Supplementary Materials for

Transcription

1 Supplementary Materials for Genetic identification of familial hypercholesterolemia within a single U.S. health care system Noura S. Abul-Husn, Kandamurugu Manickam, Laney K. Jones, Eric A. Wright, Dustin N. Hartzel, Claudia Gonzaga-Jauregui, Colm O Dushlaine, Joseph B. Leader, H. Lester Kirchner, D Andra M. Lindbuchler, Marci L. Barr, Monica A. Giovanni, Marylyn D. Ritchie, John D. Overton, Jeffrey G. Reid, Raghu P. R. Metpally, Amr H. Wardeh, Ingrid B. Borecki, George D. Yancopoulos, Aris Baras, Alan R. Shuldiner, Omri Gottesman, David H. Ledbetter, David J. Carey, Frederick E. Dewey, Michael F. Murray* This PDF file includes: Fig. S1 Tables S1 to S8 *Corresponding author. mfmurray1@geisinger.edu Published 23 December 2016, Science 354, aaf7000 (2016) DOI: /science.aaf7000

2 Table S1. Cohort demographics, clinical characteristics, and FH variant status of sequenced adult ( 18 years) MyCode participants. All sequenced participants FH variant negative FH variant positive all 41 variants* N 50,726 50, Age, median (IQR) 61 (48-73) 61 (47-72) 61 (47-72) 61 (47-72) Female, N (%) 30,028 (59.2%) 29,864 (59.2%) 164 (56.0%) 134 (58.5%) Race, N (%) Ethnicity, N (%) Asian 129 (0.3%) 129 (0.3%) Black / African American 547 (1.1%) 530 (1.1%) 17 (5.8%) 7 (3.1%) Native American / Alaskan Native 51 (0.1%) 51 (0.1%) Native Hawaiian / Other Pacific Islander 41 (0.1%) 41 (0.1%) Other 3 (0.01%) 3 (0.01%) Unknown 63 (0.1%) 63 (0.1%) White 49,892 (98.4%) 49,616 (98.4%) 276 (94.2%) 222 (96.9%) Hispanic / Latino 549 (1.1%) 544 (1.1%) 5 (1.7%) 2 (0.9%) Not Hispanic / Latino 48,477 (95.6%) 48,201 (95.6%) 276 (94.2%) 220 (96.1%) Unknown 1,700 (3.4%) 1,688 (3.3%) 12 (4.1%) 7 (3.1%) EHR-documented statin prescription (ever), N (%) 27,402 (54.0%) 27,185 (53.9%) 217 (74.1%) 179 (78.2%) LDL-C data available in EHR, N (%) 42,696 (84.2%) 42,442 (84.2%) 256 (87.3%) 204 (89.1%) Maximum EHR-documented LDL-C (mg/dl)***, N (%) LDL-C < ,512 (66.8%) 28,444 (67.0%) 70 (27.3%) 48 (23.5%) 155 LDL-C < 189 9,749 (22.8%) 9,689 (22.8%) 60 (23.4%) 44 (21.6%) 190 LDL-C < 249 4,045 (9.5%) 3,973 (9.4%) 72 (28.1%) 62 (30.4%) 250 LDL-C < (0.8%) 300 (0.7%) 37 (14.5%) 33 (16.2%) LDL-C (0.1%) 36 (0.1%) 17 (6.6%) 17 (8.3%) Severe hypercholesterolemia (LDL-C 190 mg/dl)***, N (%) 4,435 (10.4%) 4,309 (10.2%) 126 (49.2%) 112 (54.9%) EHR evidence of possible clinical FH diagnosis, N (%) ICD10 code E78.0 (Pure Hypercholesterolemia) Encounter at Lipid Clinic ICD10 code E78.0 or encounter at Lipid Clinic 1,002 (2.0%) 2,387 (4.7%) 2,632 (5.2%) 984 (2.0%) 2,352 (4.7%) 2,593 (5.1%) 18 (6.1%) 35 (11.9%) 39 (13.3%) FH variant positive strict 35 variants** 17 (7.4%) 31 (13.5%) 35 (15.3%) EHR-documented family history of high cholesterol, N (%) 1,280 (2.5%) 1,260 (2.5%) 20 (6.8%) 19 (8.3%) Abbreviations: FH, familial hypercholesterolemia; LDL-C, low-density lipoprotein cholesterol. * All 41 variants refers to the complete set of known (i.e. ClinVar documented) and predicted (i.e. protein-truncating loss of function) pathogenic FH variants identified in the Page 2 of 12

3 cohort. ** Strict 35 variants refers to the final curated set of known and predicted pathogenic FH variants. Following manual review, we removed two variants with conflicting evidence in ClinVar, two variants that were located at the same amino acid residue as a pathogenic variant but were not in ClinVar themselves, and two variants that were annotated as splice variants in an alternative transcript of LDLR (see Table S2). *** Percentages are based on number of individuals with LDL-C data available in EHR. Page 3 of 12

4 Table S2. FH variants identified in 50,726 individuals, number of carriers, associated maximum LDL-C levels, and estimated clinical penetrance based on the Dutch Lipid Clinic Network criteria. The final list of 35 pathogenic and predicted pathogenic variants excludes those shaded in gray. # Het Carriers with LDL-C data (Total #) Maximum LDL-C (mg/dl)*** Penetrance **** N (%) CHR:POS:REF:ALT* Gene Function cdna position** Protein position Loss of Function Significance (ClinVar) 1: :G:A PCSK9 Missense c.644g>a p.arg215his Pathogenic 1 (1) (100%) 1: :G:A PCSK9 Missense c.1120g>a p.asp374asn Pathogenic 2 (3) 132 ( ) 1: :C:T PCSK9 Missense c.1405c>t p.arg469trp Pathogenic 6 (7) 154 ( ) 1: :C:T PCSK9 Missense c.1486c>t p.arg496trp Pathogenic 15 (18) 172 ( ) 6 (40%) 2: :G:A APOB Missense c.10672c>t p.arg3558cys Pathogenic 39 (46) 154 ( ) 13 (33%) 2: :C:T APOB Missense c.10580g>a p.arg3527gln Pathogenic / 53 (56) 193 ( ) 33 (62%) Likely Pathogenic 2: :G:A APOB Missense c.10579c>t p.arg3527trp ***** 14 (15) 170 ( ) 4 (29%) 19: :G:A LDLR Stop gained c.131g>a p.trp44* Yes 1 (2) (100%) 19: :GT:G LDLR Frameshift c.187delt p.cys63fs Yes 1 (1) (100%) 19: :CG:C LDLR Frameshift c.233delg p.arg78fs Yes 1 (1) : :T:G LDLR Missense c.259t>g p.trp87gly Pathogenic 2 (3) 198 ( ) 1 (50%) 19: :G:A LDLR Splice donor c.313+1g>a Yes 5 (6) 293 ( ) 4 (80%) 19: :G:C LDLR Splice donor c.313+1g>c Yes 1 (1) : :T:C LDLR Splice donor c.313+2t>c Yes 9 (9) 234 ( ) 7 (78%) 19: :C:A LDLR Stop gained c.501c>a p.cys167* Yes 1 (1) (100%) 19: :GAC:G LDLR Frameshift c.680_681delac p.asp227fs Yes 2 (2) 287 ( ) 2 (100%) 19: :C:G LDLR Missense c.681c>g p.asp227glu Pathogenic 1 (1) (100%) 19: :G:A LDLR Missense c.682g>a p.glu228lys Pathogenic 1 (1) 90 19: :G:T LDLR Stop gained c.682g>t p.glu228* Yes 3 (3) 237 ( ) 2 (67%) 19: :G:A LDLR Missense c.718g>a p.glu240lys Pathogenic 1 (1) : :T:A LDLR Missense c.798t>a p.asp266glu Conflicting 23 (30) 176 ( ) 9 (39%) 19: :CAA:C LDLR Frameshift c.881_882delaa p.lys294fs Yes 1 (1) (100%) 19: :G:A LDLR Missense c.1003g>a p.gly335ser Pathogenic 4 (4) 164 ( ) 1 (25%) 19: :G:A LDLR Missense c.1024g>a p.asp342asn Conflicting 8 (11) 98 (96-140) 1 (13%) 19: :ATGCG AAGG:A LDLR Frameshift c.1056_1060+3delc GAAGGTG p.cys352fs Yes 1 (1) (100%) 19: :G:A LDLR Missense c.1217g>a p.arg406gln ***** 1 (1) (100%) 19: :T:A LDLR Splice donor c t>a Yes 3 (4) 250 ( ) 2 (67%) 19: :G:A LDLR Splice acceptor c g>a Yes 2 (3) 322 ( ) 2 (100%) Page 4 of 12

5 19: :G:T LDLR Missense c.1414g>t p.asp472tyr Pathogenic 3 (3) 183 ( ) 3 (100%) 19: :G:GC LDLR Frameshift c.1428dupc p.asp477fs Yes 2 (2) 251 ( ) 2 (100%) 19: :G:A LDLR Splice donor c g>a Yes 0 (1) -- 19: :G:A LDLR Missense c.1775g>a p.gly592glu Pathogenic / Likely Pathogenic 12 (12) 227 ( ) 10 (83%) 19: :C:A LDLR Stop gained c.2043c>a p.cys681* Yes 1 (1) (100%) 19: :C:T LDLR Missense c.2054c>t p.pro685leu Pathogenic 0 (1) -- 19: :T:TC LDLR Frameshift c.2061dupc p.asn688fs Yes 1 (1) (100%) 19: :G:A LDLR Missense c.2098g>a p.asp700asn Pathogenic 1 (1) : :C:CT LDLR Frameshift c.2259dupt p.gly754fs Yes 1 (1) (100%) 19: :T:TG LDLR Frameshift c.2416dupg p.val806fs Yes 1 (1) (100%) 19: :A:G LDLR Splice acceptor c a>g****** Yes 5 (6) 137 (67-141) 19: :G:T LDLR Splice acceptor c g>t****** Yes 1 (1) 74 dup_ex13_17 LDLR Internal duplication 26 (29) 257 ( ) 25 (96%) Abbreviations: CHR: chromosome; POS: chromosomal position based on hg19 build of the human reference genome; REF: reference allele; ALT: alternate allele; Het: heterozygous; LDL-C, low-density lipoprotein cholesterol. *Genomic coordinates based on GRCh37. **PCSK9 transcript ENST (NM_ ); APOB transcript ENST (NM_ ); LDLR transcript ENST (NM_ ). *** Median and interquartile range (IQR) of the maximum EHR-documented LDL-C levels for all carriers of each variant. **** Penetrance defined as number of carriers meeting pre-sequencing criteria for a diagnosis of possible, probable, or definite FH (using DLCN criteria). Percentages are based on number of heterozygous carriers with LDL-C data available in EHR. ***** Variant located at the same amino acid residue as a pathogenic variant in ClinVar but not itself in ClinVar. ****** Splice variant in an alternative transcript ENST (NM_ ) of LDLR. Page 5 of 12

6 Table S3. Associations between FH variants and CAD. Gene All CAD cases Number of FH variant carriers cases Number of FH variant carriers controls Cumulative allele frequency cases (%) Cumulative allele frequency controls (%) Odds ratio* (95% CI) P value* All FH variants 94/12, /35, ( ) 4.3x10-11 LDLR variants - all 52/12,298 39/35, ( ) 8.0x10-11 LDLR variants - plof 43/12,298 23/35, ( ) 7.7x10-13 APOB variants 34/12,298 58/35, ( ) 7.6x10-3 PCSK9 variants 8/12,298 20/35, ( ) 0.15 Premature CAD cases (Males: 55 years old, Females: 65 years old) All FH variants 53/4,150 77/21, ( ) 5.5x10-14 LDLR variants - all 37/4,150 23/21, ( ) 1.2x10-17 LDLR variants - plof 30/4,150 14/21, ( ) 9.8x10-19 APOB variants 13/4,150 42/21, ( ) 0.12 PCSK9 variants 3/4,150 12/21, ( ) 0.04 Abbreviations: CAD, coronary artery disease; CI, confidence interval; FH, familial hypercholesterolemia; plof, predicted loss of function (defined as variants leading to a premature stop codon, or loss of a start or stop codon; disrupting canonical splice acceptor or donor dinucleotides; or frame shifting leading to the formation of a premature stop codon). *Adjusted for age, age 2, sex, and the first five principal components of ancestry. Page 6 of 12

7 Table S4. Cause of death and evidence of FH-related comorbid disease by chart review of 14 deceased individuals with FH variants. Age of death Sex Cause of death Cardiovascular disease-related death Evidence of potential FH-related comorbid disease Ischemic cardiomyopathy Aortic valvular Cerebrovascular Other vascular 84 F CHF Yes Yes Yes No No 82 M CHF Yes Yes No Yes No 83 M CHF Yes Yes No No Yes 72 M CHF Yes Yes No No Yes 69 M CHF, hip fracture Yes Yes No No No 91 F CHF, severe aortic stenosis Yes Unknown Yes Yes Yes 67 F Cardiac arrest Yes No No No No 63 M Unknown Unknown No Yes Yes Yes 76 M Unknown Unknown No No No Yes 82 M Unknown Unknown Yes No No No 78 M Renal failure Unknown Yes Yes No No 58 F Renal failure No No No No No 77 M Gastrointestinal bleed No No No Yes Yes 83 M Gastrointestinal bleed No Yes No No Yes Abbreviations: CHF, congestive heart failure; FH, familial hypercholesterolemia Page 7 of 12

8 Table S5. Availability of DLCN diagnostic criteria for FH in EHRs of 215 living FH variant carriers. Dutch Lipid Clinic Network Criteria Points Meeting criteria in EHR, N (%) Family History First-degree relative with premature coronary and/or vascular disease (men 55 years, women 60 years) First-degree relative with known LDL-C 95th percentile for age and sex First-degree relative with tendon xanthomata and/or arcus cornealis, Children aged 18 years with known LDL-C 95th percentile for age and sex Clinical History Patient with premature coronary artery disease (men 55 years, women 60 years) Not meeting criteria in EHR, N (%) Implementation (67.9%) 69 (32.1%) - Searched for Heart Disease in family history (not limited to firstdegree relatives as this information was not always available) - Searched for Family history of premature heart disease in problem list 1 * 215 (100%) - Searched for Family history of hyperlipidemia in problem list 2 N/A N/A - Data unavailable 2 * 215 (100%) - Used known mother/child links (available since 2010) to search for children with LDL-C 95th percentile 2 44 (20.5%) 171 (79.5%) - Used electronic phenotyping to identify patients with premature coronary artery disease Patient with premature cerebral or 1 8 (3.7%) 207 (96.3%) - Searched for ICD9 diagnosis codes in encounters and problem list peripheral vascular disease (men 55 years, women 60 years) Physical Examination Tendon xanthomata 6 * 215 (100%) - Searched for ICD9 diagnosis codes in encounters and problem list Arcus cornealis at age 45 years 4 * 215 (100%) - Searched for ICD9 diagnosis codes in encounters and problem list LDL-C (mg/dl) None recorded - 23 (10.7%) - - Used maximum EHR-documented outpatient LDL-C level LDL-C < (20.5%) LDL-C < (19.1%) LDL-C < (28.4%) LDL-C < (14.0%) - LDL-C (7.4%) - DNA Analysis functional variant in LDLR, APOB or PCSK9 gene LDLR 8 96 (44.7%) - APOB 8 91 (42.3%) - PCSK (13.0%) - Abbreviations: EHR, electronic health record; FH, familial hypercholesterolemia; LDL-C, low-density lipoprotein cholesterol; N/A, not available. * This reflects the magnitude of missing data related to family history and relevant physical exam findings within the EHR. Page 8 of 12

9 Table S6. Likelihood of FH diagnosis using DLCN criteria without and with genotype information for 215 living FH variant carriers. Living FH variant carriers FH Diagnosis using EHR data alone FH diagnosis using EHR and sequencing data Unlikely (<3) Possible FH (3-5) Probable FH (6-8) Definite FH (>8) Probable FH (6-8) Definite FH (>8) N (%) (44.7%) 68 (31.6%) 35 (16.3%) 16 (7.4%) 27 (12.6%) 188 (87.4%) Maximum EHR-documented LDL-C (mg/dl) LDL-C < 155, N (%) 155 LDL-C < 189, N (%) 190 LDL-C < 249, N (%) 250 LDL-C < 329, N (%) LDL-C 330, N (%) 44 (20.5%) 41 (19.1%) 61 (28.4%) 30 (14.0%) 16 (7.4%) 43 (44.8%) 32 (33.3%) 1 (1.5%) 9 (13.2%) 52 (76.5%) 4 (5.9%) 9 (25.7%) 26 (74.3%) 16 (100%) 17 (63.0%) 27 (14.4%) 41 (21.8%) 61 (32.4%) 30 (16.0%) 16 (8.5%) No EHR-documented LDL-C data, N (%) 23 (10.7%) 21 (21.9%) 2 (2.9%) 10 (37.0%) 13 (6.9%) Clinical characteristics Age, median (IQR) Female, N (%) Body mass index, median (IQR) Ever smoker, N (%) 61 (47-40) 130 (60.5%) 30.3 ( ) 80 (37.2%) 58.5 (44-74) 58 (60.4%) 29.7 ( ) 34 (35.4%) Abbreviations: EHR, electronic health record; FH, familial hypercholesterolemia; LDL-C, low-density lipoprotein cholesterol. 61 (51-67) 71 (59.7%) 31.2 ( ) 46 (38.7%) 61 (47-40) 130 (60.5%) 30.3 ( ) 80 (37.2%) Page 9 of 12

10 Table S7. Likelihood of FH diagnosis using modified MEDPED criteria for 215 living FH variant carriers. No family history of hyperlipidemia Unlikely FH Likely FH Age < 20 and LDL-C 200 mg/dl 2 1 Age and LDL-C 220 mg/dl 7 5 Age and LDL-C 240 mg/dl 20 4 Age 40 and LDL-C 260 mg/dl No EHR-documented LDL data 23 0 Total Family history of hyperlipidemia* Unlikely FH Likely FH Age < 20 and LDL-C 155 mg/dl 0 0 Age and LDL-C 170 mg/dl 0 2 Age and LDL-C 190 mg/dl 2 3 Age 40 and LDL-C 205 mg/dl 3 9 No EHR-documented LDL data 0 0 Total 5 14 Total, N (%) 162 (75.3%) 53 (24.7%) Abbreviations: EHR, electronic health record; FH, familial hypercholesterolemia; LDL-C, low-density lipoprotein cholesterol. * MEDPED criteria rely on knowledge of a family history of FH, which is not captured effectively in the EHR; therefore, we modified these criteria to include any recorded family history of high cholesterol. Note: Excluding individuals with no EHR-documented LDL-C data (N=23), 53/192 (27.6%) carriers would meet criteria for clinical FH diagnosis. Page 10 of 12

11 Table S8. LDL-C measures and lipid-lowering therapy in living FH variant carriers (n = 215) and noncarriers (n = 46,070). Ever on lipid-lowering therapy Any lipid-lowering agent, N (%) Statin, N (%) High-intensity statin*, N (%) High- or moderate-intensity** statin, N(%) Living FH variant carriers (N = 215) 173 (80.5%) 172 (80.0%) 130 (60.5%) 170 (79.1%) Individuals with medication reconciliation from 2015 or 2016 N=189 N=41,594 Currently on lipid-lowering therapy Any lipid-lowering agent, N (%) Statin, N (%) High-intensity statin, N (%) High- or moderate-intensity statin, N(%) 110 (58.2%) 109 (57.7%) 70 (37.0%) 107 (56.6%) Individuals with LDL-C data available in EHR N = 192 N=38,944 Lowest EHR-documented LDL-C (mg/dl) Mean (SD) (48.5) LDL-C < 100 mg/dl, N (%) 99 (51.6%) Individuals with LDL-C data within 12 months of study N = 106 N = 21,515 Most recent EHR-documented LDL-C (mg/dl) Mean (SD) (63.5) LDL-C < 100 mg/dl, N (%) 41 (38.7%) Individuals currently on statin and with LDL-C data within 12 N = 63 N = 10,880 months of study Most recent EHR-documented LDL-C (mg/dl) Mean (SD) (48.3) LDL-C < 100 mg/dl, N (%) 29 (46.0%) Living noncarriers (N=46,070) 26,302 (57.1%) 24,995 (54.3%) 11,611 (25.2%) 23,674 (51.4%) 16,471 (39.6%) 15,698 (37.7%) 5,842 (14.0%) 14,265 (31.0%) 79.6 (30.0) 29,900 (76.8%) 96.1 (35.4) 12,642 (58.8%) 82.5 (30.8) 8358 (76.8%) Abbreviations: EHR, electronic health record; FH, familial hypercholesterolemia; LDL-C, low-density lipoprotein cholesterol; SD, standard deviation. * High-intensity statin refers to high-dose high-intensity statins: simvastatin 80mg, atorvastatin 40 or 80 mg, and rosuvastatin 20 or 40 mg daily. ** Moderate-intensity statin refers to moderate-dose moderate-intensity statins: atorvastatin 10 or 20 mg, rosuvastatin 5 or 10 mg, simvastatin 20 or 40 mg, pravastatin 40 or 80 mg, lovastatin 40 mg, fluvastatin XL 80 mg, and pitavastatin 2 or 4 mg daily; or fluvastatin 40 mg twice daily. Page 11 of 12

12 90, Mod LDL: 125; NA 81, None LDL: 136; NA 79, None LDL: 155; 98 58, High LDL: 142; NA 66, None LDL: 324; , High LDL: 266; , High LDL: 330; , High LDL: 293; , None LDL: 99; NA 47, None LDL: 142; NA 36, None LDL: 133; NA 28, None LDL: 191; NA 41, None LDL: 159; NA Age (years), current staen prescripeon LDL: Maximum EHR-recorded LDL-C; most recent LDL-C (mg/dl) Variant carrier No geneec data Mod: Moderate intensity staen High: High intensity staen None: No current staen NA: LDL-C not available Fig. S1. Reconstructed pedigrees containing two noncarriers and five carriers (left) and three noncarriers and three carriers (right) of the APOB p.arg3527gln variant. There was segregation of high LDL-C levels with carrier status; maximum LDL-C levels in carriers ranged between mg/dl, and in noncarriers between mg/dl. Two of the five noncarriers were currently prescribed a moderate- or high-intensity statin. Three of the carriers were currently prescribed a high-intensity statin, while the other five carriers had no current statin prescription in the EHR. In the three carriers currently prescribed a high-intensity statin, the most recent LDL-C level ranged from 109 to 187 mg/dl. There were no LDL-C levels available within the last 12 months for any of the noncarriers. EHR, electronic health record; LDL-C, low-density lipoprotein cholesterol. Page 12 of 12