(39 days)
The MolecularMD MRDx BCR-ABL Test is an in vitro diagnostic test for the quantitative detection of BCR-ABL 1 transcripts (e13a2/b2a2 and/or e14a2/63a2) and the ABL1 endogenous control mRNA in peripheral blood specimens from patients previously diagnosed with t(9:22) positive chronic myeloid leukemia (CML). The ratio of BCR-ABL1 is calculated and reported on the WHO International Scale. The test utilizes quantitative, real-time reverse transcription polymerase chain reaction performed on the Applied Biosystems 7500 Fast Dx instrument.
The MolecularMD MRDx BCR-ABL Test is intended to measure BCR-ABL mRNA transcript levels in patients diagnosed with t(9;22) positive CML during monitoring of treatment with Tyrosine Kinase Inhibitors (TKIs).
The device is also intended to be used in the serial monitoring for BCR-ABL mRNA transcript levels as an aid in identifying CML patients in the chronic phase being treated with nilotinib who may be candidates for treatment discontinuation and for monitoring of treatment-free remission.
The test does not differentiate between e13a2 or e14a2 fusion transcripts and does not monitor other rare fusion franscripts resulting from t(9:22). The test is not intended for the diagnosis of CML.
The MolecularMD MRDx® BCR-ABL Test is a quantitative, real-time polymerase chain reaction test that provides quantitation of BCR-ABL1 (hereafter BCR-ABL) transcript e13a2/b2a2 or e14a2/b3a2 and ABL1 (hereafter ABL) transcript levels in RNA extracted from peripheral blood samples collected from CML patients. Peripheral blood is collected in either EDTA or PAXgene Blood RNA Tubes. Each collection tube type requires a specified RNA extraction method and RNA input amount. Total RNA is extracted from peripheral blood and serves as the template for RT-qPCR. The test is performed using a onestep RT-qPCR protocol wherein the reverse transcription and quantitative, real-time PCR reactions are performed in the same well. BCR-ABL and ABL amplicons are generated and detected in real-time using TaqMan® MGB probes. The MolecularMD MRDx BCR-ABL Test is performed on the Applied Biosystems 7500 Fast Dx Real-Time PCR Instrument. The instrument integrates a thermal cycler, a fluorometer and application specific software. The ABI 7500 Fast Dx instrument software v1.4.1 calculates the BCR-ABL and ABL copy numbers using a standard curve generated with calibrators on each individual plate. The data are exported as a CSV file for further analysis in the MRDx® BCR-ABL Test Software. The BCR-ABL/ABL ratio is calculated and converted to the International Scale by the MRDx BCR-ABL Test Software. The MRDx BCR-ABL Test Software is used to analyze all test results. This software, provided with the MRDx BCR-ABL Test, is used to calculate the BCR-ABL/ABL % IS and MR value for patient sample using the conversion factor for the MRDx BCR-ABL Test after validating each MRDx BCR-ABL Test result against the run acceptance criteria and sample acceptance criteria.
The MolecularMD MRDx BCR-ABL Test is an in vitro diagnostic test for the quantitative detection of BCR-ABL1 transcripts and ABL1 endogenous control mRNA in peripheral blood specimens from patients previously diagnosed with t(9;22) positive chronic myeloid leukemia (CML). It calculates the ratio of BCR-ABL1 to ABL1 and reports it on the WHO International Scale. The device is intended to measure BCR-ABL mRNA transcript levels in CML patients during monitoring of treatment with Tyrosine Kinase Inhibitors (TKIs). It is also intended for serial monitoring of BCR-ABL mRNA transcript levels to aid in identifying CML patients in the chronic phase being treated with nilotinib who may be candidates for treatment discontinuation and for monitoring of treatment-free remission.
Here's an overview of the acceptance criteria and study data:
1. Table of Acceptance Criteria and Reported Device Performance
The document does not explicitly present a consolidated table of acceptance criteria for all performance characteristics. However, individual acceptance criteria are mentioned within the description of each study. Below is a summary of acceptance criteria and the device's reported performance, extracted from the document:
| Performance Characteristic | Acceptance Criteria | Reported Device Performance |
|---|---|---|
| Traceability to WHO IS | High traceability to WHO reference standards, consistent conversion factor across multiple lots. | Deming regression analyses showed a conversion factor of 1.1 for all three lots (A, B, C), demonstrating consistency. Data from all three lots showed high traceability to WHO reference standards. Slope was approximately 1.0 (0.98-1.0) and correlation was 1.0 for all lots. |
| Accuracy (Correlation to ddPCR) | High concordance between MRDx BCR-ABL Test results and the reference ddPCR test results, with acceptable predicted systematic differences at clinical decision points. (Implicit: Deming regression slope close to 1, y-intercept close to 0, high Pearson coefficient). | EDTA: Deming Slope: 0.96 (95% CI [0.94, 0.99]), Deming Y-Intercept: 0.077 (95% CI [0.0027, 0.17]), Pearson Coefficient: 0.984. Predicted difference at MR3: -0.031 (95% CI [-0.053, -0.0049]), MR4.0: -0.067 (95% CI [-0.10, -0.031]), MR4.5: -0.085 (95% CI [-0.13, -0.039]). |
| PAXgene: Deming Slope: 0.98 (95% CI [0.95, 1.0]), Deming Y-Intercept: -0.021 (95% CI [-0.11, 0.056]), Pearson Coefficient: 0.985. Predicted difference at MR3: -0.088 (95% CI [-0.11, -0.062]), MR4.0: -0.11 (95% CI [-0.15, -0.071]), MR4.5: -0.12 (95% CI [-0.17, -0.071]). Conclusion: High concordance observed for both tube types. | ||
| Limit of Blank (LoB) | Not explicitly stated as a numerical criterion, but implies no measurable BCR-ABL values in negative samples. | Out of 180 replicates from 30 BCR-ABL negative samples, 177 had no measurable BCR-ABL values. Three (1.7%) had measurements well below the LoD and were reported as undetected. |
| Limit of Detection (LoD) | Not explicitly stated as a numerical criterion, but the calculated LoD values should be acceptable for clinical use. | EDTA: Estimated LoD was 0.00029% IS (MR5.5) for both e13a2 and e14a2 transcripts. |
| PAXgene: Estimated LoD was 0.00039% IS (MR5.4) for e13a2 and 0.00029% IS (MR5.5) for e14a2. The final LoD was determined to be 0.00029% IS (MR5.5) for EDTA and 0.00039% IS (MR5.4) for PAXgene. | ||
| Limit of Quantitation (LoQ) | Total Error (TE) ≤ 0.5 log10. LoQ to be set at or below a clinically relevant threshold (e.g., MR4.5 for reporting). | EDTA: LoQ was 0.0016% IS (MR4.8). |
| PAXgene: LoQ was 0.0025% IS (MR4.6). The MRDx BCR-ABL Test Software limits the reported LoQ and quantitated results to 0.0032% IS (MR4.5) for both blood tube types. | ||
| Analytical Specificity (Interfering Substances) | No clinically significant difference in results when compared to matrix control samples. | Endogenous: Largest difference between endogenous substance spike and matrix control for PAXgene was -0.080 log10 (95% CI: -0.32 to 0.16) at MR4.5. For EDTA, it was 0.060 log10 (95% CI: -0.042 to 0.16) at MR4.5. No clinically significant difference observed. |
| Exogenous: Largest difference between exogenous substance spike and matrix control for PAXgene was 0.040 log10 (95% CI: -0.010 to 0.090) at MR3.0. For EDTA, it was 0.010 log10 (95% CI: -0.022 to 0.22) at MR4.5. No clinically significant difference observed. Conclusion: None of the potential interferents had significant interference. | ||
| Analytical Specificity (Primer Specificity) | Primers and probes amplify intended targets, no cross-reactivity with ABL2 IVT RNA. Cross-reactivity with e19a2 BCR-ABL is expected but mitigated by labeling. | Sequencing results of PCR products for BCR-ABL and ABL matched Genbank sequences, demonstrating amplification of intended targets. No cross-reactivity with ABL2 IVT RNA observed. Cross-reactivity with e19a2 BCR-ABL transcript was observed as expected, and addressed in labeling with a precaution statement. |
| Analytical Specificity (Specimen Cross Contamination) | No negative samples (0% IS) should have a BCR-ABL/ABL % IS value detectable above the LoD of the assay when juxtaposed with high positive samples. | No negative samples for either extraction method had a detectable BCR-ABL/ABL % IS value above the LoD. Conclusion: No significant carryover between wells. |
| Precision (Repeatability) | SD log10 ≤ 0.25. | PAXgene: Highest SD was 0.10 for e14a2 MR4.5. All levels for both transcripts passed (SD ≤ 0.25 log10). |
| EDTA: Highest SD was 0.087 for e14a2 MR4.5. All levels for both transcripts passed (SD ≤ 0.25 log10). | ||
| Precision (Reproducibility) | Total %CV for MR values should be within acceptable limits (typically 0.05) or acceptable Degree of Nonlinearity). Assay should be linear across the reportable range. | EDTA: e13a2 linear from MR0.93 to MR5.1 (max SD 0.22), e14a2 linear from MR0.99 to MR5.0 (max SD 0.26). |
| PAXgene: e13a2 linear from MR0.78 to MR4.8 (max SD 0.13), e14a2 linear from MR0.93 to MR4.9 (max SD 0.31). Conclusion: The assay is linear across the reportable range of MR1.0 to MR4.5 for both tube types. | ||
| Kit Stability (Real-time & Freeze-Thaw) | Stability for a specified shelf life and open-vial stability (freeze-thaw cycles and time after initial use). Acceptable performance of calibration curve, control results, and WHO secondary standards. | Passing test results data for at least 16 months for 3 lots stored at two temperatures. Shelf life of 15 months at -30 to -15°C and -80 to -65°C. Open-vial stability of 3 freeze-thaw cycles and up to 2 months following initial use. |
| Reagent Stability (In Use) | Mean of test samples ± 0.5 MR of the control. Differences between standard and extreme conditions |
§ 866.6060 BCR-ABL quantitation test.
(a)
Identification. A BCR-ABL quantitation test is identified as a reverse transcription-quantitative polymerase chain reaction (RT-qPCR) test for the quantitation of BCR-ABL1 expressed on the International Scale (IS) and control transcripts in total RNA from whole blood of diagnosed t(9;22) positive chronic myeloid leukemia (CML) patients during monitoring of treatment with tyrosine kinase inhibitors. This test is not intended for the diagnosis of CML.(b)
Classification. Class II (special controls). The special controls for this device are:(1) Premarket notification submissions must include the following information:
(i) The indication for use must indicate the variant(s) for which the assay was designed and validated, for example BCR-ABL e13a2 and/or e14a2.
(ii) A detailed description of all components in the test, including the following:
(A) A detailed description of the test components, all required reagents, instrumentation and equipment, including illustrations or photographs of non-standard equipment or methods;
(B) Detailed documentation of the device software including, but not limited to, standalone software applications and hardware-based devices that incorporate software;
(C) Methodology and protocols for control procedures for the assay to allow reporting on the International Scale;
(D) A description of the result outputs, analytical sensitivity of the assay, and the range of values that will be reported; and
(E) A description of appropriate internal and external controls that are recommended or provided. The description must identify those control elements that are incorporated into the testing procedure.
(iii) Information that demonstrates the performance characteristics of the test, including:
(A) For indications for use based on a threshold established in a predicate device of this generic type, device performance data from either a method comparison study to the predicate device or through a clinical study demonstrating clinical validity using well-characterized prospectively or retrospectively obtained clinical specimens, as appropriate, representative of the intended use population;
(B) For indications for use based on a threshold not established in a predicate device of this generic type, device performance data from a clinical study demonstrating clinical validity using well-characterized prospectively or retrospectively obtained clinical specimens, as appropriate, representative of the intended use population;
(C) Device reproducibility data generated, using a minimum of three sites, of which at least two sites must be external sites, with two operators at each site. Each site must conduct a minimum of three runs per operator over non-consecutive days evaluating a minimum of five different BCR-ABL concentrations that span and are well distributed over the measuring range and include MR3 (0.1 percent IS). Results shall be reported as the standard deviation and percentage coefficient of variation for each level tested. Prespecified acceptance criteria must be provided and followed;
(D) Device precision data using clinical samples to evaluate the within-lot, between-lot, within-run, between run, and total variation;
(E) Device linearity data using a dilution panel created from clinical samples;
(F) Device analytic sensitivity data, including limit of blank, limit of detection, and limit of quantification;
(G) Device specificity data, including interference and cross-contamination; and
(H) Device stability data, including real-time stability of samples under various storage times, temperatures, and freeze-thaw conditions.
(iv) Identification of risk mitigation elements used by your device, including a detailed description of all additional procedures, methods, and practices incorporated into the instructions for use that mitigate risks associated with testing using your device.
(2) Your 21 CFR 809.10 compliant labeling must include the following:
(i) The intended use in your 21 CFR 809.10(a)(2) and (b)(2) complaint labeling must include an indication for use statement that reads “This test is not intended for the diagnosis of CML”; and
(ii) A detailed description of the performance studies conducted to comply with paragraph (b)(1)(iii) of this section and a summary of the results.
(3) Your device output must include results on the International Scale (IS) and your assay must include multipoint calibration controls traceable to a relevant international reference panel (
e.g., the World Health Organization International Genetic Reference Panel for quantitation of BCR-ABL mRNA).