WHO Prequalification of Diagnostics Programme PUBLIC REPORT. Product: VERSANT HIV-1 RNA 1.0 Assay (kpcr) Number: PQDx

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1 WHO Prequalification of Diagnostics Programme PUBLIC REPORT Product: VERSANT HIV-1 RNA 1.0 Assay (kpcr) Number: PQDx Abstract The VERSANT HIV-1 RNA 1.0 Assay (kpcr) with product codes , , , and , manufactured by Siemens Healthcare Diagnostics, 511 Benedict Avenue, Tarrytown, NY, USA, CE-marked regulatory version, was accepted for the WHO list of prequalified diagnostics and was listed on 13 February The VERSANT HIV-1 RNA 1.0 Assay (kpcr) is an in vitro nucleic acid amplification assay for the quantitation of human immunodeficiency virus type 1 (HIV-1) RNA in plasma from HIV- 1 infected individuals, over the range of 37 to 11,000,000 copies/ml, using the VERSANT kpcr Molecular System. The assay is intended to be used in conjunction with clinical presentation and other laboratory markers of disease status, as an aid in the management of individuals infected with HIV-1. Monitoring HIV-1 RNA serial results from the assay can be used to assess the prognosis of disease progression, and as an aid in assessing viral response to antiretroviral treatment measured by changes in plasma HIV-1 RNA levels. The VERSANT HIV-1 RNA 1.0 Assay (kpcr) is a reverse transcription (RT) kinetic polymerase chain reaction (kpcr) procedure for quantifying HIV-1 RNA in human plasma. The assay releases HIV-1 RNA from the viral capsid in a solution containing chaotropic buffer and magnetic silica beads. The released HIV-1 RNA, along with other nucleic acids and the added RNA Internal Control (IC), bind to the beads and a series of washes removes the remaining plasma components. The purified RNA is eluted and added to a PCR plate containing HIV-1 Primer/Probe Mix and HIV-1 Enzyme Mix, and the wells are sealed. The HIV-1 Primer/Probe Mix contains buffer, dntps and synthetic DNA primers and probes specific for HIV-1 and the IC. These are combined with the purified RNA and the Enzyme Mix to enable the RT and kpcr reactions. The Enzyme Mix contains Uracil DNA Glycosylase (UNG) for contamination control. The HIV-1 and IC RNA are reverse transcribed to make cdna. Both targets (the HIV-1 and IC) are then simultaneously amplified and detected using kpcr methodology. The probes include fluorescent dyes and quenchers as modifiers. The dual-labeled probes specifically detect the presence of HIV-1 and IC amplicons during amplification. In their native state, the probes adopt a folded structure, positioning the quencher next to the 1 See page 2 for a list of components required to perform the assay. PQDx_107 Page 1 of 55

2 fluorescent dye. In this condition, most of the fluorescence of the dye is absorbed by the neighboring quencher, minimizing the emitted fluorescence. When amplicons are generated, fluorescent dye-labeled probes uncoil as they hybridize to the amplicons, separating the fluorescent dye from the quencher, thereby increasing the observed fluorescence. During the extension step, some of the hybridized probe is cleaved through the exonuclease activity of the polymerase. When free in solution, the fluorescence is continuously detectable. The combined fluorescence of both cleaved and bound probes correlates with the amount of amplicons generated, and is proportional to the amount of HIV-1 RNA in the sample. Refer to Figure 1 for a schematic representation of the Assay Principle. The VERSANT HIV-1 RNA 1.0 Assay (kpcr) is not intended for use as a donor screening assay for HIV-1 or as a diagnostic assay to confirm the presence of HIV-1 infection. In order to perform the assay, the following components are required: Instrumentation: VERSANT kpcr Molecular System Includes the following: o VERSANT kpcr Molecular System SP with Heater/Shaker, o VERSANT kpcr Molecular System SP Workstation w/ Barcode Scanner, o VERSANT kpcr Molecular System AD, o VERSANT kpcr Molecular System AD Workstation w/ Barcode Scanner, o VERSANT kpcr Molecular System SP System Software v1.1 CD, o VERSANT kpcr Molecular System AD System Software v1.1cd, o VERSANT kpcr Molecular System Test Definitions Non-US v1.1 CD, o VERSANT kpcr Molecular System On-Line Help v1.1 CD, o VERSANT kpcr Molecular System Software v1.1 Installation Kit, Reagents: VERSANT Sample Preparation 1.0 Reagents, IVDD (Box 1), VERSANT Sample Preparation 1.0 Reagents, IVDD (Box 2), VERSANT HIV-1 RNA Assay 1.0 (kpcr), IVDD (Box 1), VERSANT HIV-1 RNA Assay 1.0 (kpcr) IVDD (Box 2), Specific consumables: Large reagent troughs and small reagent troughs, REF μL pipette tips, REF μL pipette tips, REF well, 2-mL nuclease free, sterile deep well plates, REF Barcoded 96-well semi-skirted polypropylene plates for PCR, REF PQDx_107 Page 2 of 55

3 Optical caps, 8x strip, REF Generic consumables and general laboratory equipment: Adjustable micropipette capable of dispensing 1000 μl 1000-μL disposable, aerosol-resistant pipette tips, sterile-packaged Disposable, polypropylene sample tubes 10-mL to 25-mL serological pipettes, sterile-packaged Vortex mixer Quick-spin, bench-top microcentrifuge Bleach, unscented (0.5% sodium hypochlorite) Microcide SQ Deionized water Storage: The VERSANT HIV-1 RNA Assay 1.0 (kpcr), IVDD (Box 1), must be stored at -30 to -10 C. The VERSANT HIV-1 RNA Assay 1.0 (kpcr) IVDD (Box 2), must be stored at -90 to -60 C. The VERSANT Sample Preparation 1.0 Reagents, IVDD (Box 1), must be stored at 15 to 30 C. The VERSANT Sample Preparation 1.0 Reagents, IVDD (Box 2), must be stored at 2 to 8 C. Shelf-life: VERSANT HIV-1 RNA Assay 1.0 (kpcr), IVDD (Box 1), : 12 months VERSANT HIV-1 RNA Assay 1.0 (kpcr) IVDD (Box 2), : 12 months VERSANT Sample Preparation 1.0 Reagents, IVDD (Box 1), : 24 months VERSANT Sample Preparation 1.0 Reagents, IVDD (Box 2), : 24 months Summary of prequalification status for the VERSANT HIV-1 RNA 1.0 Assay (kpcr) Initial acceptance Date Outcome Status on PQ list 13 February 2012 listed Dossier assessment 12 December 2011 MR Inspection status 23 November 2011 MR Laboratory evaluation FT MR MR: Meets Requirements, NA: Not Applicable, FT: Fast-tracked PQDx_107 Page 3 of 55

4 The VERSANT HIV-1 RNA 1.0 Assay (kpcr) was accepted for the WHO list of prequalified diagnostics on the basis of data submitted and publicly available information. Background information Siemens Healthcare Diagnostics submitted an application for prequalification of the VERSANT HIV-1 RNA 1.0 Assay (kpcr). Based on the established prioritization criteria, the VERSANT HIV-1 RNA 1.0 Assay (kpcr) was given priority for prequalification. Product dossier assessment Siemens Healthcare Diagnostics submitted a product dossier for the VERSANT HIV-1 RNA 1.0 Assay (kpcr) as per the Instructions for compilation of a product dossier (PQDx_018 v1). The information submitted in the product dossier was reviewed by WHO staff and external experts (assessors) appointed by WHO in accordance with the internal report on the screening and assessment of a product dossier (PQDx_009 v2). Based on the product dossier screening and assessment findings, a recommendation was made to accept the product dossier for the VERSANT HIV-1 RNA 1.0 Assay (kpcr) for prequalification. Commitments for prequalification: The manufacturer committed to amend and submit additional documentation on the following issues: 1. an updated version of the risk analysis and control summary 2. shipping stability 3. independent clinical evaluation. Manufacturing site inspection An inspection was performed at the site of manufacture (333 Coney St., East Walpole, MA , USA) of the on 23, 24 September 2011 as described in Information for manufacturers on WHO prequalification inspection procedures for the sites of manufacture of diagnostics (PQDx_014 v1). The fast track inspection found that the manufacturer had a well-established quality management system and manufacturing practices in place that would ensure the manufacture of a product of consistent quality. The manufacturer has committed to respond to the observations and nonconformities identified during the inspection. Commitments for prequalification: 1. Risk assessment will be reconsidered to include risk analysis and mitigation for end users in resource limited and environmentally challenging regions to which the PQDx_107 Page 4 of 55

5 product is distributed. This is of particular importance given the deep freeze storage requirements of some of the kit components. 2. Testing for preservation of product during use and to intended destination will be extended. 3. Siemens Healthcare Diagnostics, Inc. will move towards improving customer feedback mechanisms from representative organizations of users in resource limited regions where communication may be problematic. Laboratory evaluation Given the regulatory version of the product submitted for prequalification and the quality of the data submitted as part of the product dossier to support the claims for its intended use, the the VERSANT HIV-1 RNA 1.0 Assay (kpcr) assay has been found eligible to undergo the WHO fast track procedure. Subsequently, the product will not be required to undergo a laboratory evaluation for its use with human plasma. Nevertheless, taking into consideration the needs of WHO Member States and in an effort to support current attempts to increase access to HIV Viral Load testing, a laboratory evaluation of the the VERSANT HIV-1 RNA 1.0 Assay (kpcr) will be carried out to assess its performance with the use of Dried Blood Spots (DBS). It is acknowledged that this constitutes an off-label use of the product and therefore, results from the evaluation will not impact the WHO Prequalification status for use with human plasma. The results from the evaluation will serve as a source of information in order to advise WHO Member States in their efforts to scale up HIV Viral Load Testing. PQDx_107 Page 5 of 55

6 Labelling 1. Labels 2. Instructions for use PQDx_107 Page 6 of 55

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