Who is the manufacturer of ACTOnco, ACTOnco IVD?

Act Genomics filed the 510(k) submission for ACTOnco, ACTOnco IVD (K210017).

What is the FDA product code for ACTOnco, ACTOnco IVD?

PZM. It is classified under 21 CFR 866.6080 as a Class 2 device in the Pathology panel.

What is the regulatory pathway for ACTOnco, ACTOnco IVD?

510(k) clearance (submission type: Traditional).

Who can help prepare an FDA 510(k) submission like ACTOnco, ACTOnco IVD?

Innolitics — a medical-device software consultancy — provides regulatory and engineering support for FDA 510(k) submissions. See https://innolitics.com/services/510ks/ or contact https://innolitics.com/contact.

ACTOnco, ACTOnco IVD

K210017 · Act Genomics · PZM · Dec 23, 2022 · Pathology

Device Facts

Record ID	K210017
Device Name	ACTOnco, ACTOnco IVD
Applicant	Act Genomics
Product Code	PZM · Pathology
Decision Date	Dec 23, 2022
Decision	SESE
Submission Type	Traditional
Regulation	21 CFR 866.6080
Device Class	Class 2

Intended Use

The ACTOnco IVD assay is an in vitro diagnostic test that uses targeted next generation sequencing of formalin-fixed, paraffin-embedded tumor tissue from patients with solid malignant neoplasms to detect genetic alterations in a broad multi gene panel. The test is intended to provide information on point mutations, small insertions and deletions, ERBB2 gene amplification, and tumor mutational burden for use by qualified health care professionals in accordance with professional guidelines, and is not conclusive or prescriptive for labeled use of any specific therapeutic product. ACTOnco IVD is a single-site assay performed at ACT Genomics.

Device Story

ACTOnco IVD is a targeted NGS assay for solid malignant neoplasms. It inputs DNA extracted from FFPE tumor tissue; uses amplicon-based sequencing with tiled primers (targeting ~1.8Mb of the human genome) on the Ion GeneStudio S5 Prime System. The device identifies SNVs, Indels, ERBB2 amplification, and TMB. It is a single-site assay performed at ACT Genomics. Healthcare providers use the generated variant interpretation summary—containing biologic impact and therapeutic relevance—to inform clinical decision-making in accordance with professional guidelines. The device benefits patients by providing comprehensive tumor profiling to guide potential therapeutic options.

Clinical Evidence

Bench testing only. Precision/reproducibility study (20 samples, 48 replicates) showed 98.33% call rate for SNVs/Indels. LoD study established sensitivity for various variant types. Method comparison study (438 samples) against a validated NGS assay showed 97.85% PPA and 99.97% NPA for SNVs/Indels. ERBB2 amplification comparison (129 samples) against DISH showed 91.67% PPA and 100% NPA. TMB performance compared to WES (45 samples) showed a Spearman correlation of 0.885.

Technological Characteristics

Targeted amplicon-based NGS assay. DNA extracted from FFPE tissue. Semiconductor sequencing (Ion Torrent). Targets ~1.8Mb of human genome (oncogenes, tumor suppressors, drug metabolism, immune genes). Software includes variant calling, annotation, and TMB calculation. Single-site laboratory workflow. No matched normal required; uses pooled normal baseline.

Indications for Use

Indicated for patients with solid malignant neoplasms to detect genetic alterations (SNVs, Indels, ERBB2 amplification, TMB) in FFPE tumor tissue. For prescription use by qualified healthcare professionals. Not for use as a conclusive or prescriptive test for specific therapeutic products.

Regulatory Classification

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.

Special Controls

*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.

Predicate Devices

MSK-IMPACT (Integrated Mutation Profiling of Actionable Cancer Targets) (DEN170058)

Related Devices

P240010 — MI Cancer Seek · Caris Life Sciences · Nov 5, 2024
K250003 — GENESEEQPRIME NGS Tumor Profiling Assay (FFPE) (GS6005) · Geneseeq Technology, Inc. · Aug 29, 2025
P230011 — TruSight Oncology Comprehensive · Illumina, Inc. · Aug 21, 2024
K192063 — PGDx elio tissue complete · Personal Genome Diagnostics · Apr 24, 2020

Submission Summary (Full Text)

{0}------------------------------------------------ Image /page/0/Picture/0 description: The image contains the logo of the U.S. Food and Drug Administration (FDA). The logo consists of two parts: the Department of Health & Human Services logo on the left, and the FDA logo on the right. The FDA logo features the letters "FDA" in a blue square, followed by the words "U.S. FOOD & DRUG ADMINISTRATION" in blue text. December 23, 2022 ACT Genomics % David Kern Principal and Founder K2 Regulatory Consulting, LLC 479 Cumberland Drive Burlingame, California 94010 Re: K210017 Trade/Device Name: ACTOnco, ACTOnco IVD Regulation Number: 21 CFR 866.6080 Regulation Name: Next generation sequencing based tumor profiling test Regulatory Class: Class II Product Code: PZM Dated: September 19, 2022 Received: September 26, 2022 Dear David Kern: 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. Although this letter refers to your product as a device, please be aware that some cleared products may instead be combination products. The 510(k) Premarket Notification Database located at https://www.accessdata.fda.gov/scripts/cdrh/cfdocs/cfpmn/pmn.cfm identifies combination product submissions. 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. Please note: CDRH does not evaluate information related to contract liability warranties. We remind you, however, that device labeling must be truthful and not misleading. If your device is classified (see above) into either class II (Special Controls) or class III (PMA), it may be subject to additional controls. Existing major regulations affecting your device can be found in the Code of Federal Regulations, Title 21, Parts 800 to 898. 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 {1}------------------------------------------------ requirements, including, but not limited to: registration and listing (21 CFR Part 807); labeling (21 CFR Part 801 and Part 809); medical device reporting of medical device-related adverse events) (21 CFR 803) for devices or postmarketing safety reporting (21 CFR 4, Subpart B) for combination products (see https://www.fda.gov/combination-products/guidance-regulatory-information/postmarketing-safety-reportingcombination-products); good manufacturing practice requirements as set forth in the quality systems (OS) regulation (21 CFR Part 820) for devices or current good manufacturing practices (21 CFR 4, Subpart A) for combination products; and, if applicable, the electronic product radiation control provisions (Sections 531-542 of the Act); 21 CFR 1000-1050. Also, please note the regulation entitled, "Misbranding by reference to premarket notification" (21 CFR Part 807.97). For questions regarding the reporting of adverse events under the MDR regulation (21 CFR Part 803), please go to https://www.fda.gov/medical-device-safety/medical-device-reportingmdr-how-report-medical-device-problems. For comprehensive regulatory information about mediation-emitting products, including information about labeling regulations, please see Device Advice (https://www.fda.gov/medicaldevices/device-advice-comprehensive-regulatory-assistance) and CDRH Learn (https://www.fda.gov/training-and-continuing-education/cdrh-learn). Additionally, you may contact the Division of Industry and Consumer Education (DICE) to ask a question about a specific regulatory topic. See the DICE website (https://www.fda.gov/medical-device-advice-comprehensive-regulatoryassistance/contact-us-division-industry-and-consumer-education-dice) for more information or contact DICE by email (DICE@fda.hhs.gov) or phone (1-800-638-2041 or 301-796-7100). Sincerely, # Zivana Tezak-fragale -S Zivana Tezak, Ph.D. Branch Chief Division of Molecular Genetics and Pathology OHT7: Office of In Vitro Diagnostics Office of Product Evaluation and Quality Center for Devices and Radiological Health Enclosure {2}------------------------------------------------ # Indications for Use 510(k) Number (if known) K210017 Device Name ACTOnco IVD #### Indications for Use (Describe) The ACTOnco IVD assay is an in vitro diagnostic test that uses targeted next generation sequencing of formalin-fixed. paraffin-embedded tumor tissue from patients with solid malignant neoplasms to detect genetic alterations in a broad multi gene panel. The test is intended to provide informations, small insertions and deletions, ERBB2 gene amplification, and tumor mutational burden for use by qualified health care professionals in accordance with professional guidelines, and is not conclusive or prescriptive for labeled use of any specific therapeutic product. ACTOnco IVD is a single-site assay performed at ACT Genomics. | Type of Use (Select one or both, as applicable) | |-------------------------------------------------| |-------------------------------------------------| X Prescription Use (Part 21 CFR 801 Subpart D) | Over-The-Counter Use (21 CFR 801 Subpart C) #### CONTINUE ON A SEPARATE PAGE IF NEEDED. This section applies only to requirements of the Paperwork Reduction Act of 1995. #### *DO NOT SEND YOUR COMPLETED FORM TO THE PRA STAFF EMAIL ADDRESS BELOW.* The burden time for this collection of information is estimated to average 79 hours per response, including the time to review instructions, search existing data sources, gather and maintain the data needed and complete and review the collection of information. Send comments regarding this burden estimate or any other aspect of this information collection, including suggestions for reducing this burden, to: > Department of Health and Human Services Food and Drug Administration Office of Chief Information Officer Paperwork Reduction Act (PRA) Staff PRAStaff(@fda.hhs.gov "An agency may not conduct or sponsor, and a person is not required to respond to, a collection of information unless it displays a currently valid OMB number." {3}------------------------------------------------ # 510(k) Summary for ACTOnco IVD #### 1 Submitter ACT GENOMICS Co., LTD 3F., No 345, Xinhu 2nd Rd., Neihu Dist. Taipei City 11494, Taiwan Contact: Pei-Fang Chung, MSc, MBA, RAC Associate Director, Regulatory Affairs Phone: +886-2-27953660 ext 1303 Mobile: +886-928608630 Email: peifangchung@actgenomics.com #### 2 Submission Correspondent - US K2 Regulatory Consulting, LLC 479 Cumberland Drive, Burlingame, CA 94010, USA David Kern, MBA, RAC Principal and Founder Mobile: 1.650.888.8251 Email: dkern@k2regulatory.com #### 3 Device Identification | Name of Device: | ACTOnco IVD | |----------------------------|-------------------------------------------------------| | Common or Usual Name | ACTOnco | | Classification Name: | Next generation sequencing based tumor profiling test | | Classification Regulation: | 866.6080 | | Product Code: | PZM | | Device Class: | Class II | | Classification Panel: | Pathology | #### 4 Legally Marketed Predicate Device MSK-IMPACT (Integrated Mutation Profiling of Actionable Cancer Targets) DEN170058 {4}------------------------------------------------ #### Device Description റ The ACTOnco IVD assay is an in-vitro diagnostic assay intended to provide information for use by qualified health care professionals in accordance with professional guidelines, and is not conclusive or prescriptive for labeled use of any specific therapeutic product. The assay is a custom targeted sequencing platform, utilizing amplicon-based sequencing, to detect point mutations (single nucleotide variants, or SNVs), small insertions and deletions (Indels), ERBB2 gene amplification, and tumor mutational burden (TMB) in tumor specimens. The assay uses custom DNA primers corresponding to all exons and selected introns of oncogenes, tumor suppressor genes, drug metabolism genes, and immune-related genes. Primers are synthesized by a secondary manufacturer (Thermo Fisher Scientific). An overlapping amplicon approach is utilized in which tiled primers are designed to generate multiple overlapping amplicons of the same region to avoid allele dropout. In total, the primers target approximately 1.8Mb of the human genome. Genomic DNA is extracted from FFPE tissue samples. Sequence libraries are prepared through a multiplex polymerase chain reaction (PCR) amplification step to enrich target sequences. Target sequences are tagged with index oligonucleotide to identify individual sample and adaptor oligonucleotide to anchor the amplicon to complimentary oligonucleotides embedded on the surface of the sequencing bead. Target sequences on the sequencing beads are amplified using emulsion PCR before sequencing. Multiple barcoded sequence libraries (from different patients) are pooled and then sequenced on a Thermo Fisher Ion GeneStudio™ S5 Prime System. Sequence reads are then aligned to the reference human genome. By comparing the identity of bases from the tumor DNA and the reference human genome, variant alterations are identified in the tumor. The assay system includes a sequencing instrument, reagents (DNA extraction, library preparation and sequencing), software (operation of the sequencing instrument and variant calling), and standard operating procedures (SOPs) for the use of the system. ACT Genomics takes the responsibilities in monitoring the instrument; reagents and consumable materials which will be used in the assay process. Multiple software components will be used in the assay. The NGS raw read analysis will be done using Thermo Fisher software. Variant calling for SNVs, insertions and deletions will be done using Thermo Fisher software. Mutation and variant annotation will be done using software from ACT Genomics, the Cunningham Lab and Golden Helix software. ERBB2 gene amplification will be done using software from Boeva Lab. Tumor Purity and Zygosity will be done using software from Halgamuge Lab (Kaushalya Amarasinghe). Calculations for tumor mutational burden will be done using ACT Genomics software. {5}------------------------------------------------ #### Indication for Use Statement 6 The ACTOnco IVD assay is an in vitro diagnostic test that uses targeted next generation sequencing of formalin-fixed, paraffin-embedded tumor tissue from patients with solid malignant neoplasms to detect genetic alterations in a broad multi gene panel. The test is intended to provide information on point mutations, small insertions and deletions, ERBB2 gene amplification, and tumor mutational burden for use by qualified health care professionals in accordance with professional guidelines, and is not conclusive or prescriptive for labeled use of any specific therapeutic product. ACTOnco IVD is a single-site assay performed at ACT Genomics. ### Special conditions for use statement - For prescription use only - For in vitro diagnostic use Special instrument requirements - Thermo Fisher Ion GeneStudio™ S5 Prime System (qualified by ACT Genomics). . #### 7 Comparison of Technological Characteristics with The Predicate Device The ACTOnco IVD test has technological characteristics that are substantially equivalent to the predicate device as described in the tables below. Both the subject device and the predicate device use targeted, high throughput parallel sequencing for the detection genetic alterations. The subject device uses an amplicon-based target enrichment approach and Ion Torrent's semiconductor sequencing technology, whereas the predicate device used a hybrid capture target enrichment approach and Illumina's sequencing by synthesis (SBS) technology. Both sequencing platforms have been used in IVD devices that have been either cleared or approved by the FDA. Both devices use FFPE tumor tissue samples collected from patients with solid malignant neoplasms. The predicate device also uses matched normal samples, which are not required for the ACTOnco IVD test. The ACTOnco IVD test has established a baseline using pooled normal samples, which tumor samples are then compared. {6}------------------------------------------------ | Characteristic | Predicate MSK-IMPACT (DEN170058) | Subject Device ACTOnco IVD | |---------------------------------------------------------------------------------------------------------------------------------------------------------|--------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------| | | Similarities | | | Indications for Use | The MSK-IMPACT assay is a qualitative in vitro diagnostic test that uses targeted next generation sequencing of formalin-fixed paraffin-embedded tumor tissue matched with normal specimens from patients with solid malignant neoplasms to detect tumor gene alterations in a broad multi gene panel. The test is intended to provide information on somatic mutations (point mutations and small insertions and deletions) and microsatellite instability for use by qualified health care professionals in accordance with professional guidelines and is not conclusive or prescriptive for labeled use of any specific therapeutic product. MSK-IMPACT is a single-site assay performed at Memorial Sloan Kettering Cancer Center. | The ACTOnco IVD assay is an in vitro diagnostic test that uses targeted next generation sequencing of formalin-fixed, paraffin-embedded tumor tissue from patients with solid malignant neoplasms to detect genetic alterations in a broad multi gene panel. The test is intended to provide information on point mutations, small insertions and deletions, ERBB2 amplification, and tumor mutational burden for use by qualified health care professionals in accordance with professional guidelines and is not conclusive or prescriptive for labeled use of any specific therapeutic product. ACTOnco IVD is a single-site assay performed at ACT Genomics. | | Specimen Types | Formalin-fixed, paraffin- embedded (FFPE) tumor tissue with matched normal specimens from patients with solid malignant neoplasms | Formalin-fixed, paraffin- embedded (FFPE) tumor tissue from patients with solid malignant neoplasms | | Target Population | Patients with solid malignant neoplasms | Same | | Assay cut-off | MSK-IMPACT does not report mutations below 2% for known hotspot mutations and 5% for non- hotspot mutations. | Same | | Laboratory | Single site laboratory | Single site laboratory | | Characteristic | Predicate MSK-IMPACT (DEN170058) Differences | Subject Device ACTOnco IVD | | Sequencing Instrument | Illumina HiSeqTM 2500 Sequencer | Thermo Fisher Ion GeneStudioTM S5 Prime System | | Target Enrichment Technology | Hybrid Capture | Amplicon | | Genes on Panel | 468 | 440 | | Black List | 73 exons | 650 amplicons within 199 genes excluded from reporting SNV/ Indels due to pseudo gene or consistently low coverage (≤ 35x). | | Variant Type | Intended to provide information on somatic mutations (point mutations and small insertions and deletions), and microsatellite instability | ACTOnco provides information on somatic mutations as indicated and also includes copy number alteration and provides information on tumor mutational burden (TMB). No microsatellite instability. | | Determination of Pipeline Thresholds | • Based on >200X target coverage; • 100X for ≥ 98% target exons; • hotspot mutation calling threshold (mutation coverage (DP) ≥ 20, mutant reads (AD) ≥ 8, mutation frequency (VF) ≥ 2%, and non-hotspot mutation threshold (DP ≥ 20, AD ≥ 10, VF ≥ 5%) | • Based on ≥ 500x target coverage; • 100x for ≥85% target regions; • hotspot mutation calling threshold (mutation coverage (DP) ≥ 35, mutant reads (AD) ≥ 5, strand bias (SB) < 0.9, mutation frequency (VF) ≥ 2%), and non-hotspot mutation threshold (DP ≥ 35, AD ≥ 10, SB < 0.9, VF ≥ 5%). | | Controls | • Matched normal • Positive control • Negative control • No template control (NTC) | • Positive control • Negative control • No template control (NTC) | | Clinical Evidence Curation Oncopanel results are reported under one of these two categories: | Uses OncoKB, knowledge base that includes biologic, clinical and therapeutic information curated from professional guidelines and recommendations, therapeutic labeling, disease specific expert and advocacy group | A variant interpretation summary is generated, which includes biologic impact, variant specific effect and therapeutic relevance curated from professional guidelines and recommendations, therapeutic labeling, disease | | Characteristic | Predicate MSK-IMPACT (DEN170058) | Subject Device ACTOnco IVD | | Differences | | | | • “Cancer Mutations with Evidence of Clinical Significance” or • “Cancer Mutations with Potential Clinical Significance.” | recommendations, and medical literature. Classification criteria were developed by MSK to communicate the level of clinical evidence available for individual mutations in the test report. • OncoKB undergoes periodic updates through the review of new information by a panel of experts | specific expert and advocacy group recommendations, and medical literature. Classification criteria were developed by ACT Genomics with the reference of AMP guideline, to communicate the level of clinical evidence available for individual mutations in the test report. • ACT Genomics undergoes periodic updates through the review of new information by medical informatics scientists and scientific content management team | Table 7.1 Similarities between the predicate and subject devices {7}------------------------------------------------ # Table 7.2 Differences between the predicate and subject devices {8}------------------------------------------------ #### 8 Performance Testing #### 8.1 Precision / Reproducibility The objective of this study was to assess within lab precision by evaluating sources of variability (reagent lots, operators, sequencing instruments, and days). A set of 20 unique samples comprising ten cancer types containing different variants were evaluated. Twelve (12) of the samples across ten cancer types were single clinical samples (FFPE), and eight (8) of them across four cancer types were pooled DNA from multiple FFPE blocks within the same cancer type. For SNVs, MNVs, insertions, deletions, and copy number variants, all 20 samples were evaluated. For TMB, the 12 unique single FFPE samples were evaluated. The study design utilized three reagent lots, two instruments, and two operators across twelve (12) non-consecutive days. Duplicate observations for each sample were run. This resulted in 48 observations per level (3 reagent lots x 2 operators x 2 instruments x 2 start days per instrument/operator/reagent lot combination x 2 replicates). The study design resulted in 24 degrees of freedom for the replicate variability evaluation. Prior to conducting the variance component analysis, all data were visually inspected. {9}------------------------------------------------ ### 8.1.1 SNVs, Insertions and Deletions Seven hundred fifty-one (751) variants were found within the 20 samples with 638 unique variants and 247 genes. The qualitative analysis of the data was done using the correct call rate. A correct call was defined as the same mutational variant call within each of the observations of a sample with that variant. Correct calls may be positive (mutation present) or negative (wild type present). The ACTOnco system evaluated 751 variants in the assay. There were 48 observations of each variant; this resulted in 36,048 total data points (751 x 48). Of the 36,048 observations, 32,496 (677 x 48) were from SNVs, 1,008 (21 x 48) were from MNVs, 528 (11 x 48) were from insertions, and 2,016 (42 x 48) were from deletions. Since this assay has a final QC check at the variant level, there were 142 variant observations removed from the data set due to QC failing at the variant level, resulting in 35,906 variants remaining for the study analysis (36048 - 142). Across all the mutational variant data, the call rate was 98.33% (35,308/ 35,906). The breakout for each variant type was as follows: | SNV: 98.33% 31,837 / 32,377 INS: 96.97% 512 / 528 | MNV: 97.18% 963 / 991 DEL: 99.30% 1,996 / 2,010 | |------------------------------------------------------|----------------------------------------------------| |------------------------------------------------------|----------------------------------------------------| There were 15,076 WT calls across all the samples, and given 48 observations, this resulted in 723,648 (15,076 x 48) WT calls in the data set. Across all the WT data there were 99,997% correct calls (723,628/723,648). See Table 8-1. | Mutational Variant Type | Operator | Instrument | Reagent lot | Days | Number Correct | Number Attempted | Call rate (95%CI) | |----------------------------|----------|------------|----------------|------|-------------------|---------------------|--------------------------| | All | All | All | All | All | 35308 | 35906 | 98.33 (0.982-0.985) | | Deletion | All | All | All | All | 1996 | 2010 | 99.30 (0.988-0.996) | | Insertion | All | All | All | All | 512 | 528 | 96.97 (0.951-0.981) | | MNV | All | All | All | All | 963 | 991 | 97.18 (0.959-0.980) | | SNV | All | All | All | All | 31837 | 32377 | 98.33 (0.982-0.985) | | Negative (WT) | All | All | All | All | 723628 | 723648 | 99.997 (0.99996-0.99998) | Table 8-1 Correct Calls for SNVs, Insertions and Deletions These data were also evaluated at the gene level with information on the gene, the mutation, the normalized coverage range, statistics on the MAF, and the positive call rates with a 2 sided 95% confidence interval. Table 8-2 (single samples) and Table 8-3 (mixed samples) from the study are presented below. {10}------------------------------------------------ | Tissue type | Mutation type | Gene/Exon | cDNA change | Amino acid change | Mutation frequency | FDA level | |-------------------------------|------------------|----------------|---------------------------------------|----------------------|-----------------------|--------------| | colon adenocarcinoma | SNV | BRAF exon15 | 140453136 A>T | p.V600E | 29.06% | 2 | | colon adenocarcinoma | SNV | GRIN2A exon4 | 10032000 C>A | p.G275* | 50.66% | 3 | | colon adenocarcinoma | DEL | KDM5C exon23 | 53223787_CCA>C | p.C1190fs | 58.95% | 3 | | colon adenocarcinoma | INS | KMT2D exon34 | 49432235 C>CG | p.S2969fs | 24.81% | 3 | | colon adenocarcinoma | DEL | MAP3K1 exon15 | 56179358_CAG>C | p.E1225fs | 27.16% | 3 | | colon adenocarcinoma | SNV | RECQL4 exon5 | 145741926 G>A | p.R193* | 19.53% | 3 | | colon adenocarcinoma | DEL | RNF43 exon9 | 56435160 AC>A | p.G659fs | 70.63% | 3 | | colon adenocarcinoma | DEL | SPOP exon8 | 47685274 TA>T | p.F225fs | 28.11% | 3 | | colon adenocarcinoma | SNV | TP53 exon11 | 7572929 A>G | p.*394Rext*9 | 28.98% | 3 | | colon adenocarcinoma | SNV | TP53 exon5 | 7578554 A>G | p.Y126H | 19.00% | 3 | | kidney cancer | SNV | NSD1 exon19 | 176709523 C>T | p.R1984* | 8.28% | 3 | | kidney cancer | SNV | SETD2 exon3 | 47162711 T>A | p.K1139* | 19.24% | 3 | | tumor of exocrine pancreas | SNV | IDH1 exon4 | 209113112 C>G | p.R132P | 36.71% | 3 | | tumor of exocrine pancreas | SNV | KRAS exon2 | 25398284 C>T | p.G12D | 32.13% | 3 | | tumor of exocrine pancreas | SNV | TP53 exon7 | 7577539 G>A | p.R248W | 53.06% | 3 | | liver cancer | SNV | ARID2 exon5 | 46211626 A>T | p.K198* | 30.76% | 3 | | liver cancer | DEL | ARID2 exon15 | 46245542 TC>T | p.P1213fs | 16.36% | 3 | | liver cancer | SNV | TP53 exon6 | 7578224 T>A | p.R209* | 40.45% | 3 | | cholangiocarcinoma | SNV | KRAS exon2 | 25398284 C>T | p.G12D | 20.98% | 3 | | cholangiocarcinoma | SNV | TP53 exon6 | 7578190 T>C | p.Y220C | 31.86% | 3 | | colon adenocarcinoma | DEL | B2M exon2 | 45007740 GGA>G | p.R65fs | 17.84% | 3 | | colon adenocarcinoma | SNV | CTNNB1 exon3 | 41266137_C>T | p.S45F | 35.38% | 3 | | colon adenocarcinoma | SNV | ERBB2 exon19 | 37880220 T>C | p.L755S | 18.69% | 3 | | colon adenocarcinoma | SNV | GNAS exon6 | 57480483 C>T | p.R160C | 17.13% | 3 | | colon adenocarcinoma | SNV | KRAS exon2 | 25398285_C>T | p.G12S | 17.30% | 2 | | colon adenocarcinoma | SNV | MLH1 exon4 | 37045935 C>T | p.T117M | 66.43% | 3 | | colon adenocarcinoma | DEL | NBN exon10 | 90967511 CT>C | p.R466fs | 32.29% | 3 | | colon adenocarcinoma | DEL | PIK3R1 exon3 | 67569265 GC>G | p.P129fs | 16.39% | 3 | | colon adenocarcinoma | SNV | RAF1 exon7 | 12645694 A>T | p.S259T | 17.54% | 3 | | endometrial cancer | SNV | AKT1 exon4 | 105243045 A>G | p.W80R | 25.22% | 3 | | endometrial cancer | DEL | EZH2 exon10 | 148515024 TC>T | p.G395fs | 25.03% | 3 | | endometrial cancer | SNV | KRAS exon2 | 25398284 C>T | p.G12D | 40.10% | 3 | | endometrial cancer | DEL | MAP2K4 exon4 | 11998922 CA>C | p.K143fs | 24.62% | 3 | | Tissue type | Mutation type | Gene/Exon | cDNA change | Amino acid change | Mutation frequency | FDA level | | endometrial cancer | SNV | PAX5 exon2 | 37020768 A>C | p.V26G | 27.27% | 3 | | endometrial cancer | SNV | PIK3CA exon2 | 178916876 G>A | p.R88Q | 25.20% | 3 | | endometrial cancer | SNV | PIK3CA exon8 | 178928079 G>A | p.E453K | 28.15% | 3 | | endometrial cancer | DEL | TP53 exon8 | 7577143_CAGT>C | p.L265del | 27.08% | 3 | | urinary system cancer | SNV | ERBB2 exon8 | 37868208 C>A | p.S310Y | 21.48% | 3 | | endometrial cancer | SNV | ARID1A exon18 | 27101135 C>T | p.Q1473* | 44.33% | 3 | | endometrial cancer | DEL | KDM6A exon17 | 44928975 AG>A | p.A694fs | 43.77% | 3 | | endometrial cancer | SNV | PIK3CA exon2 | 178916890 C>T | p.R93W | 44.63% | 3 | | endometrial cancer | SNV | PIK3CA exon10 | 178936092 A>G | p.E545G | 45.45% | 3 | | endometrial cancer | SNV | PTEN exon5 | 89692905 G>A | p.R130Q | 88.18% | 3 | | endometrial cancer | DEL | RNF43 exon9 | 56435160 AC>A | p.G659fs | 48.78% | 3 | | Lung cancer | DEL | CDKN2A exon2 | 21971147_TGGGC TCCGCGCCGTGG A>T | p.L65fs | 18.53% | 3 | | Lung cancer | DEL | CDKN2A exon2 | 21971176 TCCGC CACTCGGGCG>T | p.S56fs | 6.41% | 3 | | Lung cancer | SNV | PIK3CA exon21 | 178952085 A>G | p.H1047R | 19.21% | 3 | | Lung cancer | INS | TP53 exon6 | 7578186 C>CT | p.P222fs | 26.73% | 3 | | Skin cancer | SNV | BRAF exon15 | 140453136 A>T | p.V600E | 52.96% | 2 | | Skin cancer | SNV | CDKN2A exon2 | 21971179 G>C | p.A60G | 46.91% | 3 | | Skin cancer | SNV | ERBB4 exon14 | 212537975 G>A | p.R544W | 24.89% | 3 | | Skin cancer | SNV | PIK3R1 exon13 | 67591106 A>G | p.K567E | 16.22% | 3 | | Skin cancer | SNV | RAC1 exon2 | 6426893 C>T | p.P29L | 34.31% | 3 | | Skin cancer | SNV | SMARCA4 exon26 | 11141519 C>T | p.Q1166* | 27.56% | 3 | Table 8-2 Correct Calls for SNVs, Insertions and Deletions by Gene (Single Samples) {11}------------------------------------------------ Table 8-3 Correct Calls for SNVs, Insertions and Deletions by Gene (Mixed Samples) | Tissue type | Mutatio n type | Gene/Exon | cDNA change | Amino Acid change | Mutation frequency | FDA level | |--------------------------|-------------------|----------------|----------------|-------------------------|-----------------------|-----------| | Lung cancer | SNV | CTNNB1 exon3 | 41266101_C>T | p.S33F | 11.72% | 3 | | Lung cancer | SNV | EGFR exon18 | 55241708_G>C | p.G719A | 10.32% | 2 | | Lung cancer | SNV | EGFR exon20 | 55249071_C>T | p.T790M | 11.41% | 2 | | Lung cancer | SNV | EGFR exon21 | 55259515_T>G | p.L858R | 11.17% | 2 | | Lung cancer | SNV | SMARCA4 exon32 | 11169037_A>T | p.K1511* | 13.94% | 3 | | Lung cancer | SNV | TP53 exon8 | 7577106_G>C | p.P278A | 11.51% | 3 | | Lung cancer | SNV | B2M exon1 | 45003746_T>C | p.M1? | 7.48% | 3 | | Tissue type | Mutation type | Gene/Exon | cDNA change | Amino Acid change | Mutation frequency | FDA level | | Lung cancer | INS | B2M exon2 | 45007890 G>GT | p.K114* | 7.43% | 3 | | Lung cancer | SNV | BRAF exon15 | 140453136 A>T | p.V600E | 12.41% | 2 | | Lung cancer | SNV | EGFR exon18 | 55241708 G>C | p.G719A | 5.41% | 2 | | Lung cancer | SNV | EGFR exon20 | 55249071 C>T | p.T790M | 6.04% | 2 | | Lung cancer | SNV | EGFR exon21 | 55259469 G>A | p.V843I | 6.40% | 2 | | Lung cancer | SNV | EGFR exon21 | 55259515 T>G | p.L858R | 6.37% | 2 | | Lung cancer | SNV | SMARCA4 exon32 | 11169037 A>T | p.K1511* | 6.99% | 3 | | Lung cancer | SNV | TP53 exon8 | 7577106 G>C | p.P278A | 5.43% | 3 | | Breast cancer | SNV | PIK3CA exon21 | 178952085 A>G | p.H1047R | 16.57% | 2 | | Breast cancer | SNV | TP53 exon7 | 7577539 G>A | p.R248W | 17.03% | 3 | | Breast cancer | SNV | PIK3CA exon21 | 178952085 A>G | p.H1047R | 11.32% | 2 | | Breast cancer | SNV | TP53 exon7 | 7577539 G>A | p.R248W | 10.29% | 3 | | Breast cancer | SNV | TP53 exon7 | 7577556 C>T | p.C242Y | 33.94% | 3 | | Skin cancer | SNV | ARID1B exon3 | 157222648 C>T | p.Q626* | 5.96% | 3 | | Skin cancer | SNV | BRAF exon15 | 140453136 A>T | p.V600E | 7.95% | 2 | | Skin cancer | SNV | BRAF exon11 | 140481397 C>A | p.V471F | 20.06% | 3 | | Skin cancer | SNV | CARD11 exon6 | 2979559 C>T | p.D230N | 19.56% | 3 | | Skin cancer | SNV | FANCA exon27 | 89833576 G>C | p.S858R | 7.10% | 3 | | Skin cancer | SNV | NF1 exon12 | 29533315 C>T | p.R440* | 23.72% | 3 | | Skin cancer | SNV | NOTCH4 exon4 | 32188899 G>A | p.Q219* | 5.58% | 3 | | Skin cancer | SNV | NRAS exon3 | 115256529 T>C | p.Q61R | 8.71% | 2 | | Skin cancer | DEL | TP53 exon4 | 7579546 CG>C | p.P47fs | 6.82% | 3 | | Skin cancer | SNV | BRAF exon15 | 140453136 A>T | p.V600E | 2.02% | 2 | | Skin cancer | SNV | BRAF exon11 | 140481397 C>A | p.V471F | 5.56% | 3 | | Skin cancer | MNV | DNMT3A exon16 | 25466800 GG>AA | p.R635W | 26.56% | 3 | | Skin cancer | SNV | EZH2 exon16 | 148508728 A>T | p.Y646N | 24.10% | 3 | | Skin cancer | SNV | NF1 exon12 | 29533315 C>T | p.R440* | 7.87% | 3 | | Skin cancer | SNV | NRAS exon3 | 115256529 T>A | p.Q61L | 28.18% | 2 | | Skin cancer | SNV | NRAS exon3 | 115256529 T>C | p.Q61R | 2.31% | 2 | | Skin cancer | SNV | TP53 exon8 | 7577099 C>T | p.R280K | 36.85% | 3 | | urinary system cancer | INS | ARID1A exon9 | 27092804 A>AC | p.Q944fs | 12.90% | 3 | | urinary system cancer | SNV | RXRA exon10 | 137328351 C>T | p.S427F | 9.84% | 3 | | urinary system cancer | INS | ARID1A exon9 | 27092804 A>AC | p.Q944fs | 8.05% | 3 | | Tissue type | Mutatio n type | Gene/Exon | cDNA change | Amino Acid change | Mutation frequency | FDA level | | urinary system cancer | SNV | RXRA exon10 | 137328351_C>T | p.S427F | 6.52% | 3 | {12}------------------------------------------------ {13}------------------------------------------------ The positive and negative call rates for sequence mutations (SNVs, MNVs, insertions and deletions) in each sample are summarized in Table 8-7. Call rates based on the total number of mutations along with the 2-sides 95% confidence interval were calculated. Table 8-4 and Table 8-5 summarize the results of positive call rate in per clinical sample (single and mixed). Table 8-6 and Table 8-7 summarize the results of negative call rate in per clinical sample (single and mixed). Table 8-4 Positive call rate per sample (Single sample) | Sample ID | Total No unique mutations detected across all 48 replicates | Positive Call Rate per Mutation | Positive call rate (two-sided 95% CI) | |-----------|-------------------------------------------------------------------|------------------------------------|------------------------------------------| | Sample 1 | 66 | 28/48 for 1 | 3143/3166 99.27% (98.91%, 99.51%) | | | | 45/48 for 1 | | | | | 46/46 for 1 | | | | | 48/48 for 63 | | | Sample 2 | 34 | 48/48 for 34 | 1632/1632 100% (99.76%, 100%) | | Sample 3 | 14 | 48/48 for 14 | 672/672 100% (99.43%, 100%) | | Sample 4 | 52 | 48/48 for 52 | 2496/2496 100% (99.84%, 100%) | | Sample 5 | 17 | 48/48 for 17 | 816/816 100% (99.53%, 100%) | | Sample 6 | 45 | 46/46 for 1 | 2156/2157 99.95% (99.73%, 99.99%) | | | | 46/47 for 1 | | | | | 48/48 for 43 | | | Sample 7 | 35 | 47/47 for 1 | 1679/1679 100% (99.77%, 100%) | | | | 48/48 for 34 | | | Sample 8 | 13 | 5/47 for 1 | 581/623 93.25% (91.01%, 94.97%) | | | | 48/48 for 12 | | | Sample 9 | 49 | 24/48 for 1 | 2293/2352 97.49% (96.77%, 98.05%) | | | | 32/48 for 1 | | | | | 33/48 for 1 | | | | | 46/48 for 2 | | | | | 48/48 for 44 | | | Sample 10 | 11 | 48/48 for 11 | 528/528 100% (99.27%, 100%) | | Sample 11 | 16 | 34/46 for 1 | 754/766 98.43% (97.28%, 99.1%) | | | | 48/48 for 15 | | {14}------------------------------------------------ | Sample ID | Total No unique mutations detected across all 48 replicates | Positive Call Rate per Mutation | Positive call rate (two-sided 95% CI) | |-----------|-------------------------------------------------------------------|--------------------------------------------|------------------------------------------| | Sample 12 | 34 | 20/20 for 1 24/24 for 1 48/48 for 32 | 1580/1580 100% (99.75%, 100%) | * MAF = mutational allele frequency, CO = cut-off Table 8-5 Positive call rate per sample (Mixed Sample) | Sample ID | Total No unique mutations detected across all 48 replicates | Positive Call Rate per Mutation | Positive call rate (two-sided 95% CI) | |-----------|-------------------------------------------------------------------|-----------------------------------------------------------------------------------------------------------------------------------------------------------------------------------|------------------------------------------| | Sample 13 | 30 | 43/48 for 1 48/48 for 29 | 1435/1440 99.65% (99.19%, 99.85%) | | Sample 14 | 52 | 21/48 for 1 26/48 for 1 29/48 for 1 36/48 for 1 39/48 for 1 42/48 for 1 43/48 for 2 44/48 for 1 45/48 for 1 46/48 for 3 47/48 for 3 48/48 for 36 | 2375/2496 95.15% (94.24%, 95.93%) | | Sample 15 | 22 | 48/48 for 22 | 1056/1056 100% (99.64%,100%) | | Sample 16 | 32 | 46/48 for 1 47/48 for 1 48/48 for 30 | 1533/1536 99.8% (99.43%, 99.93%) | | Sample 17 | 78 | 26/48 for 2 31/48 for 1 37/47 for 1 37/48 for 1 38/48 for 1 39/48 for 1 44/44 for 1 46/48 for 4 47/47 for 4 47/48 for 3 48/48 for 59 | 3623/3735 97% (96.4%, 97.5%) | {15}------------------------------------------------ | Sample ID | Total No unique mutations detected across all 48 replicates | Positive Call Rate per Mutation | Positive call rate (two-sided 95% CI) | |-----------|-------------------------------------------------------------------|------------------------------------|------------------------------------------| | Sample 18 | 81 | 23/48 for 1 | 3786/3883 97.5% (96.96%, 97.95%) | | | | 36/48 for 1 | | | | | 37/48 for 1 | | | | | 39/48 for 1 | | | | | 40/48 for 2 | | | | | 41/48 for 1 | | | | | 43/47 for 1 | | | | | 45/45 for 1 | | | | | 45/48 for 1 | | | | | 46/47 for 1 | | | | | 46/48 for 2 | | | | | 47/48 for 5 48/48 for 63 | | | Sample 19 | 32 | 39/45 for 1 | 1503/1510 99.54% (99.05%, 99.78%) | | | | 42/42 for 1 | | | | | 47/48 for 1 | | | | | 31/31 for 1 | | | | | 48/48 for 28 | | | Sample 20 | 38 | 3/40 for 1 | 1667/1783 93.49% (92.25%, 94.55%) | | | | 17/17 for 1 | | | | | 21/48 for 1 | | | | | 33/48 for 1 | | | | | 39/48 for 1 | | | | | 42/48 for 1 | | | | | 43/48 for 2 | | | | | 44/48 for 1 | | | | | 45/48 for 1 | | | | | 46/46 for 1 | | | | | 46/48 for 3 | | | |…