(216 days)
The clonoSEQ Assay is an in vitro diagnostic that uses multiplex polymerase chain reaction (PCR) and next-generation sequencing (NGS) to identify and quantify rearranged IgH (VDJ), IgH (DJ), IgK and IgL receptor gene sequences, as well as translocated BCL1/1gH (J) and BCL2/1gH (J) sequences in DNA extracted from bone marrow from patients with B-cell acute lymphoblastic leukemia (ALL) or multiple myeloma (MM), and blood or bone marrow from patients with chronic lymphocytic leukemia (CLL).
The clonoSEQ Assay measures minimal residual disease (MRD) to monitor changes in burden of disease during and after treatment. The test is indicated for use by qualified healthcare professionals in accordance with professional guidelines for clinical decision-making and in conjunction with other clinicopathological features.
The clonoSEQ Assay is a single-site assay performed at Adaptive Biotechnologies Corporation.
The clonoSEQ Assay is a next-generation sequencing (NGS) based assay that identifies rearranged IgH (VDJ), IgH (DJ), IgK, and IgL receptor gene sequences, as well as translocated BCL1/IgH (J) and BCL2/IgH (J) sequences. The assay also includes primers that amplify specific genomic regions present as diploid copies in normal genomic DNA (gDNA) to allow determination of total nucleated cell content.
Testing begins with gDNA extracted from the specimen supplied (Figure 1). Extracted gDNA quality is assessed and rearranged immune receptors are amplified using a multiplex PCR. Reaction-specific index barcode sequences for sample identification are added to the amplified receptor sequences by PCR. Sequencing libraries are prepared from barcoded amplified DNA, which are then sequenced by synthesis using NGS. Raw sequence data are uploaded from the sequencing instrument to the Adaptive analysis pipeline. These sequence data are analyzed in a multi-step process: first, a sample's sequence data are identified using the sample index sequences. Next, data are processed using a proprietary algorithm with in-line controls to remove amplification bias. When the clonoSEQ Clonality (ID) assessment is conducted, the immune repertoire of the sample is checked for the presence of DNA sequences specific to "dominant" clone(s) consistent with the presence of a lymphoid malignancy. Each sequence that is being considered for MRD tracking is compared against a B cell repertoire database and assigned a uniqueness value that, together with its abundance relative to other sequences, is used to assign the sequence to a sensitivity bin which will be used in the estimation of the reported LoD and LoO on the patient report. During clonoSEQ Tracking (MRD) assessment, the complete immunoglobulin receptor repertoire is again assessed, and the previously identified dominant clonotype sequence(s) are detected and quantified to determine the sample MRD level. The clonoSEQ Assay MRD assessment measures residual disease in a biologic sample.
Here's a breakdown of the acceptance criteria and study details for the Adaptive Biotechnologies clonoSEQ Assay, based on the provided FDA 510(k) summary:
This device, the Adaptive Biotechnologies clonoSEQ Assay, is an in vitro diagnostic (IVD) that identifies and quantifies rearranged immune receptor gene sequences (IgH, IgK, IgL) and translocated BCL1/IgH and BCL2/IgH sequences using multiplex PCR and Next-Generation Sequencing (NGS). It measures Minimal Residual Disease (MRD) in patients with B-cell acute lymphoblastic leukemia (ALL), multiple myeloma (MM), and chronic lymphocytic leukemia (CLL) to monitor disease burden. The current submission is an expansion of indications to include blood samples from CLL patients.
1. A table of acceptance criteria and the reported device performance
The provided document doesn't explicitly list "acceptance criteria" in a single table, but rather describes the performance characteristics that were measured and the outcomes for each. I will compile these for the CLL in Blood indication, as this is the focus of the 510(k) expansion.
Acceptance Criteria & Reported Device Performance for clonoSEQ Assay (CLL in Blood)
| Performance Characteristic | Acceptance Criteria (Implicit) | Reported Device Performance (CLL in Blood) |
|---|---|---|
| Precision (MRD Frequency) | %CV within acceptable clinical/analytical limits (not explicitly stated, but inferred from successful results) | Range: 18.7% - 54.9% CV. |
- At 500 ng DNA input: 21.9% - 54.9% CV
- At 2 µg DNA input: 20.8% - 51.6% CV
- At 20 µg DNA input: 18.7% - 49.2% CV
(Comparable to BMA precision) |
| Precision (Malignant Cells Detected) | %CV within acceptable clinical/analytical limits, primarily influenced by cell number. | Range: 19% CV (at 765.70 cells) to 53% CV (at 3.10 cells).
Primarily due to residual variability; other factors (operator, instruments, reagents, day, run) contributed 0%-10% CV. |
| Linearity | Maximum deviation from linearity (based on quadratic or cubic fit) less than 5%. | Met for all DNA inputs (20µg, 2µg, 500ng) across the entire tested MRD frequency range (0 to 1x10^-3 / 4x10^-3). - Slopes: 0.989 - 0.997 (indicating strong linearity)
- Intercepts: -0.009 to -0.075 |
| Accuracy (Concordance with mpFC) | High positive percent agreement (PPA) and understanding of negative percent agreement (NPA) reflecting greater sensitivity. | PPA: 98.9% (95% CI: 94.3%-100%)
NPA: 47.5% (95% CI: 40.5%-54.6%)
(NPA lower due to clonoSEQ's higher sensitivity detecting MRD where mpFC calls negative) |
| Limit of Blank (LoB) | LoB should be zero or negligible. | LoB was confirmed as zero (95th percentile of trackable sequences in healthy blood was zero). |
| Limit of Detection (LoD) / Limit of Quantitation (LoQ) | LoD/LoQ for blood should be comparable to or lower than previously determined values for bone marrow. | LoD and LoQ for CLL in blood were lower or within the 95% CI of bootstrapped prior BMA data, confirming comparability. |
| Analytical Specificity (Interfering Substances) | Mean MRD frequency difference ± 30% when comparing with and without interferent substances. | All tested endogenous and exogenous substances met acceptance criteria. - Endogenous: Bilirubin (conjugated & unconjugated), Hemoglobin, Cholesterol, Triglycerides.
- Exogenous: K2EDTA, K3EDTA, Heparin, Chloroform.
(MRD results not substantially influenced). |
| Cross-Contamination/Sample Carryover | No significant contamination events leading to false positive ID or MRD results. | - No false calibrations for run-to-run (0/44 BMA, 0/44 BMMC). - One single well-to-well false calibration (1/44 BMA) at a very low template count (83 templates), not associated with cell lines.
- Blood: No contamination or disease clone-sharing events leading to false positive ID/MRD results.
- PCR/Library/Sequencing: No run-to-run contamination (0/36 tests). 8/712 well-to-well events (likely vendor-related primer barcode issue)
§ 866.6100 DNA-based test to measure minimal residual disease in hematological malignancies.
(a)
Identification. A DNA-based test to measure minimal residual disease in hematological malignancies is a prescription in vitro diagnostic device that identifies and quantifies specific nucleic acid sequences within human tissues to estimate the percentage of cells that harbor the specific sequence(s). The test is intended to be used as an aid to measure minimal residual disease to assess the change in burden of disease during monitoring of treatment. The test is indicated for use by qualified healthcare professionals in accordance with professional guidelines for clinical decision-making, in conjunction with other clinicopathological features.(b)
Classification. Class II (special controls). The special controls for this device are:(1) Design verification and validation must include:
(i) A detailed description of the device, including:
(A) A detailed description of all test components, reagents, instrumentation, and software, including software applications and any hardware-based devices that incorporate software.
(B) A detailed description of all genomic regions that are detected and quantified by the assay.
(C) 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, and invalids, as appropriate.
(D) Detailed specifications and procedures for sample collection, processing, and storage.
(E) A description of the internal and external controls that are recommended or provided. The description must identify those control elements that are incorporated into the testing procedure. If appropriate, this description must include a description of the controls and control procedures used during the sequencing and data analysis.
(ii) Identification of risk mitigation elements used by the device, including a detailed description of all additional procedures, methods, and practices incorporated into the instructions for use that mitigate risks associated with use of the device.
(iii) As part of the risk management activities, an appropriate end user device training program must be offered as an effort to mitigate the risk of failure from user error, as appropriate.
(iv) Description of analytical and clinical studies, including:
(A) Device performance data that demonstrates the ability to measure minimal residual disease in the claimed specimen type(s) from patients that are representative of the intended use population. Data can be obtained via:
(
1 ) A method comparison study comparing the device to a predicate device with clinical data for the specified hematological neoplastic indication using the specified specimen type(s); or(
2 ) A clinical study demonstrating clinical validity using well characterized clinical specimens from patients with known clinical outcomes using a study design deemed acceptable by FDA.(B) Device precision (repeatability and reproducibility) data using clinical samples covering the range of minimal residual disease frequencies reported by the test and covering the stated range of DNA inputs that are indicated as allowable for use with the test. Results shall be reported as the standard deviation and/or percentage coefficient of variation with the 95 percent confidence interval for each level tested. The study must evaluate all sources of variability, including, as appropriate, between-site and between operator (minimum of three sites of which two must be external with a minimum of two operators per site), between-day (minimum of 3 days), between-run, within-run, between-lot (minimum of three lots), between instrument (minimum of three instruments), and total variation.
(C) Device linearity data generated from samples covering the device measuring range using a dilution panel created from clinical samples.
(D) Device accuracy by comparison to flow cytometry across the measuring interval or to the predicate method across the measuring interval.
(E) Device analytic sensitivity data, including limit of blank, limit of detection, and limit of quantitation, using a dilution panel created from clinical samples.
(F) Analytical specificity data, including interference and cross-contamination, and index cross-contamination, as appropriate.
(G) Validation of pre-analytical methods, including DNA extraction methods and cell enrichment methods, as appropriate.
(H) Device stability data, including real-time stability of reagents under various storage times and temperatures.
(I) Specimen and prepared sample stability data established for each specimen matrix in the anticoagulant combinations and storage/use conditions that will be indicated, including specimen transport, as appropriate.
(2) The intended use for the labeling required under § 809.10(a)(4) of this chapter and for the labeling required under § 809.10(b)(5)(ii) of this chapter, as applicable, must include:
(i) The clinical hematopoietic malignancy for which the assay was designed and validated (
e.g., multiple myeloma or B-cell acute lymphoblastic leukemia);(ii) Specimen type (
e.g., bone marrow);(iii) The specific DNA regions that are being identified and quantified (
e.g., rearranged IgH (VDJ), IgH (DJ), IgK, and IgL receptor gene sequences); and(iv) A statement that the results are indicated to be interpreted by qualified healthcare professionals in accordance with professional guidelines for clinical decision-making in conjunction with other clinicopathological features.
(3) The labeling required under § 809.10(b) of this chapter must include information that demonstrates the performance characteristics of the test, including a detailed summary of the performance studies conducted and their results, as described in paragraphs (b)(1)(iv)(A) through (I) of this section.
(4) The device output, including any test report, must include the estimated minimal residual disease (MRD) frequency and an appropriate range of the uncertainty of that frequency based on the amount of DNA that was evaluated by the test and the number of specific nucleic acid sequences that were detected (
e.g., “MRD = 1.2 × 10−5 [Range = 0.8 × 10−6 to 2.0 × 10−5 ]”).