Helix Genetic Health Risk App for late-onset Alzheimer’s disease

K192073 · Helix Opco, LLC · PTA · Dec 23, 2020 · Immunology

Device Facts

• Record ID: K192073
• Device Name: Helix Genetic Health Risk App for late-onset Alzheimer’s disease
• Applicant: Helix Opco, LLC
• Product Code: PTA · Immunology
• Decision Date: Dec 23, 2020
• Decision: SESE
• Submission Type: Traditional
• Regulation: 21 CFR 866.5950
• Device Class: Class 2
• Attributes: Software as a Medical Device

Indications for Use

The Helix Genetic Health Risk App (HRA) uses qualitative genotyping to detect clinically relevant variants in genomic DNA isolated from human saliva collected from individuals ≥18 years with Oragene®Dx OGD-610 for the purpose of reporting and interpreting Genetic Health Risks (GHR): The Helix Genetic Health Risk App (HRA) for late-onset Alzheimer's disease is indicated for reporting of the e2/e2, e2/ e3, e2/e4 and e4/e4 genotypes in the APOE gene. The report describes if a person's genetic result is associated with an increased or decreased risk of developing late-onset Alzheimer's disease. The e2 and e4 variants included in this report are found and have been studied in many ethnicities. Detailed risk estimates have been studied the most in people of European descent. The Helix Genetic Health Risk App (HRA) is to be used with the Helix Laboratory Platform.

Device Story

Direct-to-consumer service; user self-collects saliva using Oragene Dx OGD-610 kit; sample shipped to CLIA-certified/CAP-accredited Helix Clinical Laboratory. DNA isolated and processed via whole exome sequencing on Helix Laboratory Platform (HLP) using Illumina NGS instrumentation. Proprietary software analyzes sequencing data to determine APOE genotypes (rs429358 and rs7412 SNPs). System categorizes genotypes into risk groups (decreased, average, increased) based on peer-reviewed literature. Personalized report provided via secure online portal. Intended to inform users of genetic risk; not a diagnostic tool. Requires physician consultation for clinical decision-making. Benefits include providing genetic risk information for late-onset Alzheimer's disease to facilitate informed discussions with healthcare providers.

Clinical Evidence

Bench testing only. Analytical performance validated via precision/reproducibility studies (n=24 samples, 100% concordance), interference studies (endogenous proteins, exogenous substances, smoking, microbial contamination), and accuracy studies (n=99 saliva samples, 100% accuracy vs. Sanger sequencing). Clinical validity supported by peer-reviewed literature meta-analyses across European, African, South Asian, and East Asian populations. User comprehension study (n=244) demonstrated 90.6–98.4% comprehension rates across categories.

Technological Characteristics

Whole exome sequencing assay; Illumina NGS instrumentation; Oragene Dx OGD-610 collection kit. Software-based analysis of rs429358 and rs7412 SNPs to determine APOE genotype. Cloud-based reporting platform. No mechanical/energy-based actuation.

Indications for Use

Indicated for individuals ≥18 years to report APOE genotypes (e2/e2, e2/e3, e3/e3, e2/e4, e3/e4, e4/e4) associated with risk of late-onset Alzheimer's disease. Not for diagnosis, treatment decisions, or determining current health status.

Regulatory Classification

Identification

A genetic health risk assessment system is a qualitative in vitro molecular diagnostic system used for detecting variants in genomic deoxyribonucleic acid (DNA) isolated from human specimens that will provide information to users about their genetic risk of developing a disease to inform lifestyle choices and/or conversations with a health care professional. This assessment system is for over-the-counter use. This device does not determine the person's overall risk of developing a disease.

Special Controls

*Classification.* Class II (special controls). The genetic health risk assessment system device, when it has previously received a first-time FDA marketing authorization (*e.g.,* 510(k) clearance) for the genetic health risk assessment system (a “one-time FDA reviewed genetic health risk assessment system”), is exempt from the premarket notification procedures in part 807, subpart E, of this chapter subject to the limitations in § 866.9. The device must comply with the following special controls:(1) The 21 CFR 809.10 compliant labeling and any prepurchase page and test report generated, unless otherwise specified, must include: (i) A section addressed to users with the following information: (A) The limiting statement explaining that this test provides genetic risk information based on assessment of specific genetic variants but does not report on a user's entire genetic profile. This test [does not/may not, as appropriate] detect all genetic variants related to a given disease, and the absence of a variant tested does not rule out the presence of other genetic variants that may be related to the disease. (B) The limiting statement explaining that other companies offering a genetic risk test may be detecting different genetic variants for the same disease, so the user may get different results using a test from a different company. (C) The limiting statement explaining that other factors such as environmental and lifestyle risk factors may affect the risk of developing a given disease. (D) The limiting statement explaining that some people may feel anxious about getting genetic test health results. This is normal. If the potential user feels very anxious, such user should speak to his or her doctor or other health care professional prior to collection of a sample for testing. This test is not a substitute for visits to a doctor or other health care professional. Users should consult with their doctor or other health care professional if they have any questions or concerns about the results of their test or their current state of health. (E) Information about how to obtain access to a genetic counselor, board-certified clinical molecular geneticist, or equivalent health care professional about the results of a user's test. (F) The limiting statement explaining that this test is not intended to diagnose a disease, tell you anything about your current state of health, or be used to make medical decisions, including whether or not you should take a medication or how much of a medication you should take. (G) A limiting statement explaining that the laboratory may not be able to process a sample, and a description of the next steps to be taken by the manufacturer and/or the customer, as applicable. (ii) A section in your 21 CFR 809.10 labeling and any test report generated that is for health care professionals who may receive the test results from their patients with the following information: (A) The limiting statement explaining that this test is not intended to diagnose a disease, determine medical treatment, or tell the user anything about their current state of health. (B) The limiting statement explaining that this test is intended to provide users with their genetic information to inform lifestyle decisions and conversations with their doctor or other health care professional. (C) The limiting statement explaining that any diagnostic or treatment decisions should be based on testing and/or other information that you determine to be appropriate for your patient. (2) The genetic test must use a sample collection device that is FDA-cleared, -approved, or -classified as 510(k) exempt, with an indication for in vitro diagnostic use in over-the-counter DNA testing. (3) The device's labeling must include a hyperlink to the manufacturer's public Web site where the manufacturer shall make the information identified in paragraph (b)(3) of this section publicly available. The manufacturer's home page, as well as the primary part of the manufacturer's Web site that discusses the device, must provide a hyperlink to the Web page containing this information and must allow unrestricted viewing access. If the device can be purchased from the Web site or testing using the device can be ordered from the Web site, the same information must be found on the Web page for ordering the device or provided in a publicly accessible hyperlink on the Web page for ordering the device. Any changes to the device that could significantly affect safety or effectiveness would require new data or information in support of such changes, which would also have to be posted on the manufacturer's Web site. The information must include: (i) An index of the material being provided to meet the requirements in paragraph (b)(3) of this section and its location. (ii) A section that highlights summary information that allows the user to understand how the test works and how to interpret the results of the test. This section must, at a minimum, be written in plain language understandable to a lay user and include: (A) Consistent explanations of the risk of disease associated with all variants included in the test. If there are different categories of risk, the manufacturer must provide literature references that support the different risk categories. If there will be multiple test reports and multiple variants, the risk categories must be defined similarly among them. For example, “increased risk” must be defined similarly between different test reports and different variant combinations. (B) Clear context for the user to understand the context in which the cited clinical performance data support the risk reported. This includes, but is not limited to, any risks that are influenced by ethnicity, age, gender, environment, and lifestyle choices. (C) Materials that explain the main concepts and terminology used in the test that include: ( *1* )*Definitions:* Scientific terms that are used in the test reports.( *2* )*Prepurchase page:* This page must contain information that informs the user about what information the test will provide. This includes, but is not limited to, variant information, the condition or disease associated with the variant(s), professional guideline recommendations for general genetic risk testing, the limitations associated with the test (*e.g.,* test does not detect all variants related to the disease) and any precautionary information about the test the user should be aware of before purchase. When the test reports the risk of a life-threatening or irreversibly debilitating disease or condition for which there are few or no options to prevent, treat, or cure the disease, a user opt-in section must be provided. This opt-in page must be provided for each disease that falls into this category and must provide specific information relevant to each test result. The opt-in page must include:( *i* ) An option to accept or decline to receive this specific test result;( *ii* ) Specification of the risk involved if the user is found to have the specific genetic test result;( *iii* ) Professional guidelines that recommend when genetic testing for the associated target condition is or is not recommended; and( *iv* ) A recommendation to speak with a health care professional, genetic counselor, or equivalent professional before getting the results of the test.( *3* )*Frequently asked questions (FAQ) page:* This page must provide information that is specific for each variant/disease pair that is reported. Information provided in this section must be scientifically valid and supported by corresponding publications. The FAQ page must explain the health condition/disease being tested, the purpose of the test, the information the test will and will not provide, the relevance of race and ethnicity to the test results, information about the population to which the variants in the test is most applicable, the meaning of the result(s), other risk factors that contribute to disease, appropriate followup procedures, how the results of the test may affect the user's family, including children, and links to resources that provide additional information.(iii) A technical information section containing the following information: (A) Gene(s) and variant(s) the test detects using standardized nomenclature, Human Genome Organization nomenclature and coordinates as well as Single Nucleotide Polymorphism Database (dbSNP) reference SNP numbers (rs#). (B) Scientifically established disease-risk association of each variant detected and reported by the test. This risk association information must include: ( *1* ) Genotype-phenotype information for the reported variants.( *2* ) Table of expected frequency and risks of developing the disease in relevant ethnic populations and the general population.( *3* ) A statement about the current professional guidelines for testing these specific gene(s) and variant(s).( *i* ) If professional guidelines are available, provide the recommendations in the professional guideline for the gene, variant, and disease, for when genetic testing should or should not be performed, and cautionary information that should be communicated when a particular gene and variant is detected.( *ii* ) If professional guidelines are not available, provide a statement that the professional guidelines are not available for these specific gene(s) and variant(s).(C) The specimen type ( *e.g.,* saliva, capillary whole blood).(D) Assay steps and technology used. (E) Specification of required ancillary reagents, instrumentation, and equipment. (F) Specification of the specimen collection, processing, storage, and preparation methods. (G) Specification of risk mitigation elements and description of all additional procedures, methods, and practices incorporated into the directions for use that mitigate risks associated with testing. (H) Information pertaining to the probability of test failure ( *i.e.,* percentage of tests that failed quality control) based on data from clinical samples, a description of scenarios in which a test can fail (*i.e.,* low sample volume, low DNA concentration, etc.), how users will be notified of a test failure, and the nature of followup actions on a failed test to be taken by the user and the manufacturer.(I) Specification of the criteria for test result interpretation and reporting. (J) Information that demonstrates the performance characteristics of the test, including: ( *1* ) Accuracy of study results for each claimed specimen type.( *i* ) Accuracy of the test shall be evaluated with fresh clinical specimens collected and processed in a manner consistent with the test's instructions for use. If this is impractical, fresh clinical samples may be substituted or supplemented with archived clinical samples. Archived samples shall have been collected previously in accordance with the instructions for use, stored appropriately, and randomly selected. In some limited circumstances, use of contrived samples or human cell line samples may also be appropriate and used as an acceptable alternative. The contrived or human cell line samples shall mimic clinical specimens as much as is feasible and provide an unbiased evaluation of the device accuracy.( *ii* ) Accuracy must be evaluated by comparison to bidirectional Sanger sequencing or other methods identified as appropriate by FDA. Performance criteria for both the comparator method and the device must be predefined and appropriate to the device's intended use. Detailed study protocols must be provided.( *iii* ) Test specimens must include all genotypes that will be included in the tests and reports. The number of samples tested in the accuracy study for each variant reported must be based on the variant frequency using either the minimum numbers of samples identified in this paragraph or, when determined appropriate and identified by FDA, a minimum number of samples determined using an alternative method. When appropriate, the same samples may be used in testing to demonstrate the accuracy of testing for multiple genotypes by generating sequence information at multiple relevant genetic locations. At least 20 unique samples representing the wild-type genotype must be tested. To test samples that are heterozygous for the reported variant(s), common variants (>0.1 percent variant frequency in the relevant population) must be tested with at least 20 unique samples. Rare variants (≤0.1 percent variant frequency in the relevant population) must be tested with at least three unique samples. To test samples that are homozygous for the reported variant(s), variants with ≥2 percent variant frequency in a relevant population must be tested with at least 20 unique samples. Variants with a frequency in the relevant population <2 percent and ≥0.5 percent must be tested with at least 10 unique samples. Variants with a frequency in the relevant population <0.5 percent must be tested with at least three unique samples. If variants with a frequency of <0.5 percent are not found within the relevant population and homozygous samples are not tested, then the test results for this homozygous rare variant must not be reported to the user.( *iv* ) Information about the accuracy study shall include the number and type of samples that were compared to bidirectional Sanger sequencing or other methods identified as appropriate by FDA. This information must either be reported in tabular format and arranged by clinically relevant variants or reported using another method identified as appropriate by FDA. As an example, for samples with different genotypes DD, Dd, and dd, the following table represents data from the accuracy study presented in tabular format: ( *v* ) The accuracy represents the degrees of agreement between the device results and the comparator results. The accuracy must be evaluated by measuring different percent agreements (PA) of device results with the comparator results and percent of 'no calls' or 'invalid calls.' Calculate the rate of 'no calls' and 'invalid calls' for each comparator output as %Inv(DD) = A4 /NDD, %Inv(Dd) = B4 /NDd , %Inv(dd) = C4 /Ndd . If 'no calls' or 'invalid calls' are required to be retested according to the device instructions for use, the percent of final 'no calls' or 'invalid calls' must be provided. In the table presenting the results of the accuracy study, use only the final results (*i.e.,* after retesting the initial 'no calls' or 'invalid calls', if required according to the instructions for use). Samples that resulted in a 'no call' or 'invalid call' after retesting must not be included in the final calculations of agreement. If the percentages of 'no calls' or 'invalid calls' for each comparator output are similar, combine these estimates as (A4 + B4 + C4 )/(NDD + NDd + Ndd ) and provide a 95 percent two-sided confidence interval. The percent of final 'no calls' or 'invalid calls' must be clinically acceptable.( *vi* ) Point estimates of percent agreement for each genotype must be calculated as the number of correct calls for that genotype divided by the number of samples known to contain that genotype excluding 'no calls' or 'invalid calls'. The calculations must be performed as follows: ( *vii* ) For percent agreements for DD, Dd and dd (PA(DD|DD), PA(Dd|Dd) and PA(dd|dd)) as described in paragraph (b)(3)(iii)(J)(*1* )(*vi* ) of this section, the 95 percent two-sided confidence intervals must be provided. The accuracy point estimates for percent agreements for DD, Dd and dd must be ≥99 percent per reported variant and overall. Any variants that have a point estimate for either PA(DD|DD), PA(Dd|Dd), or PA(dd|dd) of <99 percent compared to bidirectional sequencing or other methods identified as appropriate by FDA must not be incorporated into test claims and reports. Accuracy results generated from clinical specimens versus contrived samples or cell lines must be presented separately. Results must be summarized and presented in tabular format by sample type and by genotype or must be reported using another method identified as appropriate by FDA (see paragraph (b)(3)(iii)(J)(*1* )(*iv* ) of this section).( *viii* ) Information must be reported on the Technical Positive Predictive Value (TPPV) related to the analytical (technical) performance of the device for genotypes in each relevant subpopulation (*e.g.,* ethnicity, gender, age, geographical location, etc.). TPPV is the percentage of individuals with the genotype truly present among individuals whose test reports indicate that this genotype is present. The TPPV depends on the accuracy measures of percent agreements and on the frequency of the genotypes in the subpopulation being studied. The f(DD) is the frequency of DD and f(Dd) is the frequency of Dd in the subpopulation being studied; TPPV must be calculated as described in paragraphs (b)(3)(iii)(J)(*1* )(*ix* ) through (*xi* ) of this section.( *ix* ) For variants where the point estimates of PA(DD|DD), PA(Dd|Dd) and PA(dd|dd) are less than 100 percent, use these point estimates in TPPV calculations.( *x* ) Point estimates of 100 percent in the accuracy study may have high uncertainty about performance of the test in the population. If these variants are measured using highly multiplexed technology, calculate the random error rate for the overall device. The accuracy study described in paragraph (b)(3)(iii)(J) of this section in those cases is more to determine that there is no systematic error in such devices. In those cases, incorporate that rate in the estimation of the percent agreements as calculated in paragraph (b)(3)(iii)(J)(*1* )(*vi* ) of this section and include it in TPPV calculations.( *xi* ) The TPPV for subpopulations with genotype frequencies of f(dd), f(Dd) and f(DD) = 1−f(dd)−f(Dd) in the subpopulation is calculated as: ( *2* ) Precision and reproducibility data must be provided using multiple instruments and multiple operators, on multiple non-consecutive days, and using multiple reagent lots. The sample panel must either include specimens from the claimed sample type (*e.g.,* saliva) representing all genotypes for each variant (*e.g.,* wild type, heterozygous, and homozygous) or, if an alternative panel composition of specimens is identified by FDA as appropriate, a panel composed of those specimens FDA identified as appropriate. A detailed study protocol must be created in advance of the study and must include predetermined acceptance criteria for performance results. The percentage of samples that failed quality control must be indicated (*i.e.,* the total number of sample replicates for which a sequence variant cannot be called (no calls) or that fail sequencing quality control criteria divided by the total number of replicates tested). It must be clearly documented whether results were generated from clinical specimens, contrived samples, or cell lines. The study results shall report the variants tested in the study and the number of replicates for each variant, and what conditions were tested (*i.e.,* number of runs, days, instruments, reagent lots, operators, specimens/type, etc.). Results must be evaluated and presented in tabular format and stratified by study parameter (*e.g.,* by site, instrument(s), reagent lot, operator, and sample variant). The study must include all extraction steps from the claimed specimen type or matrix, unless a separate extraction reproducibility study for the claimed sample type is performed. If the device is to be used at more than one laboratory, different laboratories must be included in the reproducibility study and reproducibility across sites must be evaluated. Any no calls or invalid calls in the study must be listed as a part of the precision and reproducibility study results.( *3* )*Analytical specificity data:* Data must be provided that evaluates the effect of potential endogenous and exogenous interferents on test performance, including specimen extraction and variant detection. Interferents tested must include those reasonably likely to be potentially relevant to the sample type used for the device.( *4* )*Interfering variant data:* Nucleotide mutations that can interfere with the technology must be cited and evaluated. Data must be provided to demonstrate the effect of the interfering variant(s) on the performance of the correct calls. Alternatively, for each suspected interfering mutation for which data is not provided demonstrating the effect of the interfering variant, the manufacturer must identify the suspected interfering variants in the labeling and indicate that the impact that the interfering variants may have on the assay's performance has not been studied by providing a statement that reads “It is possible that the presence of [insert clearly identifying information for the suspected interfering variant] in a sample may interfere with the performance of this test. However, its effect on the performance of this test has not been studied.”( *5* )*Analytical sensitivity data:* Data must be provided demonstrating the minimum amount of DNA that will enable the test to perform correctly in 95 percent of runs.( *6* )*Reagent stability:* The manufacturer must evaluate reagent stability using wild-type, heterozygous, and homozygous samples. Reagent stability data must demonstrate that the reagents maintain the claimed accuracy and reproducibility. Data supporting such claims must be provided.( *7* )*Specimen type and matrix comparison data:* Specimen type and matrix comparison data must be generated if more than one specimen type can be tested with this device, including failure rates for the different specimens.(K) Clinical performance summary. ( *1* ) Information to support the clinical performance of each variant reported by the test must be provided.( *2* ) Manufacturers must organize information by the specific variant combination as appropriate (*e.g.,* wild type, heterozygous, homozygous, compound heterozygous, hemizygous genotypes). For each variant combination, information must be provided in the clinical performance section to support clinical performance for the risk category (*e.g.,* not at risk, increased risk). For each variant combination, a summary of key results must be provided in tabular format or using another method identified as appropriate by FDA to include the appropriate information regarding variant type, data source, definition of the target condition (*e.g.,* disease), clinical criteria for determining whether the target disease is present or absent, description of subjects with the target disease present and target disease absent (exclusion or inclusion criteria), and technical method for genotyping. When available, information on the effect of the variant on risk must be provided as the risk of a disease (lifetime risk or lifetime incidences) for an individual compared with the general population risk.( *i* ) If odds ratios are available, using information about the genotype distribution either among individuals with the target disease absent, or in the general population, or information about the risk variant frequency and odds ratios, the likelihood ratios for the corresponding device results along with 95 percent confidence intervals must be calculated. Using information about pretest risk (π), an estimate of likelihood ratio (LR), and a relationship between post-test risk R as R/(1−R) = LR·π/(1−π), the post-test risk R must be calculated.( *ii* ) When available, likelihood ratios (LR) for different test results must be presented in a tabular format along with references to the source data or using another method identified as appropriate by FDA as stated in paragraph (b)(3)(iii)(K)(2) of this section. When these values are not directly available in published literature, likelihood ratios can be separately calculated along with the 95 percent confidence interval with references to the source data. Note that a minimum requirement for the presence of the variant's effect on the risk is that a corresponding LR is statistically higher than 1 (a lower bound of 95 percent two-sided confidence interval is larger than 1). It means that the post-test risk is statistically higher than the pretest risk (an observed value of the difference between the post-test and pretest risks).(L) Materials that explain the main concepts and terminology used in the test that includes, but is not limited to: ( *1* )*Definitions:* Scientific terms that are used in the test reports.( *2* )*Prepurchase page:* This page must contain information that informs the user about what the test will provide. This includes, but is not limited to, variant information, the condition or disease associated with the variant(s), professional guideline recommendations for general genetic risk testing, the limitations associated with the test (*e.g.,* test does not detect all variants related to the disease) and any precautionary information about the test the user should be aware of before purchase. When the test reports the risk of a life-threatening or irreversibly debilitating disease or condition for which there are few or no options to prevent, treat, or cure the disease, a user opt-in section must be provided. This opt-in page must be provided for each disease that falls into this category and must provide specific information relevant to each test result. The opt-in page must include:( *i* ) An option to accept or decline to receive this specific test result;( *ii* ) Specification of the risk involved if the user is found to have the specific genetic test result;( *iii* ) Professional guidelines that recommend when genetic testing for the associated target condition is or is not recommended; and( *iv* ) A recommendation to speak with a health care professional, genetic counselor, or equivalent professional before getting the results of the test.( *3* ) Frequently asked questions (FAQ) page: This page must provide information that is specific for each variant/disease pair that is reported. Information provided in this section must be scientifically valid and supported by corresponding publications. The FAQ page must explain the health condition/disease being tested, the purpose of the test, the information the test will and will not provide, the relevance of race and ethnicity on the test results, information about the population to which the variants in the test is most applicable, the meaning of the result(s), other risks factors that contribute to disease, appropriate followup procedures, how the results of the test may affect the user's family, including children, and links to resources that provide additional information.(M) User comprehension study: Information on a study that assesses comprehension of the test process and results by potential users of the test must be provided. ( *1* ) The test manufacturer must provide a genetic risk education module to naïve user comprehension study participants prior to their participation in the user comprehension study. The module must define terms that are used in the test reports and explain the significance of genetic risk reports.( *2* ) The test manufacturer must perform pre- and post-test user comprehension studies. The comprehension test questions must include directly evaluating a representative sample of the material being presented to the user as described in paragraph (b)(3)(ii) of this section.( *3* ) The manufacturer must provide a justification from a physician and/or genetic counselor that identifies the appropriate general and variant-specific concepts contained within the material being tested in the user comprehension study to ensure that all relevant concepts are incorporated in the study.( *4* ) The user study must meet the following criteria:( *i* ) The study participants must comprise a statistically sufficient sample size and demographically diverse population (determined using methods such as quota-based sampling) that is representative of the intended user population. Furthermore, the study participants must comprise a diverse range of age and educational levels and have no prior experience with the test or its manufacturer. These factors shall be well defined in the inclusion and exclusion criteria.( *ii* ) All sources of bias must be predefined and accounted for in the study results with regard to both responders and non-responders.( *iii* ) The testing must follow a format where users have limited time to complete the studies (such as an onsite survey format and a one-time visit with a cap on the maximum amount of time that a participant has to complete the tests).( *iv* ) Users must be randomly assigned to study arms. Test reports in the user comprehension study given to users must define the target condition being tested and related symptoms, explain the intended use and limitations of the test, explain the relevant ethnicities in regard to the variant tested, explain genetic health risks and relevance to the user's ethnicity, and assess participants' ability to understand the following comprehension concepts: The test's limitations, purpose, appropriate action, test results, and other factors that may have an impact on the test results.( *v* ) Study participants must be untrained, be naïve to the test subject of the study, and be provided the labeling prior to the start of the user comprehension study.( *vi* ) The user comprehension study must meet the predefined primary endpoint criteria, including a minimum of a 90 percent or greater overall comprehension rate (*i.e.,* selection of the correct answer) for each comprehension concept. Other acceptance criteria may be acceptable depending on the concept being tested. Meeting or exceeding this overall comprehension rate demonstrates that the materials presented to the user are adequate for over-the-counter use.( *vii* ) The analysis of the user comprehension results must include results regarding reports that are provided for each gene/variant/ethnicity tested, statistical methods used to analyze all data sets, and completion rate, non-responder rate, and reasons for nonresponse/data exclusion. A summary table of comprehension rates regarding comprehension concepts (*e.g.,* purpose of test, test results, test limitations, ethnicity relevance for the test results, etc.) for each study report must be included.(4) The intended use of the device must not include the following indications for use: (i) Prenatal testing; (ii) Determining predisposition for cancer where the result of the test may lead to prophylactic screening, confirmatory procedures, or treatments that may incur morbidity or mortality to the patient; (iii) Assessing the presence of genetic variants that impact the metabolism, exposure, response, risk of adverse events, dosing, or mechanisms of prescription or over-the-counter medications; or (iv) Assessing the presence of deterministic autosomal dominant variants.

Predicate Devices

• 23andMe PGS Genetic Health Risk Report for Late-onset Alzheimer's Disease (DEN160026)

Submission Summary (Full Text)

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Food and Drug Administration
10903 New Hampshire Avenue
Silver Spring, MD 20993-0002
www.fda.gov

# 510(k) SUBSTANTIAL EQUIVALENCE DETERMINATION DECISION SUMMARY

ASSAY ONLY

## I Background Information:

A 510(k) Number

K192073

B Applicant

Helix OpCo, LLC

C Proprietary and Established Names

The Helix Genetic Health Risk App for late-onset Alzheimer's disease

D Regulatory Information

| Product Code(s) | Classification | Regulation Section | Panel |
| --- | --- | --- | --- |
| PTA | Class II | 21 CFR 866.5950 - Genetic Health Risk Assessment System | IM - Immunology |

## II Submission/Device Overview:

A Purpose for Submission:

New Device

B Measurand:

rs7412 SNP and rs429358 SNP in genomic DNA obtained from a human saliva sample

C Type of Test:

The Helix Genetic Health Risk App for late-onset Alzheimer's disease is intended to provide the late-onset Alzheimer's disease risk report. The report is based on a qualitative genetic test for detecting single nucleotide polymorphisms (SNP), rs429358 and rs7412, and for reporting e2, e3 and e4 alleles in the APOE gene. The Helix Genetic Health Risk App is for over-the-counter.

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## III Intended Use/Indications for Use:

### A Intended Use / Indications for Use:

The Helix Genetic Health Risk App (HRA) uses qualitative genotyping to detect clinically relevant variants in genomic DNA isolated from human saliva collected from individuals ≥18 years with Oragene Dx OGD-610 for the purpose of reporting and interpreting Genetic Health Risks (GHR):

The Helix Genetic Health Risk App (HRA) for late-onset Alzheimer’s disease is indicated for reporting of the e2/e2, e2/e3, e3/e3, e2/e4, e3/e4 and e4/e4 genotypes in the APOE gene. The report describes if a person's genetic result is associated with an increased or decreased risk of developing late-onset Alzheimer’s disease. The e2 and e4 variants included in this report are found and have been studied in many ethnicities. Detailed risk estimates have been studied the most in people of European descent.

The Helix Genetic Health Risk App (HRA) is to be used with the Helix Laboratory Platform.

### B Special Conditions for Use Statement(s):

- For over-the-counter (OTC) use.
- The customer must first opt-in to receive the late-onset Alzheimer’s disease risk report.
- The test is intended for users ≥ 18 years old.
- This test is not intended to diagnose a disease, determine medical treatment or other medical intervention, or tell the user anything about their current state of health.
- Any diagnostic or treatment decisions must be based on confirmatory prescription testing and/or other information that a healthcare professional determines to be appropriate for the patient, such as additional clinical testing and other risk factors that may affect individual risk and health care.
- This test is not a substitute for visits to a healthcare provider. It is recommended that you consult with a healthcare provider if you have any questions or concerns about your results.
- Subject to meeting the limitations contained in the special controls under regulation 21 CFR 866.5950.
- The laboratory may not be able to process a user’s sample. The probability that the laboratory cannot process a sample can be up to 2.5%.
- A user’s race, ethnicity, age, and sex may affect how the genetic test results are interpreted.
- The test does not detect all genetic variants associated with late-onset Alzheimer’s Disease (AD). The absence of a variant tested does not rule out the presence of other genetic variants that may be disease-related.
- The test does not describe a person's overall risk of developing Alzheimer’s Disease. In addition, other genetic and all non-genetic factors should be considered.
- Third-party application developers must discuss with FDA the usability of the original data generated on the Helix Laboratory Platform (HLP).

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C Special Instrument Requirements:

Helix Laboratory Platform

IV Device/System Characteristics:

A Device Description:

The HRA is an OTC and direct-to-consumer DNA genetic testing service. The HRA for late-onset Alzheimer’s disease reports the lifetime risk of developing Alzheimer’s disease at or above age 65 years based on six genotypes (e2/e2, e2/e3, e2/e4, e3/e3, e3/e4 and e4/e4) of the APOE gene combining qualitative genotyping data with descriptive information derived from peer-reviewed published genetic research studies. A customer’s saliva is self-collected using the Oragene Dx OGD-610 (K192920) manufactured by DNA Genotek, Inc., which consists of a sealable collection tube containing a stabilizing buffer solution. Once the saliva sample is collected, it is shipped to the Clinical Laboratory Improvement Amendments (CLIA)-certified and College of American Pathologists (CAP)-accredited Helix Clinical Laboratory for testing.

DNA is isolated from the saliva and tested using Helix’s proprietary whole exome sequencing assay in the Helix clinical laboratory. The genomic DNA is processed and sequenced using next generation sequencing (NGS) reagents and instrumentation manufactured by Illumina. The sequencing data is analyzed using Helix’s proprietary software, where the genetic variants of interest are determined. All samples must pass Helix’s quality control metrics prior to analysis. Samples that do not pass quality thresholds undergo re-sequencing and/or sample re-collection.

The APOE genotypes are used to generate personalized reports for each user. These reports tell the user which genetic variant(s) has/have been detected in their sample and provide information on the risk of disease associated with the genetic variants. If no variant was detected, that information is also provided. The personalized reports are designed to present scientific concepts to users in an easy-to-understand format. The reports provide scientifically valid information about the risks associated with the presence of a particular genetic variant. The reports are designed to help users understand the meaning of their results and any appropriate actions that may be taken based on their results.

B Principle of Operation:

DNA is isolated from a user’s saliva sample and tested on the HLP. The HLP detects the rs429358 and rs7412 variants in the APOE gene and determines each user’s APOE genotype. The HRA interprets and categorizes each user’s APOE genotype (e2/e2, e2/e3, e2/e4, e3/e3, e3/e4 and e4/e4) into risk groups (decreased, average, and increased) based on the estimated lifetime risk of developing late-onset Alzheimer’s disease.

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V Substantial Equivalence Information:

A Predicate Device Name(s):

23andMe PGS Genetic Health Risk Report for Late-onset Alzheimer's Disease

B Predicate 510(k) Number(s):

DEN160026

C Comparison with Predicate(s):

| Device & Predicate Device(s): | Device K192073 | Predicate DEN160026 |
| --- | --- | --- |
| Device Trade Name | The Helix Genetic Health Risk App for late-onset Alzheimer's disease | 23andMe PGS Genetic Health Risk Report for Late-onset Alzheimer's Disease |
| General Device Characteristic Similarities | | |
| Intended Use/Indications For Use | The Helix Genetic Health Risk App (HRA) uses qualitative genotyping to detect clinically relevant variants in genomic DNA isolated from human saliva collected from individuals ≥18 years with Oragene Dx OGD-610 for the purpose of reporting and interpreting Genetic Health Risks (GHR):
The Helix Genetic Health Risk App (HRA) for late-onset Alzheimer's disease is indicated for reporting of the e2/e2, e2/e3, e3/e3, e2/e4, e3/e4 and e4/e4 genotypes in the APOE gene. The report describes if a person's genetic result is associated with an increased or decreased risk of developing late-onset Alzheimer's disease. The e2 and e4 variants included in this report are found and have been studied in many ethnicities. Detailed risk estimates have been studied the most in people of European descent. | The 23andMe Personal Genome Service (PGS) Test uses qualitative genotyping to detect the following clinically relevant variants in genomic DNA isolated from human saliva collected from individuals ≥18 years with the Oragene Dx model OGD-500.001 for the purpose of reporting and interpreting Genetic Health Risks (GHR):
The 23andMe PGS Genetic Health Risk Report for Late-onset Alzheimer's Disease is indicated for reporting of the ε4 variant in the APOE gene. The report describes if a person's genetic result is associated with an increased risk of developing Late-onset Alzheimer's Disease, but it does not describe a person's overall risk of developing Alzheimer's Disease. The ε4 variant included in this report is found and has been studied in many ethnicities. Detailed risk estimates have been studied the |

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| Device & Predicate Device(s): | Device K192073 | Predicate DEN160026 |
| --- | --- | --- |
| | The Helix Genetic Health Risk App (HRA) is to be used with the Helix Laboratory Platform. | most in people of European descent. |
| Classification | Class II | Same |
| Product code | PTA | Same |
| Regulation | 21 CFR 866.5950 | Same |
| Target population | ≥ 18 years old | Same |
| Interpretation of results | For over the counter use (OTC). Specialized interpretation by a physician not required | Same |
| Human factors | User comprehension testing | Same |
| Design | Software application that includes product information page, e-commerce (registration and order DNA kit), secure login, download genetic report | Same |
| Measurand | DNA | Same |
| Sample type | Saliva | Same |
| General Device Characteristic Differences | | |
| Variants detected and alleles reported | APOE genotype (e2) is the result of a ‘T’ nucleotide at the rs7412 SNP and a ‘T’ nucleotide at the rs429358 SNP.
APOE genotype (e3) is the result of a ‘C’ nucleotide at the rs7412 SNP and a ‘T’ nucleotide at the rs429358 SNP.
APOE genotype (e4) is the result of a ‘C’ nucleotide at the rs7412 SNP and a ‘C’ nucleotide at the rs429358 SNP. | rs429358 SNP for APOE e4 allele |
| Technology | Next Generation Sequencing | Microarray genotyping |
| Method and sequencing platform | Whole exome sequencing is conducted using Helix Laboratory Platform (HLP) and pipeline analysis using Helix Bioinformatics Pipeline (FDA-cleared under DEN190035). | Magnetic bead DNA extraction is performed using Tecan Evo.
Genotyping and PCR analysis is conducted using a chip-based method with Illumina Infinium’s BeadChip v4 assay, the Illumina iScan System for detection and |

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| Device & Predicate Device(s): | Device K192073 | Predicate DEN160026 |
| --- | --- | --- |
| | | analysis is facilitated with the use of GenomeStudio and Coregen software. |
| Specimen collection kit | DNA Genotek Inc., Oragene Dx OGD-610 | DNA Genotek Inc., Oragene Dx OGD500.001 |

VI Standards/Guidance Documents Referenced:

- Guidance for Industry and Food and Drug Administration Staff - Format for Traditional and Abbreviated 510(k)s (2019)
- Guidance for the Content of Premarket Submissions for Software Contained in Medical Devices (2005)
- General Principles of Software Validation Guidance for Industry and FDA Staff (2002)
- Off-The-Shelf Software Use in Medical Devices Guidance for Industry and Food and Drug Administration Staff (2019)
- Content of Premarket Submissions for Management of Cybersecurity in Medical Devices (2014)
- Class II Special Control: 866.5950 - Code of Federal Regulations Title 21
- Direct-to-Consumer test (https://www.fda.gov/medical-devices/vitro-diagnostics/direct-consumer-tests)

VII Performance Characteristics (if/when applicable):

A Analytical Performance:

All results presented below met the sponsor’s pre-defined acceptance criteria outlined in the Special Controls of 21 CFR 866.5950. Information regarding samples that failed quality control (FQC) was also evaluated and presented in each study below.

1. Precision / Reproducibility:

Reproducibility of the APOE genotype calls made by the HRA for late-onset Alzheimer’s disease was assessed by testing 24 samples that include six human B-lymphocyte cell lines (6 cell lines hereafter) and 18 unique saliva-derived DNA samples) in two independent studies (Study 1 and Study 2). The APOE genotype calls for the in the well-characterized cell line samples were compared to known APOE genotypes.

Study 1 tested 24 samples (6 cell lines and 18 unique saliva-derived DNA samples) with up to 72 replicates (3 replicates / sample / library prep plate x 3 plates x 2 enrichments x 4 independent runs of cBot and HiSeq instruments) for the APOE genotype calls. The test results are summarized below:

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| Cell Line | N | APOE Genotype | No. of replicates | No. of Correct Genotype Calls | No. of Incorrect Genotype Calls | No. of FQC | Percent of FQC (%) | Percent Correct Genotype Calls (%) |
| --- | --- | --- | --- | --- | --- | --- | --- | --- |
| Study 1: Cell lines | | | | | | | | |
| NA24385 | 1 | e2/e3 | 72 | 72 | 0 | 0 | 0 | 100 |
| NA24149 | 1 | e2/e4 | 72 | 72 | 0 | 0 | 0 | 100 |
| | 1 | e3/e3 | 72 | 72 | 0 | 0 | 0 | 100 |
| NA12877 | 1 | e3/e4 | 72 | 72 | 0 | 0 | 0 | 100 |
| NA24143 | 1 | e3/e4 | 72 | 72 | 0 | 0 | 0 | 100 |
| NA24631 | 1 | e4/e4 | 72 | 72 | 0 | 0 | 0 | 100 |
| Study 1: Clinical Samples | | | | | | | | |
| | 4 | e2/e3 | 288 | 276 | 0 | 12 | 4.2 | 100 |
| | 2 | e2/e4 | 144 | 141 | 0 | 3 | 2.1 | 100 |
| | 7 | e3/e3 | 504 | 484 | 0 | 20 | 4.0 | 100 |
| | 2 | e3/e4 | 144 | 136 | 0 | 8 | 5.6 | 100 |
| | 2 | e4/e4 | 144 | 141 | 0 | 3 | 2.1 | 100 |
| | 1 | Unknown | 72 | N/A | N/A | 72 | 100 | N/A |

Study 2 tested 24 samples (6 cell lines and 18 unique saliva-derived DNA samples) with up to 54 replicates (3 replicates / sample / library prep plate x 3 plates x 2 enrichments x 3 reagent lots) for the APOE genotype calls. The test results are summarized below:

| Cell Line | N | APOE Genotype | No. of replicates | No. of Correct Genotype Calls | No. of Incorrect Genotype Calls | No. of FQC | Percent of FQC (%) | Percent Correct Genotype Calls (%) |
| --- | --- | --- | --- | --- | --- | --- | --- | --- |
| Study 2: Cell lines | | | | | | | | |
| NA24385 | 1 | e2/e3 | 54 | 54 | 0 | 0 | 0 | 100 |
| NA24149 | 1 | e2/e4 | 54 | 54 | 0 | 0 | 0 | 100 |
| NA12878 | 1 | e3/e3 | 54 | 54 | 0 | 0 | 0 | 100 |
| NA12877 | 1 | e3/e4 | 54 | 54 | 0 | 0 | 0 | 100 |
| NA24143 | 1 | e3/e4 | 54 | 54 | 0 | 0 | 0 | 100 |

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| Cell Line | N | APOE Genotype | No. of replicates | No. of Correct Genotype Calls | No. of Incorrect Genotype Calls | No. of FQC | Percent of FQC (%) | Percent Correct Genotype Calls (%) |
| --- | --- | --- | --- | --- | --- | --- | --- | --- |
| NA24631 | 1 | e4/e4 | 54 | 48 | 0 | 6 | 11.1 | 100 |
| Stud 2: Clinical Samples | | | | | | | | |
| | 2 | e2/e3 | 108 | 108 | 0 | 0 | 0 | 100 |
| | 7 | e3/e3 | 378 | 377 | 0 | 1 | 0.3 | 100 |
| | 7 | e3/e4 | 378 | 376 | 0 | 2 | 0.5 | 100 |
| | 2 | e4/e4 | 108 | 108 | 0 | 0 | 0 | 100 |

The calls for concordance the APOE genotypes were analyzed in the clinical samples. Genotyping results produced 100% of replicates that were called correctly for all APOE genotypes.

2. Linearity:

Not applicable

3. Analytical Specificity / Interference:

Four studies were performed to determine the effects of substances found in saliva that may interfere with the performance of the HRA for late-onset Alzheimer's disease. Per study protocol, for sequenced samples to be included in the data analyses (e.g., evaluable samples), the sample(s) must pass pre-defined QC thresholds.

a. Study 1 evaluated four endogenous proteins commonly found in saliva. Each of the following were added to saliva samples: alpha-amylase (395 U/mL), hemoglobin (20 mg/mL), immunoglobulin A (IgA) (0.43 mg/mL), and albumin (2.67mg/mL). The study showed that these proteins did not affect test performance for saliva samples (n=29–30 evaluable samples across four endogenous proteins). The test results are shown below.

| APOE Genotype | No. of replicates | No. of Correct Genotype Calls | No. of Incorrect Genotype Calls | No. of FQC | Percent of FQC (%) | Percent Correct Genotype (%) |
| --- | --- | --- | --- | --- | --- | --- |
| e2/e3 | 18 | 18 | 0 | 0 | 0 | 100 |
| e2/e4 | 4 | 4 | 0 | 0 | 0 | 100 |
| e3/e3 | 76 | 75 | 0 | 1 | 1.3 | 100 |

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| APOE Genotype | No. of replicates | No. of Correct Genotype Calls | No. of Incorrect Genotype Calls | No. of FQC | Percent of FQC (%) | Percent Correct Genotype (%) |
| --- | --- | --- | --- | --- | --- | --- |
| e3/e4 | 18 | 18 | 0 | 0 | 0 | 100 |
| e4/e4 | 4 | 4 | 0 | 0 | 0 | 100 |

b. In study 2, saliva samples were tested before and after (either immediately or 30 minutes after) exposure to one of four exogenous substances: eating food, drinking liquids, using mouthwash, or chewing gum. The study showed that exogenous substances did not interfere with test performance for saliva samples (n=12–15 evaluable samples across testing time for four exogenous substances. The test results are shown below.

| APOE Genotype | No. of replicates | No. of Correct Genotype Calls | No. of Incorrect Genotype Calls | No. of FQC | Percent of FQC (%) | Percent Correct Genotype Calls (%) |
| --- | --- | --- | --- | --- | --- | --- |
| e2/e4 | 12 | 12 | 0 | 0 | 0 | 100 |
| e3/e3 | 90 | 83 | 0 | 7 | 7.8 | 100 |
| e3/e4 | 24 | 21 | 0 | 3 | 12.5 | 100 |
| Unknown | 6 | N/A | N/A | 6 | 100 | N/A |

c. Study 3 evaluated saliva samples tested at 60 minutes before smoking, immediately after smoking, and 30 minutes after smoking and showed that smoking did not interfere with test performance (n=15 evaluable samples across three smoking conditions). The test results are shown below.

| APOE Genotype | No. of replicates | No. of Correct Genotype Calls | No. of Incorrect Genotype Calls | No. of FQC | Percent of FQC (%) | Percent Correct Genotype Calls (%) |
| --- | --- | --- | --- | --- | --- | --- |
| e3/e3 | 18 | 18 | 0 | 0 | 0 | 100 |
| e3/e4 | 12 | 12 | 0 | 0 | 0 | 100 |

d. In study 4, bacterial DNA from a commercial source (American Type Culture Collection) was added in various amounts into six cell line DNA samples (NA24385, NA24149, NA12878, NA12877, NA24143, and NA24631) to evaluate the effects of bacterial contamination in the test performance. This bacterial sample (ATCC MSA-1003) is comprised of 20 fully sequenced cultures that encompass a variety of characteristics including bacterial species found in

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mouse and oral cavity. The six cell lines were tested across five levels of bacterial content (0%, 10%, 20%, 30%, and 50%). This study showed that microbial DNA did not interfere with test performance (n=11–18 evaluable samples across five levels of bacterial content). Study 4 also tested various amounts of microbial and yeast DNA added to the saliva samples from three volunteers (0%, 10% Candida albicans, and 30% of ATCC MSA-1003). This study showed that yeast DNA did not interfere with test performance in clinical samples. The test results are shown below.

| APOE Genotype | No. of replicates | No. of Correct Genotype Calls | No. of Incorrect Genotype Calls | No. of FQC | Percent of FQC (%) | Percent Correct Genotype Calls (%) |
| --- | --- | --- | --- | --- | --- | --- |
| Cell lines | | | | | | |
| e2/e3 | 15 | 15 | 0 | 0 | 0 | 100 |
| e2/e4 | 15 | 13 | 0 | 2 | 13.3 | 100 |
| e3/e3 | 15 | 11 | 0 | 4 | 26.7 | 100 |
| e3/e4 | 30 | 28 | 0 | 2 | 6.7 | 100 |
| e4/e4 | 15 | 14 | 0 | 1 | 6.7 | 100 |
| Clinical samples | | | | | | |
| e3/e3 | 12 | 12 | 0 | 0 | 0 | 100 |
| e3/e4 | 6 | 6 | 0 | 0 | 0 | 100 |

4. Assay Reportable Range: Not applicable

5. Traceability, Stability, Expected Values (Controls, Calibrators, or Methods):

Refer to K192920 for pre-collection shelf-life stability of the collection device, stability of samples post-saliva collection, and freeze-thaw stability of samples stored in the Oragene Dx collection device.

6. Detection Limit:

The detection limit study evaluated the impact of different levels of DNA input on the performance of the HRA for late-onset Alzheimer's disease. The study yielded concordant test results for all 19 saliva samples with known APOE genotypes when tested at sample DNA concentrations between 3.5 to 10 ng/μL, an input corresponding to a range of 35 to 100 ng of DNA in the library preparation.

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| APOE Genotype | No. of replicates | No. of Correct Genotype Calls | No. of Incorrect Genotype Calls | No. of FQC | Percent of FQC (%) | Percent Correct Genotype Calls (%) |
| --- | --- | --- | --- | --- | --- | --- |
| e2/e3 | 12 | 12 | 0 | 0 | 0 | 100 |
| e3/e3 | 84 | 79 | 0 | 5 | 6 | 100 |
| e3/e4 | 18 | 18 | 0 | 0 | 0 | 100 |

## B Comparison Studies:

### 1. Method Comparison:

Three method comparison studies were conducted to evaluate accuracy: the first study was conducted with human saliva samples with known genotypes and a second study was conducted with human cell line samples with known genotypes to determine the rates of correct APOE genotype calls. A third study was conducted with human saliva and/or human cell line samples with known variants in genes other than the APOE gene to determine the agreement of the genotype calls.

#### a) Accuracy study with human saliva samples:

Accuracy of the HRA for late-onset Alzheimer's disease was evaluated by testing human saliva samples with known APOE genotypes. The presence of the two variants in the APOE gene was analyzed on the HLP and the genotyping results were compared to the known genotypes confirmed by Sanger sequencing (comparator). The accuracy for detecting the two variants in the APOE gene on the HLP (genotype call) using DNA isolated from human saliva samples (n=99) was 100% with a lower bound of 95% CI as 96.3% for all samples tested. The test results are shown in the table below.

| APOE Genotype | N | No. of Correct Genotype Calls | No. of Incorrect Genotype Calls | No. of FQC | Percent of FQC (%) | Percent Correct Genotype Calls (%) |
| --- | --- | --- | --- | --- | --- | --- |
| e2/e2 | 12 | 12 | 0 | 0 | 0 | 100 |
| e2/e3 | 17 | 17 | 0 | 0 | 0 | 100 |
| e2/e4 | 10 | 10 | 0 | 0 | 0 | 100 |
| e3/e3 | 20 | 20 | 0 | 0 | 0 | 100 |
| e3/e4 | 20 | 20 | 0 | 0 | 0 | 100 |
| e4/e4 | 20 | 20 | 0 | 0 | 0 | 100 |
| All | 99 | 99 | 0 | 0 | 0 | 100 |

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b) Accuracy study with human cell line samples:

Accuracy for detecting two variants, rs7412 and rs429358, in the APOE gene on the HLP was evaluated by using six cell lines with known APOE genotypes (e2/e3, e2/e4, e3/e3, e3/e4, e3/e4, and e4/e4). The accuracy for detecting the two variants in the APOE gene was 100% for all samples tested.

| Cell Line | N | APOE Genotype | No. of replicates | No. of Correct Genotype Calls | No. of Incorrect Genotype Calls | No. of FQC | Percent of FQC (%) | Percent Correct Genotype Calls (%) |
| --- | --- | --- | --- | --- | --- | --- | --- | --- |
| NA24385 | 1 | e2/e3 | 1 | 1 | 0 | 0 | 0 | 100 |
| NA24149 | 1 | e2/e4 | 1 | 1 | 0 | 0 | 0 | 100 |
| NA12878 | 1 | e3/e3 | 1 | 1 | 0 | 0 | 0 | 100 |
| NA12877 | 1 | e3/e4 | 1 | 1 | 0 | 0 | 0 | 100 |
| NA24143 | 1 | e3/e4 | 1 | 1 | 0 | 0 | 0 | 100 |
| NA24631 | 1 | e4/e4 | 1 | 1 | 0 | 0 | 0 | 100 |

c) Accuracy study with 3,295 human saliva and cell line samples with known variants in genes other than APOE:

In addition to the two variants in the APOE gene, accuracy of the HRA for late-onset Alzheimer's disease for detecting variants in the genes associated with other clinical conditions included in DEN160026 was evaluated on the HLP using specimens carrying unique genetic variants linked to the specific clinical conditions as listed in the table below:

| Clinical condition | Gene | SNP |
| --- | --- | --- |
| Hereditary Thrombophilia | FS | rs6025 |
| Hereditary Thrombophilia | F2 | rs1799963 |
| Alpha-1 Antitrypsin Deficiency | SERPINA1 | rs28929474 |
| Alpha-1 Antitrypsin Deficiency | SERPINA1 | rs17580 |
| Late-Onset Alzheimer's Disease | APOE | rs429358 |
| Parkinson's Disease | LRRK2 | rs34637584 |
| Parkinson's Disease; Gaucher Disease Type 1 | GBA | rs76763715 |
| Gaucher Disease Type 1 | GBA | rs387906315 |
| Gaucher Disease Type 1 | GBA | rs80356769 |
| Factor XI Deficiency | FXI | rs121965064 |
| Factor XI Deficiency | FXI | rs121965063 |
| Factor XI Deficiency | FXI | rs373297713 |
| Celiac Disease | HLA-DQA1 | rs2187668 |
| Glucose-6-Phosphate-Dehydrogenase Deficiency | G6PD | rs1050828 |
| Hereditary Hemochromatosis | HFE | rs1800562 |
| Hereditary Hemochromatosis | HFE | rs1799945 |
| Early-Onset Primary Dystonia | DYT1 | rs724159981 |

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The overall number of true positive, false positives, true negative and false negatives were analyzed for 17 variants in 11 genes. A total of 4,282 true positive and 51,731 true negative calls were reported. There were no false positive and no false negative results reported for the known variants tested and one no call was reported each for rs76763715 and rs429358.

2. Matrix Comparison:
Not applicable

C Clinical Studies:

1. Clinical Sensitivity:
Not applicable

2. Clinical Specificity:
Not applicable

3. Other Clinical Supportive Data (When 1. and 2. Are Not Applicable):

1) Risk of developing late-onset Alzheimer's disease and APOE allele frequency

Late-onset Alzheimer's disease is characterized by onset of the disease in people at or older than age 65 years. Many factors have been suggested to be associated with a decreased or increased risk of developing late-onset Alzheimer's disease, including age, genetic variants, family history of dementia, and lifestyle. The association of the APOE e2 and e4 alleles with the modified risk of developing late-onset Alzheimer's disease is one of the well-known genetic determinants across various ethnicities based on peer-reviewed scientific literature. The clinical performance and genetic variant detection by the HRA for late-onset Alzheimer's disease are supported by peer-reviewed scientific literature. The APOE e2 allele is associated with a decreased risk to develop late-onset Alzheimer's disease at or above age 65 years, whereas the APOE e4 allele is associated with an increased risk to develop late-onset Alzheimer's disease at or above age 65 years. The HLP detects the two single nucleotide polymorphisms (SNPs), rs429358 and rs7412, that determine the status of the e2, e3, and e4 alleles of the APOE gene.

The different combinations of the APOE alleles (e2, e3 and e4) were grouped based on the risk of developing late-onset Alzheimer's disease as follows:

a. Individuals with the APOE e3/e3 genotype have an average risk of developing late-onset Alzheimer's disease.

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b. Individuals with the APOE e2/e2 or e2/e3 genotype have a decreased risk of developing late-onset Alzheimer's disease compared to individuals with the APOE e3/e3 genotype for the European, African, and South Asian descent.

c. Individuals with the APOE e2/e4, e3/e4, or e4/e4 genotype have an increased risk of developing late-onset Alzheimer's disease compared to individuals with the APOE e3/e3 genotype.

d. Individuals who have none of these APOE genotypes will not receive information on their relative risk to develop late-onset Alzheimer's disease due to the lack of studies in the scientific literature.

Three APOE alleles covered by this test are found in people of all ethnicities. The percentage of the population carrying each of the six possible APOE genotypes will vary from study to study based on the enrollment criteria of the genetic study. A summary of the frequency of each APOE genotype in different ethnicities collected from multiple published peer-reviewed scientific studies is presented in the table below:

| | | Allele frequency (%) | | | | | | |
| --- | --- | --- | --- | --- | --- | --- | --- | --- |
| | | e2/e2 | e2/e3 | e3/e3 | e2/e4 | e3/e4 | e4/e4 | Reference |
| | | Decreased risk | | Average risk | Increased risk | | | |
| European descent | AD* | 0.2 | 4.8 | 36.4 | 2.6 | 41.1 | 14.8 | Farrer et al., 1997 |
| | controls | 0.8 | 12.7 | 60.9 | 2.6 | 21.3 | 1.8 | |
| African American descent | AD | 0.6 | 7.4 | 37.7 | 3.7 | 37.7 | 13.0 | Murrell et al., 2006 |
| | controls | 1.9 | 15.1 | 51.3 | 4.1 | 24.2 | 3.5 | |
| South Asian descent | AD | 0.2 | 6.2 | 44.8 | 6.2 | 37.2 | 5.3 | Agarwal et al., 2014 |
| | controls | 2.0 | 11.8 | 72.0 | 1.5 | 11.7 | 0.9 | |
| East Asian descent | AD | 0.9 | 7.9 | 51.0 | 3.3 | 29.3 | 7.6 | Liu et al., 2014 |
| | controls | 0.8 | 11.8 | 73.1 | 1.7 | 12.4 | 0.3 | |

*AD = Alzheimer's disease

Using the same references, the likelihood ratios (LR) for APOE e2, e3 and e4 combinations in different ethnicities, which represent an estimate of how the test results affects the chances of a condition was calculated as shown in the table below.

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| Ethnicity | Test result | Genotype | LR | 95% CI for LR | References | Study summary |
| --- | --- | --- | --- | --- | --- | --- |
| European descent | Decreased risk | e2/e2 | 0.3 | 0.1–0.5 | Farrer et al., 1997 | A meta-analysis of 5,930 patients who met criteria for probable or definite Alzheimer’s disease and 8,607 controls. Among study participants, there were 5,107 Alzheimer’s disease patients from European descent, and 6,262 controls from European descent. |
| | | e2/e3 | 0.4 | 0.3–0.4 | | |
| | Average risk | e3/e3 | 0.6 | 0.57–0.62 | | |
| | Increased risk | e2/e4 | 1 | 0.8–1.3 | | |
| | | e3/e4 | 1.9 | 1.8–2.1 | | |
| | | e4/e4 | 8.2 | 6.8–10.0 | | |
| African American descent | Decreased risk | e2/e2 | 0.3 | 0.04–2.7 | Murrell et al., 2006 | This study included 162 individuals from African descent (African American) with Alzheimer’s disease and 318 controls from African descent (African American). |
| | | e2/e3 | 0.5 | 0.3–0.9 | | |
| | Average risk | e3/e3 | 0.7 | 0.6–0.9 | | |
| | Increased risk | e2/e4 | 0.9 | 0.4–2.3 | | |
| | | e3/e4 | 1.6 | 1.2–2.1 | | |
| | | e4/e4 | 3.8 | 1.9–7.6 | | |
| South Asian descent | Decreased risk | e2/e2 | 0.1 | 0.02–0.9 | Agarwal et al., 2014 | A meta-analysis of 417 individuals from South Asian descent with Alzheimer’s disease and 651 controls from South Asian descent. |
| | | e2/e3 | 0.5 | 0.3–0.8 | | |
| | Average risk | e3/e3 | 0.6 | 0.6–0.7 | | |
| | Increased risk | e2/e4 | 4.1 | 2.0–8.3 | | |
| | | e3/e4 | 3.2 | 2.5–4.1 | | |
| | | e4/e4 | 5.7 | 2.3–14.0 | | |
| East Asian descent | Average risk | e2/e2 | 1.1* | 0.5– 2.3 | Liu et al., 2014 | A meta-analysis of 1,576 individuals from East Asian descent with Alzheimer’s disease and 1,741 controls from East Asian descent. (*limited number of e2/e2 samples used for analysis) |
| | | e2/e3 | 0.7 | 0.5–0.8 | | |
| | | e3/e3 | 0.7 | 0.6–0.7 | | |
| | Increased risk | e2/e4 | 1.9 | 1.2–3.0 | | |
| | | e3/e4 | 2.5 | 2.2–2.9 | | |
| | | e4/e4 | 25.6 | 10.5–62.6 | | |

2) User comprehension study

a. Saliva collection kit user study:
Refer to K192920 for the saliva collection device (Oragene Dx OGD-610) instructions for use and to assess the ability of lay users to provide samples adequate for testing.

b. HRA user comprehension study:

Objectives:
The user comprehension study was performed to assess user comprehension of the HRA reports. The study was performed with a demographically diverse group of study participants (e.g. age, gender, ethnicity, and education level) by asking questions for five comprehension categories (purpose of test, limitation, relevant ethnicities, meaning of results, and appropriate follow-up) of the HRA in the pre-purchase page and the test reports in a controlled online setting.

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# Methods:

Quota-based sampling per U.S. Census was employed to recruit a diverse set of study participants based on their demographic variables in age, gender, ethnicity, and education level. Study participants who enrolled into the user comprehension study were randomly assigned into one of four risk category groups (increased, average, decrease, and result not available). Study participants took the test on their personal desktop. Voice and screen recordings of each study participant are captured during the test. The user comprehension was tested through a two-step process. First, participants' understanding of genetics was tested prior to viewing the pre-purchase page and test reports. Second, participants were shown the pre-purchase page and the test reports and completed the comprehension assessment. A total of 303 study participants met the inclusion criteria and were enrolled in the study. Fifty-nine (59) participants were non-complete users and removed from final analysis due to one of the following reasons across four risk category groups.

- Exited test: an automated reason generated by Usertesting.com when their software detects there was some type of technical difficulty that resulted in the participant leaving the study prior to completion.
- Session Expired is an automated reason generated by Usertesting.com when their software detects that a participant does not start the study within 30 minutes.
- Uploading Issue is an automated reason generated by Usertesting.com when their software detects that a participant had issues uploading the test answers to the testing site.

The number of non-complete users in each risk category group were as follows: 16 users (20%) in the increased risk group, 9 users (12.8%) in the average risk group, 12 users (16.4%) in the decreased risk group, and 22 users (26.5%) in the result not available group. There were no timed-out users in this study. A total of 244 study participants assigned to a risk category group across all risk categories (increased, average, decrease, and result not available) were included in the comprehension rate analysis. The completion rate was 100% for all 244 study participants.

# Results:

The overall comprehension rate per comprehension category ranged 90.6 to $98.4\%$ across all risk categories. The comprehension rate within the risk category ranged from 85.2 to $100\%$ per comprehension category. The user comprehension study results are shown in the table below.

| Comprehension category | Comprehension rates (%) within Risk category | | | | Overall comprehension rate (%) |
| --- | --- | --- | --- | --- | --- |
| | Increased Risk | Average Risk | Decreased Risk | Result not available | |
| Purpose of Test | 96.7 | 98.4 | 100 | 98.4 | 98.4 |
| Limitation | 93.4 | 92.6 | 100 | 96.7 | 95.7 |
| Relevant Ethnicities | 90.2 | 91.8 | 93.4 | 91.8 | 91.8 |
| Meaning of Results | 85.2 | 89.6 | 90.7 | 96.7 | 90.6 |
| Appropriate Follow-up | 96.7 | 95.9 | 100 | 96.7 | 97.3 |

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The comprehension rates were also analyzed by age, gender, race, and education level for each comprehension category. The comprehension rates ranged 87–100% across ages, 93–99% across genders, 79–100% across ethnicities, and 87–100% across education levels as summarized in the table as below.

| | Purpose of Test | | Limitations | | Relevant Ethnicities | | Meaning of Results | | Appropriate follow-up | |
| --- | --- | --- | --- | --- | --- | --- | --- | --- | --- | --- |
| | % | correct responses / total responses | % | correct responses / total responses | % | correct responses / total responses | % | correct responses / total responses | % | correct responses / total responses |
| Age (group) | | | | | | | | | | |
| 18-24 | 98 | 39/40 | 93 | 74/80 | 87 | 35/40 | 94 | 75/80 | 98 | 78/80 |
| 25-34 | 100 | 43/43 | 94 | 81/86 | 95 | 41/43 | 99 | 85/85 | 100 | 96/86 |
| 35-44 | 100 | 42/42 | 98 | 82/84 | 93 | 39/42 | 96 | 81/84 | 100 | 84/84 |
| 45-54 | 100 | 44/44 | 98 | 86/88 | 89 | 39/44 | 95 | 84/88 | 100 | 88/88 |
| 55-64 | 100 | 35/36 | 97 | 70/72 | 92 | 33/36 | 94 | 68/72 | 97 | 70/72 |
| 65+ | 95 | 37/39 | 96 | 74/78 | 94 | 37/39 | 97 | 76/78 | 95 | 74/78 |
| Gender (male or female) | | | | | | | | | | |
| Male | 98 | 123/124 | 97 | 233/240 | 93 | 112/120 | 97 | 232/240 | 98 | 234/240 |
| Female | 98 | 117/120 | 94 | 234/248 | 90 | 112/124 | 96 | 237/248 | 99 | 246/248 |
| Race/Ethnicity | | | | | | | | | | |
| Caucasian | 99 | 139/140 | 97 | 272/280 | 94 | 131/140 | 97 | 272/281 | 99 | 278/280 |
| Black | 94 | 32/34 | 93 | 63/68 | 79 | 27/34 | 91 | 62/68 | 94 | 64/68 |
| Hispanic / Latino | 98 | 40/41 | 94 | 77/82 | 95 | 39/41 | 95 | 77/81 | 98 | 80/82 |
| All other ethnicities | 100 | 29/29 | 95 | 55/58 | 93 | 27/29 | 100 | 58/58 | 100 | 58/58 |
| Education Level | | | | | | | | | | |
| High School | 99 | 83/84 | 94 | 158/168 | 90 | 76/84 | 94 | 158/168 | 99 | 166/168 |
| Some College | 98 | 53/54 | 97 | 105/108 | 93 | 50/54 | 93 | 100/107 | 98 | 106/108 |
| College or equivalent | 97 | 71/73 | 97 | 142/146 | 95 | 69/74 | 100 | 144/144 | 97 | 142/146 |
| Post Graduate | 100 | 33/33 | 94 | 62/66 | 87 | 29/33 | 97 | 67/69 | 100 | 66/66 |

c. Frequently Asked Questions Material:

A Frequently Asked Questions (FAQ) section was developed and included in each Genetic Health Risk Report. The FAQ section was to provide users information to adequately understand the purpose, limitations, and the meaning of the results of the test, and was developed using methodology consistent with the Manufacturer's labeling design, identification of communication messages, and label comprehension. The concepts covered in the FAQ section include: the test results, the purpose of the test, limitations of the test, relevance of race and ethnicity on test results, the meaning of the result, other risks factors that contribute to disease, appropriate follow-up procedures, how the results of the test may affect the user's family and children, and links to resources that provide additional information. The questions included in the FAQ section for each of the tests report included the following:

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- What does this test do?
- What does this test not do?
- My report says my result is e2/e2 what does this mean about my late onset Alzheimer's disease risk?
- What does "decreased risk" mean?
- Based on my result, what are some things I could do?
- The report says that estimates about risk are best studied in people of European descent. What if I am not of European descent?
- How could these results affect my family?
- Where can I find references for this information?
- How private is my result?

Each Genetic Health Report has answers to the FAQs that are specific to the variant(s) and disease being reported, where applicable.

d. User Opt-In page:

Due to the nature of late-onset Alzheimer's disease, users are informed on the pre-purchase page, which shows information on the user opt-in section prior to adding the HRA for late-onset Alzheimer's disease to the (purchase) cart as indicated below.

- Are you sure you want to view your Helix Genetic Health Risk App for late-onset Alzheimer's Disease test report?
- By purchasing this product, you are opting-in to receive personal health information for Alzheimer's disease alone. This test will indicate whether you are at an increased, decreased or average risk of developing late-onset Alzheimer's disease. Note that there are currently no genetic testing guidelines or treatment options available for Alzheimer's disease. Talk to a healthcare provider if you have any questions or concerns.

Before the report is generated, users are asked to opt-in for viewing the report as below.

- Are you sure you want to view your Alzheimer's Disease Gene Test report?
- This test will indicate whether you are at an increased, decreased or average risk of developing Alzheimer's disease. Note that there are currently no genetic testing guidelines or treatment options available for Alzheimer's disease. Talk to a healthcare provider if you have any questions or concerns.

i. Yes, I want to view my report
ii. No, I do not want to view my report
iii. Ask me again later

D. Clinical Cut-off:

Not applicable

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E Expected Values/Reference Range

Not applicable

VIII Proposed Labeling:

The labeling supports the finding of substantial equivalence for this device.

IX Conclusion:

The submitted information in this premarket notification is complete and supports a substantial equivalence decision.