(265 days)
The Verigene® CYP2C19 Nucleic Acid Test (CYP2C19 Test), performed using the sample-to-result Verigene System, is a qualitative multiplexed in vitro diagnostic test for the simultaneous detection and identification of an individual's CYP450 2C19 genotype in genomic deoxyribonucleic acid (DNA) obtained from EDTA-anticoagulated whole blood samples. The Verigene CYP2C19 Nucleic Acid Test (CYP2C19 Test) is indicated for use in clinical laboratories upon prescription by the attending physician as an aid to clinicians in determining therapeutic strategy for therapeutics that are metabolized by the CYP450 2C19 gene product, specifically *2, *3, and *17. The Verigene CYP2C19 Nucleic Acid Test (CYP2C19 Test) is not indicated to be used to predict drug response or non-response.
The Verigene® System is comprised of test consumables and shared instrumentation. All Verigene tests are formatted in self-contained test-specific Verigene Test Cartridges which serve to analyze a nucleic acid sample that is presented to them. Nucleic acids are prepared directly from a whole blood specimen using magnetic glass particles and input automatically into a Test Cartridge inside the Verigene Processor SP. Test progress is tracked and directed by the Verigene Reader instrument, which serves as a central control unit for each Verigene System. Genomic DNA is extracted from the white blood cells in a whole blood specimen, fragmented and denatured. This fragmented, single-stranded genomic DNA hybridizes to complementary sequence-specific DNA oligonucleotides, known as capture oligonucleotides, arrayed on the surface of a substrate (glass slide). A second DNA oligonucleotide is then hybridized to the captured genomic DNA that was captured initially. This oligonucleotide is known as a mediator oligonucleotide containing two sequence domain is complementary to the genomic DNA target and a second domain is complementary to a common oligonucleotide attached to a signal generating gold nanoparticle probe. After washing away any DNA not affixed to the captures, the probe is exposed to the captured mediator/target compound where it hybridizes to any captured mediators. Presence of the gold nanoparticle probes at a particular location on the substrate is assessed optically. The Verigene CYP2C19 Nucleic Acid Test is designed to detect and genotype the CYP450 2C19 *2, *3 and *17 alleles. The test report lists the alleles and provides which genotype was detected in the specimen. The CYP2C19 Test algorithm automatically calculates each of the allele results using a preset normalized ratio of the signal of wild type capture locations on the microarray to the mutant capture locations on the microarray.
This submission focuses on the analytical and clinical performance of the Verigene® CYP2C19 Nucleic Acid Test. Here's a breakdown based on your request:
1. Table of Acceptance Criteria and Reported Device Performance
| Acceptance Criteria Category | Acceptance Criteria (Implicit/Explicit) | Reported Device Performance |
|---|---|---|
| Analytical Sensitivity / Limit of Detection (LOD) | Initial call rate > 90% and 100% accuracy vs. BDS for detected genotypes within specified sample volume range. | Initial call rate > 90% and 100% accuracy vs. BDS for all tested genotypes between 800µL - 1200µL sample input volume. |
| Interference | Consistent detection and 100% accuracy vs. BDS in presence of elevated interfering substances. | All genotypes detected consistently with 100% accuracy vs. BDS in the presence of albumin, bilirubin (conjugated and unconjugated), triglycerides, and cholesterol. |
| Specimen Stability | No degradation (e.g., no decrease in extracted DNA concentration/purity, high daily call rate, 100% genotype accuracy) for designated storage conditions. | No signs of degradation, daily call rate > 97%, and 100% genotype accuracy observed for 10 days at refrigerated storage (2 to 8°C). |
| Carry-over / Cross-contamination | 100% genotyping accuracy and no evidence of carry-over/cross-contamination. | 100% genotyping accuracy and no evidence of carry-over/cross-contamination (initial call rates > 93%) during sequential testing of different genotypes. |
| Precision | High percent agreement compared to bi-directional sequencing, with a high initial and final call rate. | Initial Call Rate: 97.7% (375/384). Final Call Rate: 100% (384/384). Agreement vs. BDS: 100% (384/384). |
| Reproducibility | High percent agreement compared to bi-directional sequencing, with a high initial and final call rate across multiple sites. | Initial Call Rate: 96.9% (465/480). Final Call Rate: 99.6% (478/480). Agreement vs. BDS: 99.6% (478/480). |
| Method Comparison | High percent agreement compared to bi-directional sequencing, with high initial and final call rates. | Initial Call Rate: 94.8% (635/670). Final Call Rate: 99.9% (669/670). Agreement vs. BDS: 99.6% (667/670). |
2. Sample Size Used for the Test Set and Data Provenance
The document describes several test sets used for different studies:
- Analytical Sensitivity / LOD: 7 individual whole blood samples, each with a different genotype. Tested in replicates of 40 (total 280 tests). The data provenance is not specified, but it refers to "individual whole blood samples," suggesting clinical samples. The study appears to be prospective in nature, designed specifically for this validation.
- Interference Testing: 5 individual whole blood samples, each with a different genotype. Tested in 30 replicates per specimen for each interfering substance (total 150 tests per substance, with 5 substances and a control, leading to 900 measurements in Table 2). Data provenance is not specified, but it refers to "EDTA-anticoagulated whole blood samples," suggesting clinical samples. This study appears to be prospective.
- Specimen Stability Study: 35 EDTA whole blood samples. Tested once at 5, 10, 12, and 15 day time points (total 140 tests). Data provenance is not specified (e.g., "freshly-collected whole blood samples"). This study appears to be prospective.
- Carry-over / Cross-contamination: Whole blood specimens containing different genotypes. Tested sequentially on ten Verigene instruments, repeated in triplicate (total number of samples not explicitly stated, but includes "*1/*2, followed by *1/*1, then *1/*17, then *1/*1" repeated in triplicate). Data provenance is not specified. This study appears to be prospective.
- Precision Study: 8 unique whole blood specimens. Each tested in duplicate twice daily by two operators over 12 non-consecutive days at one site (48 replicates per specimen, total 384 data points). Data provenance is not specified. This study appears to be prospective.
- Reproducibility Study: The same 8-member panel of specimens as the Precision Study. Tested in duplicate twice daily by two operators over 5 non-consecutive days at three sites (60 replicates per specimen, total 480 data points). Data provenance is not specified. This study appears to be prospective.
- Method Comparison Study: 670 unique human whole blood samples, collected in EDTA. Data provenance is not specified, but the samples are "human whole blood samples," implying clinical origin. The study appears to be prospective, specifically for method comparison.
3. Number of Experts Used to Establish the Ground Truth for the Test Set and Qualifications of Those Experts
The ground truth for all performance studies (Analytical Sensitivity, Interference, Precision, Reproducibility, Method Comparison) was established by bi-directional sequencing (BDS). This is a laboratory method, not reliant on human experts for interpretation in the same way imaging studies might be. Therefore, the concept of "number of experts" and their "qualifications" for ground truth establishment is not applicable here.
4. Adjudication Method for the Test Set
Not applicable for this type of laboratory test. The ground truth (bi-directional sequencing) is considered definitive. When the device produced "No Calls" initially, these samples were re-tested, and if successful, contributed to the "Final Call Rate." In the Reproducibility study, the two final "No Call" results were considered discordant. For the Method Comparison study, one sample with an initial and final no-call on the Verigene test had a BDS result of *1/*2, indicating this was a definite discrepancy.
5. If a Multi Reader Multi Case (MRMC) Comparative Effectiveness Study was done
No, an MRMC comparative effectiveness study was not conducted. This device is a molecular diagnostic test that provides a genotype result, not an imaging device requiring human reader interpretation.
6. If a Standalone (i.e. algorithm only without human-in-the-loop performance) was done
Yes, the performance studies detailed (Analytical Sensitivity, Interference, Stability, Carry-over, Precision, Reproducibility, Method Comparison) represent the standalone performance of the Verigene® CYP2C19 Nucleic Acid Test. The system is described as "sample-to-result" with "automated DNA extraction" and the "Test algorithm automatically calculates each of the allele results." This indicates minimal human intervention in the final result generation once the sample is loaded.
7. The Type of Ground Truth Used
The primary ground truth used for all performance evaluations was bi-directional sequencing (BDS). This is a highly accurate molecular method for determining specific DNA sequences, considered the gold standard for genotyping.
8. The Sample Size for the Training Set
The document does not specify a separate training set or its sample size. This is common for diagnostic tests like this, especially when the underlying technology (genotyping microarray and algorithm) is based on established scientific principles rather than a machine learning model that requires explicit training data in the context of regulatory submissions. The algorithm's parameters are likely "preset" based on scientific design and internal development/optimization rather than a distinct, large-scale training dataset as seen with AI/ML systems.
9. How the Ground Truth for the Training Set was Established
Since no explicit "training set" is described in the context of this 510(k) summary, the method for establishing its ground truth is also not provided. The device's "preset normalized ratio of the signal of wild type capture locations on the microarray to the mutant capture locations on the microarray" implies that the underlying genetic science and expected signal profiles for specific alleles dictate the algorithm's basis, rather than being "trained" on a dataset in the AI/ML sense.
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510(K) Summary
510(k) Number:
K120466: Verigene® CYP2C19 Nucleic Acid Test (CYP2C19)
Summary Preparation Date:
October 31, 2012
Submitted by:
Nanosphere, Inc. 4088 Commercial Avenue Northbrook, IL 60062 Phone: 847-400-9000 Fax: 847-400-9176
Contact:
Mark A. Del Vecchio Vice President, Regulatory Affairs
Proprietary Names:
For the instrument: Verigene® System
For the assay: Verigene® CYP2C19 Nucleic Acid Test (CYP2C19 Test)
Common Names:
For the instrument
Bench-top molecular diagnostics workstation
For the assay:
Cytochrome P450 CYP2C19 Drug Metabolizing Test
NOV 6 2012
Nanosphere, Inc.
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Regulatory Information:
Regulation section:
862.3360 Drug Metabolizing Enzyme Genotyping System 862.2570 Instrumentation for Clinical Multiplex Test Systems
Classification:
Class II
Panel:
Toxicology (91) Chemistry (75)
Product Code(s):
NTI Drug Metabolizing Enzyme Genotyping System NSU Instrumentation for Clinical Multiplex Test Systems Other codes used by predicate devices:
None
Predicate Device(s):
INFINITI®CYP2C19 Assay (K101683)
Indications for Use:
The Verigene® CYP2C19 Nucleic Acid Test (CYP2C19 Test), performed using the sample-to-result Verigene System, is a qualitative multiplexed in vitro diagnostic test for the simultaneous detection and identification of an individual's CYP450 2C19 genotype in genomic deoxyribonucleic acid (DNA) obtained from EDTA-anticoagulated whole blood samples. The Verigene CYP2C19 Nucleic Acid Test (CYP2C19 Test) is indicated for use in clinical laboratories upon prescription by the attending physician as an aid to clinicians in determining therapeutic strategy for therapeutics that are metabolized by the CYP450 2C19 gene product, specifically *2, *3, and *17. The Verigene CYP2C19 Nucleic Acid Test (CYP2C19 Test) is not indicated to be used to predict drug response or non-response.
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Technological Characteristics:
The Verigene® System is comprised of test consumables and shared instrumentation. All Verigene tests are formatted in self-contained test-specific Verigene Test Cartridges which serve to analyze a nucleic acid sample that is presented to them. Nucleic acids are prepared directly from a whole blood specimen using magnetic glass particles and input automatically into a Test Cartridge inside the Verigene Processor SP. Test progress is tracked and directed by the Verigene Reader instrument, which serves as a central control unit for each Verigene System.
Genomic DNA is extracted from the white blood cells in a whole blood specimen, fragmented and denatured. This fragmented, single-stranded genomic DNA hybridizes to complementary sequencespecific DNA oligonucleotides, known as capture oligonucleotides, arrayed on the surface of a substrate (glass slide). A second DNA oligonucleotide is then hybridized to the captured genomic DNA that was captured initially. This oligonucleotide is known as a mediator oligonucleotide containing two sequence domain is complementary to the genomic DNA target and a second domain is complementary to a common oligonucleotide attached to a signal generating gold nanoparticle probe. After washing away any DNA not affixed to the captures, the probe is exposed to the captured mediator/target compound where it hybridizes to any captured mediators. Presence of the gold nanoparticle probes at a particular location on the substrate is assessed optically.
The Verigene CYP2C19 Nucleic Acid Test is designed to detect and genotype the CYP450 2C19 *2, 1 *3 and * 17 alleles. The test report lists the alleles and provides which genotype was detected in the specimen. The CYP2C19 Test algorithm automatically calculates each of the allele results using a preset normalized ratio of the signal of wild type capture locations on the microarray to the mutant capture locations on the microarray.
Substantial Equivalence:
The Verigene® CYP2C19 Nucleic Acid Test (CYP2C19) has the same Intended Use and Indications for Use, similar technological and performance characteristics, and similar principles of operation as its predicate device, the INFINITI®CYP2C19 Assay (K101683), that the Food and Drug Administration ("FDA") has cleared for the identification of a patient's CYP450 2C19 genotype in genomic deoxyribonucleic acid (DNA) obtained from EDTA-anticoagulated whole blood samples (see Table 1). The INFINITI CYP2C19 Assay is a qualitative in vitro assay for use in clinical laboratories upon prescription by the attending physician and is indicated for use as an aid to clinicians in determining therapeutic strategy for therapeutics that are metabolized by CYP450 enzymes known to be affected by mutations in the *2, *3 and *17 alleles of the CYP2C19 gene. The INFINITI CYP2C19 Assay (K101683) is not indicated to be used to predict drug response or nonresponse. The minor differences between the CYP2C19 test and its predicate device raise no new issues of safety or effectiveness. The analytical and clinical performance data demonstrate that the CYP2C19 test is as safe and effective as the predicate device. Therefore, the Verigene CYP2C19 Nucleic Acid Test is substantially equivalent to the INFINITI CYP2C19 Assay.
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| Items | Device | Predicate |
|---|---|---|
| Verigene CYP2C19 Test | INFINITI CYP2C19 Assay | |
| 510(k)# | K120466 | K101683 |
| Regulations | 862.3360 and 862.2570 | Same |
| Product Code | NTI, NSU | Same |
| Device Class | II | Same |
| Intended Use | Identification of an individual's CYP450 2C19genotype | Same |
| Indications for Use | To aid in determining therapeutic strategy fortherapeutics metabolized by the CYP450 2C19 geneproduct, specifically *2, *3, and *17 alleles. | Same |
| Warnings andPrecautions | For use in clinical laboratories upon prescription bythe attending physician | Same |
| Contraindication (s) | Assay is not intended to be used to predict drugresponse or non-response. | Same |
| Test Cartridge | Disposable single-use, multi- chambered fluidiccartridge. | Same |
| Sample Type | EDTA-anticoagulated whole blood | Same |
| Sample Prep | On-board, automated DNA extraction | Same |
| Quality control | Internal procedural/instrument quality controls;Internal Negative Control, Sample processingcontrol, external positive and negative assay controls | Same |
| Interpretation of Results | Diagnostic Software/Decision Algorithm | Same |
| Target Mutations | *2, *3, and *17 genotype of CYP 2C19 | Same |
| Type of Test | Genotyping microarray | Same |
| Detection Method | Gold/Ag nanoparticle probe detection of DNA oncomplementary oligo- microarray | Measures fluorescent signals oflabeled DNA target hybridizedto the microarray. |
| DNA Extraction Method | Automated DNA extraction with "blood sample toresult system" | Manual, "off-line" DNAextraction |
| Sample Carry-over | None observed | Same |
| Interference | None observed for bilirubin, triglyceride, cholesterol,and albumin | Same |
| Precision | ||
| Initial Call Rate | 97.7% | Not Done |
| Final Call Rate | 100% | |
| Agreement vs BDS | 100% | |
| Reproducibility | ||
| Initial Call Rate | 96.9% | 96.5% |
| Final Call Rate | 99.6% | 100% |
| Agreement vs BDS | 99.6% | 99.8% |
| Method Comparison | ||
| Initial Call Rate | 94.8% | 98.1% |
| Final Call Rate | 99.9% | 100% |
| Agreement vs BDS | 99.6% | 100% |
Table 1: Comparative Characteristics: CYP2C19 Test and Predicate Device
:
.
Nanosphere, Inc.
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- . . . . . . .
Performance Data Summary - Analytical Testing
Analytical Sensitivity / Limit of Detection (LOD)
To evaluate the upper and lower limits of the assay sample volume between which the CYP2C19 Test performs properly, seven individual whole blood samples, each containing a different genotype, * 1/* 1, * 1/* 2, * 2/* 17, * 1/* 3, * 1/* 17, and * 17/* 17, were tested in replicates of 40 at sample input volumes of 800uL, 900uL, 1000uL, 1100uL, and 1200uL. The study demonstrated that the CYP2C19 Test detected each genotype consistently (initial call rate >90%) and accurately (100% vs. BDS) for all genotypes tested between the specimen volume range of 1000 + 200μL (800μL - 1200μL). These results establish 1.0 + 0.1mL of whole blood as the sample volume tolerance for the assay. The ability of the CYP2C19 test to consistently detect the genotype of a specimen will be negatively impacted at whole blood sample input volumes of less than 750pL.
Interference Testing
The potential inhibitory effects of substances that may be present in whole blood were evaluated at biologically relevant but elevated concentrations, by individually adding albumin (@6000 mg/dL), bilirubin (conjugated and unconjugated @20 mg/dL), triglycerides (@3000 mg/dL), and cholesterol (@500 mg/dL) directly into EDTA-anticoagulated whole blood samples, each individually containing genotypes * 1/* 1, * 1/* 2, * 2/* 17, * 1/* 17, and * 17/* 17, A total of 30 replicates per specimen containing each interfering substance were tested and the results compared to unspiked control samples. In the presence of these substances, each genotype was detected consistently with 100% accuracy vs. BDS, demonstrating that these substances do not interfere with the CYP2C19 Test.
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| Initial Results | Final Results | |||||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|
| Genotype | No. Tested | Interfering Substance | Reps per Interferent | No. of Genotype Calls Correct | No. of Genotype Calls Incorrect | % Calls Agreement | 95% One-sided CI LL | No. of Genotype Calls Correct | No. of Genotype Calls Incorrect | % Calls Agreement | 95% One-sided CI LL | |
| *1/*1 | 1 | Control | 30 | 29 | 0 | 97 | 85.1 | 30 | 0 | 100 | 90.5 | |
| Cholesterol | 30 | 28 | 0 | 93 | 80.5 | 30 | 0 | 100 | 90.5 | |||
| Triglycerides | 30 | 29 | 0 | 97 | 85.1 | 30 | 0 | 100 | 90.5 | |||
| Unconj. Bilirubin | 30 | 29 | 0 | 97 | 85.1 | 30 | 0 | 100 | 90.5 | |||
| Conj. Bilirubin | 30 | 29 | 0 | 97 | 85.1 | 30 | 0 | 100 | 90.5 | |||
| Albumin | 30 | 28 | 0 | 93 | 80.5 | 30 | 0 | 100 | 90.5 | |||
| TOTAL | 180 | 172 | 0 | 96 | 92.1 | 180 | 0 | 100 | 98.4 | |||
| *1/*2 | 1 | Control | 30 | 30 | 0 | 100 | 90.5 | 30 | 0 | 100 | 90.5 | |
| Cholesterol | 30 | 29 | 0 | 97 | 85.1 | 30 | 0 | 100 | 90.5 | |||
| Triglycerides | 30 | 30 | 0 | 100 | 90.5 | 30 | 0 | 100 | 90.5 | |||
| Unconj. Bilirubin | 30 | 29 | 0 | 97 | 85.1 | 30 | 0 | 100 | 90.5 | |||
| Conj. Bilirubin | 30 | 30 | 0 | 100 | 90.5 | 30 | 0 | 100 | 90.5 | |||
| Albumin | 30 | 30 | 0 | 100 | 90.5 | 30 | 0 | 100 | 90.5 | |||
| TOTAL | 180 | 178 | 0 | 99 | 96.5 | 180 | 0 | 100 | 98.4 | |||
| *1/*3 | 1 | Control | 30 | 30 | 0 | 100 | 90.5 | 30 | 0 | 100 | 90.5 | |
| Cholesterol | 30 | 30 | 0 | 100 | 90.5 | 30 | 0 | 100 | 90.5 | |||
| Triglycerides | 30 | 28 | 0 | 93 | 80.5 | 30 | 0 | 100 | 90.5 | |||
| Unconj. Bilirubin | 30 | 30 | 0 | 100 | 90.5 | 30 | 0 | 100 | 90.5 | |||
| Conj. Bilirubin | 30 | 30 | 0 | 100 | 90.5 | 30 | 0 | 100 | 90.5 | |||
| Albumin | 30 | 27 | 0 | 90 | 76.1 | 29 | 0 | 97 | 85.1 | |||
| TOTAL | 180 | 175 | 0 | 97 | 94.3 | 179 | 0 | 99 | 97.4 | |||
| *1/*17 | 1 | Control | 30 | 30 | 0 | 100 | 90.5 | 30 | 0 | 100 | 90.5 | |
| Cholesterol | 30 | 30 | 0 | 100 | 90.5 | 30 | 0 | 100 | 90.5 | |||
| Triglycerides | 30 | 27 | 0 | 90 | 76.1 | 30 | 0 | 100 | 90.5 | |||
| Unconj. Bilirubin | 30 | 28 | 0 | 93 | 80.5 | 30 | 0 | 100 | 90.5 | |||
| Conj. Bilirubin | 30 | 26 | 0 | 87 | 72.0 | 30 | 0 | 100 | 90.5 | |||
| Albumin | 30 | 29 | 0 | 97 | 85.1 | 30 | 0 | 100 | 90.5 | |||
| TOTAL | 180 | 170 | 0 | 94 | 90.8 | 180 | 0 | 100 | 98.4 | |||
| *2/*2 | 1 | Control | 30 | 30 | 0 | 100 | 90.5 | 30 | 0 | 100 | 90.5 | |
| Cholesterol | 30 | 29 | 0 | 97 | 85.1 | 30 | 0 | 100 | 90.5 | |||
| Triglycerides | 30 | 30 | 0 | 100 | 90.5 | 30 | 0 | 100 | 90.5 | |||
| Unconj. Bilirubin | 30 | 30 | 0 | 100 | 90.5 | 30 | 0 | 100 | 90.5 | |||
| Conj. Bilirubin | 30 | 30 | 0 | 100 | 90.5 | 30 | 0 | 100 | 90.5 | |||
| Albumin | 30 | 30 | 0 | 100 | 90.5 | 30 | 0 | 100 | 90.5 | |||
| TOTAL | 180 | 179 | 0 | 99 | 97.4 | 180 | 0 | 100 | 98.4 | |||
| *2/*17 | 1 | Control | 30 | 27 | 0 | 90 | 76.1 | 30 | 0 | 100 | 90.5 | |
| Cholesterol | 30 | 28 | 0 | 93 | 80.5 | 30 | 0 | 100 | 90.5 | |||
| Triglycerides | 30 | 27 | 0 | 90 | 76.1 | 30 | 0 | 100 | 90.5 | |||
| Unconj. Bilirubin | 30 | 29 | 0 | 97 | 85.1 | 30 | 0 | 100 | 90.5 | |||
| Conj. Bilirubin | 30 | 28 | 0 | 93 | 80.5 | 29 | 0 | 97 | 85.1 | |||
| Albumin | 30 | 27 | 0 | 90 | 76.1 | 30 | 0 | 100 | 90.5 | |||
| TOTAL | 180 | 166 | 0 | 92 | 88.1 | 179 | 0 | 99 | 97.4 | |||
| *17/*17 | 1 | Control | 30 | 29 | 0 | 97 | 85.1 | 29 | 0 | 97 | 85.1 | |
| Cholesterol | 30 | 28 | 0 | 93 | 80.5 | 29 | 0 | 97 | 85.1 | |||
| Triglycerides | 30 | 29 | 0 | 97 | 85.1 | 30 | 0 | 100 | 90.5 | |||
| Unconj. Bilirubin | 30 | 30 | 0 | 100 | 90.5 | 30 | 0 | 100 | 90.5 | |||
| Conj. Bilirubin | 30 | 30 | 0 | 100 | 90.5 | 30 | 0 | 100 | 90.5 | |||
| Albumin | 30 | 30 | 0 | 100 | 90.5 | 30 | 0 | 100 | 90.5 | |||
| TOTAL | 180 | 176 | 0 | 98 | 95.0 | 178 | 0 | 99 | 96.5 |
Table 2: Interfering Substance Testing - Initial and Final Call Rates by Genotype
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1 / 2 / 2 / 2 / 2 / 2 / 2 / 2 / 2 / 2 / 2 / 2 / 2 / 2 / 2 / 2 / 2 / 2 / 2 / 2 / 2 / 2 / 2 / 2 / 2 / 2 / 2 / 2 / 2 / 2 / 2 / 2 / 2 / 2 / 2 / 2 / 2 / 2 / 2 / 2 / 2 / 2 / 2 / 2
Specimen Stability Study
The stability of freshly-collected whole blood samples, stored refrigerated for up to 15 days following collection, was evaluated by testing with the CYP2C19 Test. Aliquots of thirty-five (35) EDTA whole blood samples, containing individually genotypes * 1/* 1 (n=4), *2/*2 (n=1), *1/*17 (n=11), and *17/*17 (n=3), were tested once at 5, 10, 12, and 15 day time points with the CYP2C19 Test and the extracted DNA concentration and purity measured for each sample. No signs of degradation (e.g., no decrease in extracted DNA concentration and/or purity, observed daily call rate for all 35 samples tested >97%, and genotype accuracy of 100%) were observed for any sample during the study, establishing 10 days at refrigerated storage (2 to 8℃) as the whole blood specimen stability claim for the CYP2C19 Test.
Carry-over / Cross-contamination
In order to demonstrate that the CYP2C19 Test, when performed on the Verigene System, is not susceptible to sample carry-over or cross-contamination, testing of whole blood specimens containing different genotypes was performed sequentially on ten Verigene instruments (e.g.;*1/*2, followed by *1/*1, then *1/*17, then *1/*1); this testing was repeated in triplicate. Genotyping accuracy was 100% and no evidence of carryover and/or cross-contamination was observed (initial call rates >93% for each of the three genotypes tested.)
Precision
Precision of the CYP2C19 Test was evaluated by testing eight unique whole blood specimens (containing genotypes 1/ 1, *1/*2, *2/*2, 2/ 17, *1/3 (2 tested), 1/ 17, and * 17/ 17) in duplicate twice daily by two operators for twelve non-consecutive days at one testing site. This testing regimen generated a total of forty-eight (48) replicates per specimen and an overall total of 384 data points. Of the 384 samples tested, the percent agreement for all panel members as compared to bi-directional sequencing was 100% (384/384). There were nine (9) "No Calls" in the study for an initial call rate of 97.7% (375/384). All nine of these samples were repeat tested successfully for a final call rate of 100% (384/384).
| Genotype | No. Tested | Reps per Sample | Initial Results | Final Results | ||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|
| No. of Genotype CallsCorrect | No. of Genotype CallsIncorrect | No Calls | Agreement | 95% One-sided CILL | No. of Genotype CallsCorrect | No. of Genotype CallsIncorrect | No Calls | Agreement | 95% One-sided CILL | |||
| *1/*1 | 1 | 48 | 47 | 0 | 1 | 97.9 | 90.5 | 48 | 0 | 0 | 100 | 94.0 |
| *1/*2 | 1 | 48 | 47 | 0 | 1 | 97.9 | 90.5 | 48 | 0 | 0 | 100 | 94.0 |
| *1/*3 | 1 | 48 | 46 | 0 | 2 | 95.8 | 87.5 | 48 | 0 | 0 | 100 | 94.0 |
| *1/*3 | 1 | 48 | 46 | 0 | 2 | 95.8 | 87.5 | 48 | 0 | 0 | 100 | 94.0 |
| *1/*17 | 1 | 48 | 47 | 0 | 1 | 97.9 | 90.5 | 48 | 0 | 0 | 100 | 94.0 |
| *2/*2 | 1 | 48 | 48 | 0 | 0 | 100 | 94.0 | 48 | 0 | 0 | 100 | 94.0 |
| *2/*17 | 1 | 48 | 47 | 0 | 1 | 97.9 | 90.5 | 48 | 0 | 0 | 100 | 94.0 |
| *17/*17 | 1 | 48 | 47 | 0 | 1 | 97.9 | 90.5 | 48 | 0 | 0 | 100 | 94.0 |
| ALL | 8 | 384 | 375 | 0 | 9 | 97.7 | 96.0 | 384 | 0 | 0 | 100 | 99.2 |
Table 3: CYP2C19 Test Precision Study Results
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Performance Data Summary - Clinical Testing
Reproducibility
Reproducibility of the CYP2C19 Test was evaluated at three sites (two external and one internal) using the same eight-member panel as for the Precision Study. These eight specimens were tested in duplicate twice daily by two operators for five (5) non-consecutive days. This testing regimen generated a total of sixty (60) replicates (20 replicates/site) per specimen for an overall total of 480 data points. A summary of the Reproducibility Study results stratified by investigational site is provided in Table 7. There were fifteen (15) initial "No Calls" in the study for an initial call rate of 96.9% (465/480). Thirteen (13) of these samples were repeat tested successfully for a final call rate of 99.6% (478/480). Considering the two final "No Call" results as discordant results, the percent agreement for all panel members as compared to bi-directional sequencing was 99.6% (478/480).
| Genotype | No. Tested | Reps per Sample | Initial Results | Final Results | ||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|
| No. of Genotype CallsCorrect | No. of Genotype CallsIncorrect | No. Calls | Agreement | 95% One-sided CI LL | No. of Genotype CallsCorrect | No. of Genotype CallsIncorrect | No. Calls | Agreement | 95% One-sided CI LL | |||
| *1/*1 | 1 | 60 | 59 | 0 | 1 | 98.3 | 92.3 | 60 | 0 | 0 | 100 | 95.1 |
| *1/*2 | 1 | 60 | 59 | 0 | 1 | 98.3 | 92.3 | 60 | 0 | 0 | 100 | 95.1 |
| *1/*3 | 1 | 60 | 56 | 0 | 4 | 93.3 | 85.4 | 60 | 0 | 0 | 100 | 95.1 |
| *1/*3 | 1 | 60 | 57 | 0 | 3 | 95.0 | 87.6 | 59 | 0 | 1 | 98.3 | 92.3 |
| *1/*17 | 1 | 60 | 59 | 0 | 1 | 98.3 | 92.3 | 60 | 0 | 0 | 100 | 95.1 |
| *2/*2 | 1 | 60 | 59 | 0 | 1 | 98.3 | 92.3 | 60 | 0 | 0 | 100 | 95.1 |
| *2/*17 | 1 | 60 | 57 | 0 | 3 | 95.0 | 87.6 | 59 | 0 | 1 | 98.3 | 92.3 |
| *17/*17 | 1 | 60 | 59 | 0 | 1 | 98.3 | 92.3 | 60 | 0 | 0 | 100 | 95.1 |
| ALL | 8 | 480 | 465 | 0 | 15 | 96.9 | 95.2 | 478 | 0 | 2 | 99.6 | 98.7 |
Table 4: CYP2C19 Test Reproducibility Study Results
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Method Comparison Study
A total of 670 unique human whole blood samples, collected in EDTA, were tested at three sites using the Verigene CYP2C19 Nucleic Acid Test. Table 5 provides agreement with bi-directional sequencing and the number of correct calls observed in the study. Table 6 shows the initial and final call rates at each of the sites and the combined statistics for each, in addition to agreement with bi-directional sequencing. There were 35 initial no calls of which 34 were successfully repeated. Therefore the initial call rate was 94.8% (635/670) and the final call rate was 99.9% (669/670).
| Genotype (a) | No.Tested | Reps perSample | No. CorrectGenotypeCalls | No.IncorrectCalls | No Calls | Agreement | 95% Two-sidedConfidenceInterval LowerLimit |
|---|---|---|---|---|---|---|---|
| *1/*1 (b) | 260 | 1 | 260 | 0 | 0 | 100% | 98.6% |
| *1/*2 | 177 | 1 | 176 | 0 | 1(c) | 99.4% | 96.9% |
| *1/*3 | 24 | 1 | 23 | 1 | 0 | 95.8% | 78.9% |
| *1/*17 | 114 | 1 | 113 | 1 | 0 | 99.1% | 95.2% |
| *2/*2 | 32 | 1 | 32 | 0 | 0 | 100% | 89.1% |
| *2/*3 | 13 | 1 | 13 | 0 | 0 | 100% | 75.3% |
| *2/*17 | 30 | 1 | 30 | 0 | 0 | 100% | 88.4% |
| *3/*17 | 1 | 1 | 1 | 0 | 0 | 100% | 2.5% |
| *3/*3 | 1 | 1 | 1 | 0 | 0 | 100% | 2.5% |
| *17/*17 | 18 | 1 | 18 | 0 | 0 | 100% | 81.5% |
| Total | 670 | 1 | 667 | 2(c) | 1 | 99.6% | 98.7% |
Table 5: Method Comparison Results: Agreement with bi-directional sequencing (n=670)
a) Genotype determined by bi-directional sequencing
b) *1/*1 samples are inferred if they are wild-type for *2, *3 and *17
c) CYP2C19 result initial and final no-call; BDS result *1/*2.
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| Call Rate Per Genotype(95% Two-Sided Confidence Interval) | Agreement withReference Method | ||||||||
|---|---|---|---|---|---|---|---|---|---|
| Site 1(a) | Site 2(b) | Site 3(c) | All Sites | Bi-DirectionalSequencing Result | |||||
| n= | 281 | 264 | 125 | 670 | |||||
| Genotype | Initial | Final | Initial | Final | Initial | Final | Initial | Final | |
| *1/*1 | 100%113/113(96.8-100) | 100%113/113(96.8-100) | 94.9%92/97(88.4-98.3) | 100%97/97(96.3-100) | 92.0%46/50(80.8-97.8) | 100%50/50(92.9-100) | 96.5%251/260(93.5-98.4) | 100%260/260(98.6-100) | 100%260/260(98.6-100) |
| *1/*2 | 95.9%70/73(88.5-99.1) | 98.6%72/73(92.6-99.9) | 91.3%63/69(82.0-96.7) | 100%69/69(94.8-100) | 94.4%34/36(91.3-99.3) | 100%36/36(90.3-100) | 93.8%167/178(89.2-96.9) | 99.4%177/178(96.9-99.9) | 99.4%176/177(d)(96.9-99.9) |
| *1/*3 | 100%8/8(63.1-100) | 100%8/8(63.1-100) | 88.9%8/9(51.6-99.7) | 100%9/9(66.4-99.9) | 100%6/6(54.1-100) | 100%6/6(54.1-100) | 95.7%22/23(78.1-99.9) | 100%23/23(85.2-100) | 95.8%23/24(78.9-99.9) |
| *1/*17 | 96.2%51/53(87.0-99.5) | 100%53/53(93.3-100) | 93.0%40/43(80.9-98.5) | 100%43/43(91.8-100) | 100%17/17(90.5-100) | 100%17/17(90.5-100) | 95.6%108/113(90.0-98.6) | 100%113/113(96.8-100) | 99.1%113/114(95.2-99.9) |
| *2/*2 | 100%11/11(71.5-100) | 100%11/11(71.5-100) | 100%16/16(79.4-100) | 100%16/16(79.4-100) | 100%5/5(47.8-100) | 100%5/5(47.8-100) | 100%32/32(89.1-100) | 100%32/32(89.1-100) | 100%32/32(89.1-100) |
| *2/*3 | 100%3/3(29.2-100) | 100%3/3(29.2-100) | 100%5/5(47.8-100) | 100%5/5(47.8-100) | 80.0%4/5(28.4-99.5) | 100%5/5(47.8-100) | 92.3%12/13(64.0-99.8) | 100%13/13(75.3-100) | 100%13/13(75.3-100) |
| *2/*17 | 83.3%10/12(51.6-97.9) | 100%12/12(73.5-100) | 85.7%12/14(57.2-98.2) | 100%14/14(76.8-100) | 75.0%3/4(19.4-99.4) | 100%4/4(39.8-100) | 83.3%25/30(65.3-94.4) | 100%30/30(88.4-100) | 100%30/30(88.4-100) |
| *3/*17 | 0%0/1(0-97.5) | 100%1/1(2.5-100) | 0%0/1(0-97.5) | 100%1/1(2.5-100) | 100%1/1(2.5-100) | ||||
| *3/*3 | 100%1/1(2.5-100) | 100%1/1(2.5-100) | 100%1/1(2.5-100) | 100%1/1(2.5-100) | 100%2/2(15.8-100) | 100%2/2(15.8-100) | 100.0%1/1(c)(2.5-100) | ||
| *17/*17 | 100%6/6(54.1-100) | 100%6/6(54.1-100) | 81.8%9/11(48.2-97.7) | 100%11/11(71.5-100) | 100%1/1(2.5-100) | 100%1/1(2.5-100) | 88.9%16/18(65.3-98.6) | 100%18/18(81.5-100) | 100%18/18(81.5-100) |
| Total BySite | 97.2%273/281(94.5-98.8) | 99.6%280/281(98.0-99.9) | 92.8%245/264(89.0-95.6) | 100%264/264(98.6-100) | 93.6%117/125(87.8-97.2) | 100%125/125(97.1-100) | 94.8%635/670(92.8-96.3) | 99.9%669/670(99.2-100) | 99.6%667/670(98.7-99.9) |
| Table 6: Method Comparison Results, Initial and Final Call Rates and Accuracy by Site and Genotype | |||
|---|---|---|---|
| ---------------------------------------------------------------------------------------------------- | -- | -- | -- |
(a) 8 samples - "no-call" repeat testing of two * 1/* 17, three * 1/*2 two 2/ 17, and one 3/ 17 genotypes.
19 samples - "no-call" repeat testing of five * 1/* 2, one * 1/* 3, three * 1/* 17, two 2/ 17, and two (b) *17/*17genotypes.
8 samples - "no-call" repeat testing of four * 1/* 1, two * 1/* 2, one 2/ 17 genotypes. (c)
(d) CYP2C19 result initial and final no-call; BDS result *1/*2.
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Image /page/10/Picture/0 description: The image shows the logo for the U.S. Department of Health & Human Services. The logo features the department's name in a circular arrangement around a stylized symbol. The symbol resembles a stylized caduceus, with a central staff and a winding, ribbon-like design.
DEPARTMENT OF HEALTH & HUMAN SERVICES
Public Health Service
Food and Drug Administration 10903 New Hampshire Avenue Document Control Center - WO66-G609 Silver Spring, MD 20993-002
Letter date: November 6, 2012
Nanosphere, Inc. c/o Mark A. Del Vecchio 4088 Commercial Avenue Northbrook, IL 60062
Re: K120466
Trade Name: Verigene® CYP2C19 Nucleic Acid Test (CYP2C19) Regulation Number: 21 CFR §862.3360 Regulation Name: Drug Metabolizing enzyme genotyping system Regulatory Class: Class II Product Codes: NTI, NSU Dated: September 20, 2012 Received: September 21, 2012
Dear Mr. Del Vecchio:
We have reviewed your Section 510(k) premarket notification of intent to market the device referenced above and have determined the device is substantially equivalent (for the indications for use stated in the enclosure) to legally marketed predicate devices marketed in interstate commerce prior to May 28, 1976, the enactment date of the Medical Device Amendments, or to devices that have been reclassified in accordance with the provisions of the Federal Food, Drug, and Cosmetic Act (Act) that do not require approval of a premarket approval application (PMA). You may, therefore, market the device, subject to the general controls provisions of the Act. The general controls provisions of the Act include requirements for annual registration, listing of devices, good manufacturing practice, labeling, and prohibitions against misbranding and adulteration.
If your device is classified (see above) into either class II (Special Controls) or class III (PMA), it may be subject to such additional controls. Existing major regulations affecting your device can be found in Title 21, Code of Federal Regulations (CFR), Parts 800 to 895. In addition, FDA may publish further announcements concerning your device in the Federal Register.
Please be advised that FDA's issuance of a substantial equivalence determination does not mean that FDA has made a determination that your device complies with other requirements of the Act or any Federal statutes and regulations administered by other Federal agencies. You must comply with all the Act's requirements, including, but not limited to: registration and listing (21 CFR Part 807); labeling (21 CFR Parts 801 and 809); medical device reporting (reporting of medical device-related adverse events) (21 CFR 803); and good manufacturing practice requirements as set forth in the quality systems (QS) regulation (21 CFR Part 820); and if
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Page 2 – Mark Del Vecchio
applicable, the electronic product radiation control provisions (Sections 531-542 of the Act); 21 CFR 1000-1050.
If you desire specific advice for your device on our labeling regulation (21 CFR Part 801), please contact the Office of In Vitro Diagnostic Device Evaluation and Safety at (301) 796-5450. Also, please note the regulation entitled, "Misbranding by reference to premarket notification" (21 CFR Part 807.97). For questions regarding postmarket surveillance, please contact CDRH's Office of Surveillance and Biometric's (OSB's) Division of Postmarket Surveillance at (301) 796-5760. For questions regarding the reporting of adverse events under the MDR regulation (21 CFR Part 803), please go to
http://www.fda.gov/MedicalDevices/Safety/ReportaProblem/default.htm for the CDRH's Office of Surveillance and Biometrics/Division of Postmarket Surveillance.
You may obtain other general information on your responsibilities under the Act from the Division of Small Manufacturers, International and Consumer Assistance at its toll-free number (800) 638-2041 or ( 301 ) 796-5680 or at its Internet address http://www.fda.gov/MedicalDevices/ResourcesforYou/Industry/default.htm.
Sincerely yours,
Carol C. Benson of
Digitally signed by Carol C. Benson DN: c=US. o=U.S. Government, ou=HHS DA, ou=People, cn=Carol C. Benso 0.9.2342.19200300.100.1.1=1300086490 Date: 2012.11.06 14:51:34 -05'00'
for
Courtney H. Lias, Ph.D. Director Division of Chemistry and Toxicology Devices Office of In Vitro Diagnostics and Radiological Health Center for Devices and Radiological Health
Enclosure
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Indications for Use Statement
K120466 510(k) Number (if known):
Device Name: Verigene® CYP2C19 Nucleic Acid Test (CYP2C19 Test)
Indications for Use:
The Verigene® CYP2C19 Nucleic Acid Test (CYP2C19 Test), performed using the sample-toresult Verigene System, is a qualitative multiplexed in vitro diagnostic test for the simultaneous detection and identification of an individual's CYP450 2C19 genotype in genomic deoxyribonucleic acid (DNA) obtained from EDTA-anticoagulated whole blood samples. The Verigene CYP2C19 Nucleic Acid Test (CYP2C19 Test) is indicated for use in clinical laboratories upon prescription by the attending physician as an aid to clinicians in determining therapeutic strategy for therapeutics that are metabolized by the CYP450 2C19 gene product, specifically *2, *3, and *17. The Verigene CYP2C19 Nucleic Acid Test (CYP2C19 Test) is not indicated to be used to predict drug response or non-response.
Prescription Use V (Part 21 CFR 801 Subpart D)
and/or.
Over-The-Counter Use (21 CFR 801 Subpart C)
(PLEASE DO NOT WRITE BELOW THIS LINE-CONTINUE ON ANOTHER PAGE IF NEEDED)
Concurrence of CDRH, Office of In Vitro Diagnostic Device Evaluation and Safety (OIR)
Qute Clu
Division Sign-Off Office of In Vitro Diagnostic Device Evaluation and Safety
× 120466 510(k)
.......
§ 862.3360 Drug metabolizing enzyme genotyping system.
(a)
Identification. A drug metabolizing enzyme genotyping system is a device intended for use in testing deoxyribonucleic acid (DNA) extracted from clinical samples to identify the presence or absence of human genotypic markers encoding a drug metabolizing enzyme. This device is used as an aid in determining treatment choice and individualizing treatment dose for therapeutics that are metabolized primarily by the specific enzyme about which the system provides genotypic information.(b)
Classification. Class II (special controls). The special control is FDA's guidance document entitled “Class II Special Controls Guidance Document: Drug Metabolizing Enzyme Genotyping Test System.” See § 862.1(d) for the availability of this guidance document.