoted as V_R. PI Z-containing compound heterozygotes (ZS/ZV_R; n = 7) had lower FEV₁/FVC (P = 0.02) and forced expiratory flow, midexpiratory phase (P = 0.009). Nineteen white heterozygotes for five non-S/Z coding variants associated with lower alpha-1 antitrypsin had greater computed tomography-based emphysema compared with those without rare variants. In African Americans, a 5' untranslated region insertion (rs568223361) was associated with lower alpha-1 antitrypsin and functional small airway disease (P = 0.007).Conclusions: In this integrative deep sequencing study of SERPINA1 with alpha-1 antitrypsin concentrations in a heavy smoker and chronic obstructive pulmonary disease cohort, we confirmed the effects of PI Z heterozygote and compound heterozygote genotypes. We demonstrate the cumulative effects of multiple SERPINA1 variants on alpha-1 antitrypsin deficiency, lung function, and emphysema, thus significantly increasing the frequency of SERPINA1 variation associated with respiratory disease in at-risk smokers.

Keywords: SERPINA1; alpha-1 antitrypsin; chronic obstructive pulmonary disease; emphysema; rare variant.

Figure 1.

SERPINA1 genetic study flow diagram summarizes the sequential analytical steps for characterizing rare SERPINA1 variants in SPIROMICS. First, burden tests for PIs (protease inhibitors) Z, S, and 21 additional rare exonic coding variants and 1 frameshift insertion, all with a minor allele frequency of less than 0.05, were performed by self-reported ethnic group to evaluate for gene-level associations with the co–primary outcomes (30). Second, individual rare variants were analyzed with regression-based allelic association tests to further evaluate significant associations. Based on the strong effect of PI type Z variant on the co–primary outcomes in white individuals, we performed association testing of this variant conditional on additional less common (PI S) and rare SERPINA1 variants. We compared white individuals without a PI Z, S, or additional rare variant allele (no Z/S/V_R, where V_R designates all non-Z and non-S rare variants), heterozygotes for non–PI Z rare variation (MS/MV_R), PI Z heterozygotes without another rare variant (MZ), compound heterozygotes with non–PI Z rare variation (SS/SV_R/V_RV_R), PI Z–containing compound heterozygotes with PI S or additional rare variants (ZS/ZV_R), and PI Z homozygotes (ZZ) to evaluate the effects of PI Z heterozygotes with the confirmed absence of other rare variants and PI Z in compound heterozygotes with another rare variant. We performed similar analyses for secondary related outcomes. BODE = body mass index, airflow obstruction, dyspnea, and exercise; FEF_25–75 = forced expiratory flow, midexpiratory phase; SGRQ = St. George’s Respiratory Questionnaire; SPIROMICS = Subpopulations and Intermediate Outcomes Measures in Chronic Obstructive Pulmonary Disease Study.

Figure 2.

SERPINA1 variants identified in the promoter and coding exons in the SPIROMICS multiethnic cohort. Missense and frameshift polymorphisms identified through resequencing of a 16.9 kB region of SERPINA1 containing the untranslated promoter and five coding exons of SERPINA1 in the SPIROMICS multiethnic cohort are shown by amino acid position, coding change, and protease inhibitor type where available. Of the 26 exonic coding variants identified with resequencing, three were the common and benign protease inhibitor types M1Ala/Val (Val²³⁷Ala, rs6647), M2/M4 (Arg¹²⁵His, rs709932), and M3 (Glu⁴⁰⁰Asp, rs1303). A newly identified missense variant is underlined. Variants only found in white individuals are in blue text, those only in African Americans in red text (including an ethnic-specific G-nucleotide 5′ untranslated region insertion [underlined] evaluated with functional studies), and those only in Hispanics in green text. SPIROMICS = Subpopulations and Intermediate Outcomes Measures in Chronic Obstructive Pulmonary Disease Study.

Figure 3.

Effects of rare SERPINA1 variant–containing genotypes on lung function and computed tomography scan–based emphysema. Regression-based association testing compared white individuals without PIs (protease inhibitors) Z, S, or other rare variants (No Z/S/V_R); heterozygotes for non–PI Z rare variants (MS [n = 132] and MV_R [n = 53] heterozygotes); PI Z heterozygotes without another rare variant (MZ); PI S and V_R compound heterozygotes (SS, SV_R, and V_RV_R); PI Z–containing compound heterozygotes with PI S or additional rare variants (ZS/ZV_R); and PI Z homozygotes (ZZ). Compound heterozygotes with PI type Z and S or additional rare variants include three PI type ZS, one PI type Z/PI M_6Passau, one PI type Z/PI I, and two PI type Z/Ala³⁰⁸Ser. Compound heterozygotes without PI type Z included two PI type S homozygotes (SS), one PI type S/Ile³¹⁷Thr, one PI type S/PI type F, and one PI type F/Ala³⁰⁸Ser. Bars represent the means. Regression-based analyses adjusted by study site, age, sex, body mass index, and pack-years smoking history for lung function ([A] FEV₁% predicted and [B] FEV₁/FVC ratio). Models also included height for log-transformed computed tomography scan–based emphysema based on percentage of the area in bilateral lungs with density of −950 Hounsfield units (HFU) or less ([C] log % bilateral area ≤ −950 HFU).

Figure 4.

Rare SERPINA1 variant genotypes and alpha-1 antitrypsin levels in non-Hispanic white individuals and effects on lung function and computed tomography scan–based emphysema. Detailed genotypes in white individuals are shown as those without a rare SERPINA1 variant (MM), those with one rare variant (V_R) that is not PI (protease inhibitor) type S or Z (MV_R), those with one PI type S allele and no other variant (MS), PI type Z heterozygotes (MZ), PI type S homozygotes (SS), compound heterozygotes with PI type S except PI type Z (SV_R), PI type S and Z-containing compound heterozygotes (SZ), Z-containing compound heterozygotes with additional rare variants except PI type S (ZV_R), and PI type Z homozygotes (ZZ), as shown. MV_R heterozygotes included those with Lys⁴⁰⁴Arg (n = 1), Pro³⁹³Ser (PI M_wurzburg; n = 3), Ile³⁶⁴Val (n = 1), Ser³⁵⁴Phe (PI S_Munich; n = 2), Pro³⁵⁰Ser (n = 1), Ala³⁰⁸Ser (n = 7), Asp²⁸⁰Val (PI P_Lowell, Q0_Cardiff; n = 1), Arg²⁴⁷Cys (PI F; n = 8), Tyr²¹¹Cys (n = 1), Tyr¹⁸⁴Terfs (PI Q0_{Granite Falls}; n = 1), Gly¹⁷²Trp (PI M2_Obernburg; n = 5), Ser⁶⁹Phe (n = 1), and Arg⁶³Ser (PI I; n = 6). ZV_R compound heterozygotes included one PI Z/PI M_6Passau, one PI Z/PI I, and one PI Z/Ala³⁰⁸Ser. SV_R compound heterozygotes included one PI S/Ile³¹⁷Thr. Regression-based analyses were adjusted by study site, age, sex, body mass index, and pack-years smoking history for lung function ([A] FEV₁% and [B] FEV₁/FVC ratio). Models also included height for (C) log-transformed computed tomography scan–based emphysema based on percentage of the area in bilateral lungs with density of −950 Hounsfield units (HFU) or less (% area ≤ −950 HFU). α1AT = alpha-1 antitrypsin.

Figure 5.

Effects on non-PI (protease inhibitor) type S or Z rare SERPINA1 variants associated with alpha-1 antitrypsin concentrations on computed tomography scan–based emphysema in non-Hispanic white individuals. Regression-based analyses were adjusted by study site, age, sex, pack-years smoking history, body mass index, and height. Bars represent the means, red dots on the right column represents subjects without rare SERPINA1 variants (common, major allele homozygotes), and the dots of multiple colors on the right column represent different heterozygotes for one of five different rare SERPINA1 variants associated with lower alpha-1 antitrypsin concentrations (P = 0.05) or with mean concentrations ≤1.5 mg/ml, excluding PI type S and Z. HFU = Hounsfield units.