(267 days)
The PGDx elio™ tissue complete assay is a qualitative in vitro diagnostic device that uses targeted next generation sequencing of DNA isolated from formalin-fixed, paraffin-embedded tumor tissue from patients with solid malignant neoplasms to detect tumor gene alterations in a broad multi-gene panel.
PGDx elio tissue complete is intended to provide tumor mutation profiling information on somatic alterations (SNVs, small insertions and deletions, one amplification and four translocations), microsatellite instability (MSI) and tumor mutation burden (TMB) for use by qualified healthcare professionals in accordance with professional guidelines in oncology for previously diagnosed cancer patients, and is not conclusive or prescriptive for labeled use of any specific therapeutic product.
PGDx elio tissue complete is an in vitro diagnostic assay that uses NGS to detect turnor gene alterations in genomic DNA isolated from formalin-fixed, paraffin-embedded (FFPE) tumor tissue from a variety of tumor types, using a targeted panel (505 genes). The assay takes less than 7 days from DNA to report and provides information on single nucleotide variants (SNVs) in a range of GC content and genomic contexts, insertion/ deletions (indels), 1 amplification as well as 4 translocations. It also identifies microsatellite instability based on select mononucleotide tracts and signatures of sequence mutations. The PGDx elio tissue complete assay utilizes a ~1.3 Mb region of interest (ROI) to calculate tumor mutation burden (TMB).
The provided text describes the analytical validation studies for the PGDx elio tissue complete assay, a qualitative in vitro diagnostic device for tumor profiling.
Here's an analysis of the acceptance criteria and the study that proves the device meets them:
1. Table of Acceptance Criteria & Reported Device Performance
The acceptance criteria are generally established as target performance metrics (e.g., call rates, concordance rates, %CV, false positive rates). The reported device performance is the outcome of the analytical studies.
| Performance Metric Category | Specific Metric (Acceptance Criteria Implicitly Defined by Study Design/Results) | Reported Device Performance |
|---|---|---|
| Specificity | False Positive Rate (SNVs and Indels) | 99.9% |
| TMB Mean Absolute Percent Error (MAPE) vs. 100 ng | 1.8% to 11.8% | |
| Contamination Control | Absence of false positives in negative samples from carryover/cross-contamination | No positive variant results observed in known negative samples. |
| Exogenous Interference | Concordance (PPA) with interfering substances vs. baseline | > 97.2% |
| Concordance (NPA) with interfering substances vs. baseline | > 99.9% | |
| TMB MAPE with interfering substances | 0% to 6.0% | |
| Overall Sample Acceptance Rate | First Pass Rate | 81.8% (2352/2874) |
| Overall Pass Rate (allowing single repeat) | 92.9% (2671/2874) | |
| Accuracy (Concordance to Orthogonal Methods) | PPA/NPA for various variant types (SNVs, indels, amplifications, translocations) | See Table 1.10 for detailed SNV/Indel PPAs (range 80.8% - 100%) and NPAs (all 99.9% or 100%) |
| ERBB2 Amplifications: PPA 75.0% (all cases), 87.0% (excluding borderline); NPA 96.7% (all cases), 95.9% (excluding borderline) | ||
| ALK Translocations: PPA 92.9%; NPA 98.2% | ||
| RET Translocations: PPA 55.6%; NPA 100% | ||
| TMB: Spearman correlation coefficient 0.903 vs WES | ||
| MSI: Overall PPA 98.8% (excluding failures), 94.0% (accounting for failures); NPA 99.3% (excluding failures), 77.6% (accounting for failures) | ||
| Interlaboratory Reproducibility | First Pass Rate | 90.3% (455/504) |
| Overall Pass Rate | 98.2% (495/504) | |
| Overall Positive Call Rate (All Variants) | 86.2% | |
| APA (all variants) | > 92% | |
| ANA (all variants) | > 99% | |
| TMB %CV | 3.5% | |
| Lot to Lot Precision | APA (Variants with Evidence of Clinical Significance) | 96.1% to 98.7% |
| ANA (Variants with Evidence of Clinical Significance) | 99.8% to 99.9% | |
| TMB %CV | 40 tumor types. |
- Accuracy: 582 samples with PGDx elio tissue complete data and orthogonal data. Specifically:
- SNVs/Indels: 582 samples.
- ERBB2 Amplifications (FISH comparison): 147 cases (all); 120 cases (excluding borderline).
- ALK Translocations (FISH comparison): 71 cases. Additional in silico simulation on 10 clinical samples (410 observations).
- RET Translocations (FISH comparison): 27 cases. 3 RET translocation-positive cell lines also tested.
- TMB: 118 cases across 8 tumor types.
- MSI: 283 samples across 18 tumor types.
- Wild Type Calls: 112 specimens.
- Interlaboratory Reproducibility: 13 FFPE tissue specimens and 1 cell line (14 samples total), tested in duplicate by 2 operators on 12 sequencing runs across 3 non-consecutive days at 3 independent laboratory sites (504 total replicates).
- Lot to Lot Precision: 5 test cases in triplicate across 3 unique kit lots (45 observations).
Data Provenance: The document does not explicitly state the country of origin for the clinical samples. However, it mentions "clinical FFPE specimens," implying real-world patient data. The studies are described in a factual manner, suggesting they are retrospective analyses of collected samples for analytical validation purposes.
3. Number of Experts Used to Establish Ground Truth and Qualifications
The document does not specify the number or qualifications of experts involved in establishing the ground truth for the test set.
Instead, the ground truth for performance evaluation (accuracy) is established by:
- Orthogonal methods:
- "2 NGS targeted panels" and "PCR" (for SNVs and Indels).
- "ERBB2 FISH," "ALK FISH," "RET FISH" (for amplifications and translocations).
- "matched tumor-normal whole exome sequencing results" (for TMB).
- "MSI PCR" (for MSI).
- Validated assays/literature: For the 3 RET translocation-positive cell lines.
4. Adjudication Method for the Test Set
The document does not describe any expert adjudication process for resolving discrepancies or establishing ground truth for the test set. The ground truth is primarily based on the results from the orthogonal methods. For the in silico studies, the ground truth is derived from down-sampling "clinical samples."
5. MRMC Comparative Effectiveness Study
No multi-reader multi-case (MRMC) comparative effectiveness study was mentioned or performed. This device is a molecular diagnostic assay, not an imaging AI tool, and thus comparative effectiveness with human readers improving with AI assistance is not applicable in this context.
6. Standalone Performance
Yes, the entire document describes studies of the standalone (algorithm only, without human-in-the-loop performance) of the PGDx elio tissue complete assay. The goal is to demonstrate the analytical performance of the automated workflow, from sample preparation to data analysis and variant calling, against established ground truth methods.
7. Type of Ground Truth Used
The primary type of ground truth used is orthogonal methods, specifically:
- Other NGS targeted panels: For SNVs and indels.
- PCR: For certain SNVs/indels and MSI.
- FISH (Fluorescence In-Situ Hybridization): For gene amplifications (ERBB2) and translocations (ALK, RET).
- Whole Exome Sequencing (WES): For Tumor Mutation Burden (TMB).
- Cell line characterization/literature: For known variants in control cell lines.
This is a form of reference standard ground truth, where the device's performance is compared against established, validated measurement techniques.
8. Sample Size for the Training Set
The document details analytical validation (testing) and reproducibility studies. It does not provide information about a "training set" size. As a molecular diagnostic assay, its development likely involves internal data for algorithm development and optimization, but specific training set sizes are not mentioned in this regulatory submission, which focuses on validation data.
9. How the Ground Truth for the Training Set Was Established
Since the document does not discuss a specific "training set" or its size, it also does not describe how ground truth for such a set was established. The focus is on the performance of the final, locked-down algorithm against independent test data with ground truth established by orthogonal methods.
§ 866.6080 Next generation sequencing based tumor profiling test.
(a)
Identification. A next generation sequencing (NGS) based tumor profiling test is a qualitative in vitro diagnostic test intended for NGS analysis of tissue specimens from malignant solid neoplasms to detect somatic mutations in a broad panel of targeted genes to aid in the management of previously diagnosed cancer patients by qualified health care professionals.(b)
Classification. Class II (special controls). The special controls for this device are:(1) Premarket notification submissions must include the following information:
(i) A detailed description of all somatic mutations that are intended to be detected by the test and that are adequately supported in accordance with paragraph (b)(1)(v) of this section and reported in the test results in accordance with paragraph (b)(2)(iv) of this section, including:
(A) A listing of mutations that are cancer mutations with evidence of clinical significance.
(B) As appropriate, a listing of mutations that are cancer mutations with potential clinical significance.
(ii) The indications for use must specify the following:
(A) The test is indicated for previously diagnosed cancer patients.
(B) The intended specimen type(s) and matrix (
e.g., formalin-fixed, paraffin-embedded tumor tissue).(C) The mutation types (
e.g., single nucleotide variant, insertion, deletion, copy number variation or gene rearrangement) for which validation data has been provided.(D) The name of the testing facility or facilities, as applicable.
(iii) A detailed device description including the following:
(A) A description of the test in terms of genomic coverage, as follows:
(
1 ) Tabulated summary of all mutations reported, grouped according to gene and target region within each gene, along with the specific cDNA and amino acid positions for each mutation.(
2 ) A description of any within-gene targeted regions that cannot be reported and the data behind such conclusion.(B) Specifications for specimen requirements including any specimen collection devices and preservatives, specimen volume, minimum tumor content, specimen handling, DNA extraction, and criteria for DNA quality and quantity metrics that are prerequisite to performing the assay.
(C) A detailed description of all test components, reagents, instrumentation, and software required. Detailed documentation of the device software including but not limited to, software applications and hardware-based devices that incorporate software.
(D) A detailed description of the methodology and protocols for each step of the test, including description of the quality metrics, thresholds, and filters at each step of the test that are implemented for final result reporting and a description of the metrics for run-failures, specimen-failures, invalids, as applicable.
(E) A list of links provided by the device to the user or accessed by the device for internal or external information (
e.g., decision rules or databases) supporting clinical significance of test results for the panel or its elements in accordance with paragraphs (b)(1)(v) and (b)(2)(vi) of this section.(F) A description of internal and external controls that are recommended or provided and control procedures. The description must identify those control elements that are incorporated into the testing procedure.
(iv) Information demonstrating analytical validity of the device according to analytical performance characteristics, evaluated either specifically for each gene/mutation or, when clinically and practically justified, using a representative approach based on other mutations of the same type, including:
(A) Data that adequately supports the intended specimen type (
e.g., formalin-fixed, paraffin-embedded tumor tissue), specimen handling protocol, and nucleic acid purification for specific tumor types or for a pan-tumor claim.(B) A summary of the empirical evidence obtained to demonstrate how the analytical quality metrics and thresholds were optimized.
(C) Device precision data using clinical samples to adequately evaluate intra-run, inter-run, and total variability. The samples must cover all mutation types tested (both positive and negative samples) and include samples near the limit of detection of the device. Precision must be assessed by agreement within replicates on the assay final result for each representative mutation, as applicable, and also supported by sequencing quality metrics for targeted regions across the panel.
(D) Description of the protocols and/or data adequately demonstrating the interchangeability of reagent lots and multiplexing barcodes.
(E) A description of the nucleic acid assay input concentration range and the evidence to adequately support the range.
(F) A description of the data adequately supporting the limit of detection of the device.
(G) A description of the data to adequately support device accuracy using clinical specimens representing the intended specimen type and range of tumor types, as applicable.
(
1 ) Clinical specimens tested to support device accuracy must adequately represent the list of cancer mutations with evidence of clinical significance to be detected by the device.(
2 ) For mutations that are designated as cancer mutations with evidence of clinical significance and that are based on evidence established in the intended specimen type (e.g., tumor tissues) but for a different analyte type (e.g., protein, RNA) and/or a measurement (e.g., incorporating a score or copy number) and/or with an alternative technology (e.g., IHC, RT-qPCR, FISH), evidence of accuracy must include clinically adequate concordance between results for the mutation and the medically established biomarker test (e.g., evidence generated from an appropriately sized method comparison study using clinical specimens from the target population).(
3 ) For qualitative DNA mutations not described in paragraph (b)(1)(iv)(G)(2 ) of this section, accuracy studies must include both mutation-positive and wild-type results.(H) Adequate device stability information.
(v) Information that adequately supports the clinical significance of the panel must include:
(A) Criteria established on what types and levels of evidence will clinically validate a mutation as a cancer mutation with evidence of clinical significance versus a cancer mutation with potential clinical significance.
(B) For representative mutations of those designated as cancer mutations with evidence of clinical significance, a description of the clinical evidence associated with such mutations, such as clinical evidence presented in professional guidelines, as appropriate, with method comparison performance data as described in paragraph (b)(1)(iv)(G) of this section.
(C) For all other mutations designated as cancer mutations with potential clinical significance, a description of the rationale for reporting.
(2) The 21 CFR 809.10 compliant labeling and any product information and test report generated, must include the following, as applicable:
(i) The intended use statement must specify the following:
(A) The test is indicated for previously diagnosed cancer patients.
(B) The intended specimen type(s) and matrix (
e.g., formalin-fixed, paraffin-embedded tumor tissue).(C) The mutation types (
e.g., single nucleotide variant, insertion, deletion, copy number variation or gene rearrangement) for which validation data has been provided.(D) The name of the testing facility or facilities, as applicable.
(ii) A description of the device and summary of the results of the performance studies performed in accordance with paragraphs (b)(1)(iii), (b)(1)(iv), and (b)(1)(v) of this section.
(iii) A description of applicable test limitations, including, for device specific mutations validated with method comparison data to a medically established test in the same intended specimen type, appropriate description of the level of evidence and/or the differences between next generation sequencing results and results from the medically established test (
e.g., as described in professional guidelines).(iv) A listing of all somatic mutations that are intended to be detected by the device and that are reported in the test results under the following two categories or equivalent designations, as appropriate: “cancer mutations panel with evidence of clinical significance” or “cancer mutations panel with potential clinical significance.”
(v) For mutations reported under the category of “cancer mutations panel with potential clinical significance,” a limiting statement that states “For the mutations listed in [cancer mutations panel with potential clinical significance or equivalent designation], the clinical significance has not been demonstrated [with adequate clinical evidence (
e.g., by professional guidelines) in accordance with paragraph (b)(1)(v) of this section] or with this test.”(vi) For mutations under the category of “cancer mutations panel with evidence of clinical significance,” or equivalent designation, link(s) for physicians to access internal or external information concerning decision rules or conclusions about the level of evidence for clinical significance that is associated with the marker in accordance with paragraph (b)(1)(v) of this section.