GWAS have been conducted in hundreds of thousands of research participants and are currently underway in many other populations, with increasingly large sample sizes being employed and combined to maximize statistical power. We sought to identify and characterize the potentially “notifiable” genetic variants residing on commercially available SNP arrays used for GWAS. Since the current guidelines for the use of genetic tests from research findings that we adopted indicate these tests should be conducted in CLIA-certified facilities, we considered known genes tested by these laboratories6
. From our review of all types of genetic tests currently measured for 1,362 genes, it appears that only 11.7% of the genes are tested for targeted mutation analysis in ≥2 CLIA-certified laboratories. Using a variety of bioinformatics methods, we confirmed only 12 SNPs tested in ≥2 CLIA-certified laboratories for 9 diseases/genes are currently found on ≥1SNP array used for GWAS. Because GWAS studies increasingly impute millions of SNPs from genotyped SNPs using HapMap information11
, it might be possible that many additional potentially notifiable SNPs could be found among the imputed SNPs. However, we confirmed only 4 additional SNPs found on ≥1 commercially available SNP scan that are “perfect proxies” for the 12 potentially notifiable SNPs based on the HapMap. Further, we found 6 additional HapMap SNPs tested for 6 diseases/genes in ≥2 CLIA-certified laboratories that are not currently found on any SNP arrays. Thus, our findings suggest that SNP arrays used for GWAS, and the HapMap itself, generally harbor very few potentially notifiable genetic variants.
Potential Notifiable SNPs
For a genetic result to be considered notifiable to research participants from community-based populations, a number of criteria have been suggested, including evidence that the notifiable disease has important health implications, that penetrance is relatively high though not necessarily complete (RR>2.0), the risk for disease is strong, the magnitude of risk conferred by the genetic variant is significant, and there are proven therapeutic or preventive interventions available, or there are significant reproductive implications6
. When we assessed the state of the literature for each of the genes (and the specific associated variants available on the commercially-available SNP arrays) to determine whether they met the criteria and might qualify for notification, we found that few, if any, potentially notifiable variants that reside on arrays used in GWAS meet these criteria. Three disorders – congenital adrenal hyperplasia due to 21-hydroxylase deficiency (CYP21A2
), biotinidase insufficiency (BTD
) and galactosemia (GALT
) – primarily display onset in newborns or infants and are identified with high sensitivity by newborn screening (National Newborn Screening and Genetics Resource Center). A fourth condition, methylenetetrahydrofolate reductase (MTHFR) deficiency, is associated with developmental delays in physical and cognitive functions, as well as mental retardation and various psychiatric disturbances17
. However, this condition is incompletely penetrant and routine newborn screening is not conducted.
Similarly, it is unclear whether most of these variants are strongly associated with adult diseases for which treatments are available and that might qualify them for genetic notification. For the two separate amino acid substitutions detectable for MTHFR, Ala222Val (677C>T) and Glu429Ala (1298A>C), the evidence for association with cardiovascular disease remains uncertain and the indication for notification is weak. Glucose-6-phosphate dehydrogenase deficiency (Val68Met in G6PD, not measured in FHS), Familial Mediterranean fever (FMF, Pro369Ser, Arg408Gln in MEFV, for both FHS MAF=0.8%), the nonclassic form of congenital adrenal hyperplasia (Val237Glu, FHS MAF=0.02%), and hereditary pancreatitis (Asn34Ser in SPINK1, FHS MAF=1.0%) may manifest postnatally and are uncommon. In the FHS sample we observed a total of 6 homozygotes for the minor alleles of variants in these genes, for which the disease-related gene variants are incompletely penetrant. Knowledge regarding genetic status relating to these conditions might be beneficial, specifically for avoiding triggers for the severe episodic illnesses in hereditary pancreatitis and G6PD deficiency, colchicine prophylaxis in FMF, and steroid replacement in congenital adrenal hyperplasia (CAH); however, there is insufficient evidence regarding the penetrance of these conditions in general community-based populations, and whether there are significant benefits from genetic notification.
We examined evidence for return of results for each of these four conditions. For glucose-6-phosphate dehydrogenase deficiency, a G6PD
mutation, Val68Met, is present on genotyping arrays. However, this mutation has only been found to cause a disease phenotype in combination with another mutation, Asn126Asp, which is not present on any of the arrays18,19
, making interpretation of the status for Val68Met alone unclear. The MEFV
mutations present on arrays and related to the autosomal recessive FMF, for which 3 homozygotes have been potentially identified in FHS. Since onset of FMF typically occurs in adulthood, often via a difficult diagnosis after multiple exploratory surgeries, there may be a case for notification. However, the mutations identified here (Pro369Ser, Arg408Gln) are not among the most commonly observed disease variants, their role in FMF has not been clearly elucidated, and some studies suggest incomplete penetrance of these and other MEFV
. Thus, a careful clinical review and consideration of further mutational screening would likely be necessary in cases where notification regarding MEFV
mutation homozygosity was considered.
Individuals with the nonclassic form of CAH present postnatally and exhibit moderate enzyme deficiency and in some cases signs of hyperandrogenism. They may be heterozygous for one or more mutations or deletions (compound heterozygotes), including the Val237Glu mutation that is among a cluster of tested mutations in exon 6, and which is present on the array used in the FHS GWAS. In FHS, there were 3 potential heterozygotes for the Val237Glu mutation, which appears to be a functional null24
, raising the question of whether such participants might benefit from notification as it could explain and possibly lead to treatment of symptoms if there is also a second undetected mutation, which in combination, might lead to undiagnosed nonclassical CAH. Finally, for hereditary pancreatitis, while both heterozygosity and, particularly, homozygosity for the SPINK1
mutation (Asn34Ser) is clearly and strongly associated with symptomatic disease25
, the pancreatitis shows highly incomplete penetrance, likely due to required environmental triggers, such as infection and there may be little impact on treatment26
, although avoidance of alcohol and some drugs may be recommended and earlier recognition of first pancreatitis episode could be a benefit.
Genetic testing for two other common genetic mutations, F5
(Arg506Gln) and HFE
(Cys282Tyr), is sometimes conducted in adults with clinical manifestations of venous thromboembolism or hemochromatosis, respectively. Carriers of the Factor V Leiden variant (Arg506Gln) with a history of venous thromboembolism are at increased risk for a second thromboembolic event27
. However, there is no evidence of a clear benefit from screening asymptomatic persons for variants in the F5
. Furthermore, from available clinical trial evidence, there is little evidence that genetic testing predicts responsiveness or aids in decisions regarding use of more intensive anticoagulation in persons with recurrent venous thromboembolism29
, or that long-term prophylactic anticoagulation is beneficial for asymptomatic, heterozygous individuals. However, knowledge of Factor V Leiden carrier status might result in modifying exposure to thromboembolic risk factors, such as smoking or prophylactic aspirin use for sedentary periods (e.g., airplane flights). Factor V Leiden may contribute to pregnancy loss, however, testing for this variant is a usual part of the evaluation for recurrent pregnancy loss.
homozygotes have an increased prevalence of liver enzyme abnormalities with increased hepatic iron stores30
. There are no randomized trials assessing long term outcomes of phlebotomy in HFE
Cys282Tyr homozygotes. Since homozygous individuals may or may not have biochemical expression of iron overload, and many will not develop disease and end-organ damage, the use of phlebotomy is reserved for homozygotes with abnormal iron levels. In our current study, we identified 31 potential FHS homozygotes for the rare allele of Cys282Tyr in HFE
(FHS MAF=5.8%). While available clinical trial data are limited in asymptomatic homozygotes for HFE
variants, it is possible that manifestations of iron overload can be delayed or averted by interventions to reduce iron intake or overload31
. Best practice guidelines exist which do recommend a predictive referral of Cys282Tyr homozygotes for hemochromatosis examination in a clinical setting, but they do not suggest general population screening due to issues of incomplete penetrance32
. Even though HFE
is incompletely penetrant and a test has not been conducted in a CLIA lab, the measurement of iron levels is routine and safe, and if a clinical imbalance was detected a safe and effective treatment exists with routine phlebotomy. Thus, further consideration may be warranted of the potential benefits versus risks for reporting variants in the HFE
gene to research participants depending on the specific context of the research study in question. In some contexts important clinical information may be available to supplement knowledge of the genotype in deciding whether to return results (e.g., for F5
, if a medical history of DVT, or high iron levels, is known, respectively), though such situations invoke the boundary between research and clinical practice, how they are defined and what is expected in each.
While not an explicit goal of this study, we briefly consider the next steps, ethical obligations and caveats in the potential incidental notification process, a subject that has been discussed extensively elsewhere5,6,33,34
. If there is a clear medical benefit that outweighs the risks of notifying participants who have consented to such notification, then there seems to be an ethical obligation to inform. We did not immediately deem any of the variants considered here to clearly meet notifiability criteria. However, we decided to bring these findings to our independent Ethics Advisory Board to solicit input and a recommendation was made for further consultation with outside experts. If variants are deemed notifiable a number of additional practical steps might be considered, including: 1) consulting the consent form for the study to assess whether individuals clearly indicated an opt-in or opt-out preference to be informed, in recognition of individuals who would not want this information, 2) reviewing available genotype quality assessments (e.g., cluster plots), if any, for the variants in question to determine potential false positives, and considering additional validation genotyping in a CLIA-certified lab, 3) if clinical information is available and consent provided, conducting a clinical review for evidence of related, possibly undiagnosed disorders if this is deemed appropriate to the situation at hand, and 4) verifying that the participants are alive and can be contacted. Identification of resources including a specialized care provider, such as a genetic counselor and/or medical geneticist, and development of educational materials may also be warranted.