Learning Objectives Case
4/20/2015
New HIV Tests and Algorithm: A change we can believe in
Esther Babady, PhD, D (ABMM) Memorial Sloan-Kettering Cancer Center New York, New York
Memorial Sloan-Kettering Cancer Center Learning Objectives
After this presentation you should be able to: List new methods used for diagnosis of HIV infections Explain the advantages of new methods in diagnosis of HIV infections Describe the most recent CDC/APHL laboratory testing recommendations for HIV diagnosis
Memorial Sloan-Kettering Cancer Center Case
68 yo woman Developed a large parotid mass on neck Radiologic studies: Abnormal Fine needle aspirate of parotid Enlarged, atypical cells and abundant necrosis Suggestive of B cell lymphoma
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Memorial Sloan-Kettering Cancer Center Case
Non-Hodgkin’s Lymphoma confirmed Pre-chemotherapy Infectious Diseases Evaluation Hepatitis serologies HIV serologies
Memorial Sloan-Kettering Cancer Center Case
Tests Results Reference range Hepatitis B surface antigen Negative Negative Hepatitis B surface antigen Positive Positive* antibody Hepatitis B core antibody (Total) Positive Negative Hepatitis B core antibody IgM Negative Negative Hepatitis B quantitative PCR Negative [20-170,000,000 IU/mL] Hepatitis C antibody Negative Negative HIV-1 and HIV-2 antibodies Negative Negative
Memorial Sloan-Kettering Cancer Center Case
Patient admitted 1 week later Started on chemotherapy protocol
Tests Results Reference range HIV-1 and HIV-2 Negative Negative Testsantibodies Results Reference range HIV-1 quantitative PCR 120,000 [20-10,000,000 copies/mL]
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Memorial Sloan-Kettering Cancer Center
http://www.cdc.gov/hiv/default.html
Memorial Sloan-Kettering Cancer Center Background: The virus
Enveloped positive-sense ss RNA virus Retroviridae Lentivirus ~100 nm diameter 9.7 kb genome 5’ and 3’ LTR, 9-10 genes
http://www.niaid.nih.gov/factsheets/graphics/howhiv.jpg
Memorial Sloan-Kettering Cancer Center Background: HIV Phylogeny
Groups Subtypes Major (M) A, B, C, D, E,F, G, H, J, K
Non M/O (N) HIV-1 Outlier (O)
“Pending” (P)
Circulating recombinant forms
A
HIV-2 B
C,D,E,F,G,H
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Memorial Sloan-Kettering Cancer Center Background: HIV Distribution
Subtypes Geography Subtype A West Africa Subtype B North America and Europe Subtype C South Africa and S/E Asia Subtype D Eastern and central Africa Subtype F Central Africa, South America and Eastern Europe Subtype G Africa and Central Europe Subtype H Central Africa Subtype J Africa and Caribbean Subtype K DRC and Cameroon HIV-2 West Africa
McCutchan, F.E. JMV 2006
Memorial Sloan-Kettering Cancer Center Background: Stages of Infection
Acute Established AIDS Flu-like symptoms Mild or No Low CD4 count Highly infectious symptoms Opportunistic Weeks Infectious infections Years Years
Naif, H.M. Infect Dis Rep. 2013
Memorial Sloan-Kettering Cancer Center Background: Epidemiology
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Memorial Sloan-Kettering Cancer Center Background: Epidemiology
Memorial Sloan-Kettering Cancer Center Trends in Annual Age-Adjusted* Rate of Death Due to HIV Infection, United States, 1987−2010
Note: For comparison with data for 1999 and later years, data for 1987−1998 were modified to account for ICD-10 rules instead of ICD-9 rules. *Standard: age distribution of 2000 US population
Memorial Sloan-Kettering Cancer Center Stage 3 (AIDS) Classifications and Deaths of Persons with HIV Infection Ever Classified as Stage 3 (AIDS), among Adults and Adolescents, 1985–2012—United States and 6 Dependent Areas
Note. All displayed data have been statistically adjusted to account for reporting delays, but not for incomplete reporting. Deaths of persons with HIV infection, stage 3 (AIDS) may be due to any cause.
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Memorial Sloan-Kettering Cancer Center Need for new recommendations?
1989 2015
Memorial Sloan-Kettering Cancer Center Need for new recommendations?
Better tests Simpler tests More commercially available tests More treatment options
Memorial Sloan-Kettering Cancer Center Laboratory testing sequence
CDC and APHL. Laboratory Testing for the Diagnosis of HIV Infection: Updated Recommendations. June 27, 2014. Accessed [04/2015]
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Memorial Sloan-Kettering Cancer Center
CDC and APHL. Laboratory Testing for the Diagnosis of HIV Infection: Updated Recommendations. June 27, 2014. Accessed [04/2015]
Memorial Sloan-Kettering Cancer Center Laboratory testing: Immunoassays
Utility: Screening Diagnostics Additional testing: Acute vs Established
tat 5’LTR gag pol vif/vpr/tat env 3’LTR rev/vpu nef
Memorial Sloan-Kettering Cancer Center Laboratory Testing: Immunoassays
1st generation HIV-1 IgG ab Viral culture cell lysates
Viral culture cell lysates 2nd generation HIV-1/2 IgG ab Synthetic/Recombinant ag
3rd generation HIV-1/2 IgM/IgG ab Synthetic/Recombinant ag
HIV-1/2 IgM/IgG ab 4th generation Synthetic/Recombinant ag p24 ag
CDC and APHL. Laboratory Testing for the Diagnosis of HIV Infection: Updated Recommendations. June 27, 2014. Accessed [04/2015]
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Memorial Sloan-Kettering Cancer Center Laboratory Testing: Western blot
Cambridge Biotech HIV-1 Western Blot Kit PI
Memorial Sloan-Kettering Cancer Center Laboratory Testing: Rapid HIV tests
Manufacturer Rapid test: Test CLIA waived Methods Target Date Alere Alere Lateral flow HIV-1 2013 Scarborough Determine™ HIV- Immunoassay HIV-2 1/2 Ag/Ab Combo p24 Bio-Rad Geenius HIV 1/2 Immuno- HIV-1 2014 Laboratories, Supplemental chromatograph HIV-2 Inc Assay
Accessed 4/2015 http://www.fda.gov/downloads/BiologicsBloodVaccines/BloodBloodProducts/ApprovedProducts/PremarketApprovalsPMAs/UCM149971.pdf
Memorial Sloan-Kettering Cancer Center Laboratory Testing: Rapid HIV tests
Manufacturer Test Methods Target Date Bio-Rad Multispot HIV- Immuno- HIV-1 2004 Laboratories, 1/HIV-2 Rapid concentration HIV-2 Inc. Test Microparticle OraSure OraQuick Lateral flow HIV-1 2002 Technologies ADVANCE® Immunoassay HIV-2 Rapid HIV-1/2 Antibody
Accessed 4/2015 http://www.fda.gov/downloads/BiologicsBloodVaccines/BloodBloodProducts/ApprovedProducts/PremarketApprovalsPMAs/UCM149971.pdf
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Memorial Sloan-Kettering Cancer Center Laboratory Testing: Immunoassays
Manufacturer Test Methods Generation Date cleared Abbott Architect HIV Chemiluminescence 4th 2010 Diagnostics Ag/Ab combo Microparticle Immunoassay Bio-Rad GS HIV Enzyme 4th 2011 Laboratories, Combo Ag/Ab Immunoassay Inc. Ortho Clinical Vitros Anti-HIV Chemiluminescence 3rd 2008 Diagnostics 1+2 Immunoassay Siemens Advia Centaur Chemiluminescence 3rd 2006 Healthcare HIV 1/0/2 Immunoassay diagnostics Accessed 4/2015 http://www.fda.gov/downloads/BiologicsBloodVaccines/BloodBloodProducts/ApprovedProducts/PremarketApprovalsPMAs/UCM149971.pdf
Memorial Sloan-Kettering Cancer Center Laboratory Testing: Molecular tests
Utility Diagnosis of HIV infection (Qualitative) Baseline viral load before initiation of treatment (Quantitative) Monitoring of response to antiviral treatment (Quantitative)
tat 5’LTR gag pol vif/vpr/tat env 3’LTR rev/vpu nef
Memorial Sloan-Kettering Cancer Center Laboratory Testing: Molecular tests
Examples of Qualitative Molecular Methods
Manufacturer Test Methods LOD Hologic Aptima HIV-1 RNA TMA 100 qualitative assay HPA copies/mL
Accessed 4/2015 http://www.fda.gov/downloads/BiologicsBloodVaccines/BloodBloodProducts/ApprovedProducts/PremarketApprovalsPMAs/UCM149971.pdf
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Memorial Sloan-Kettering Cancer Center Laboratory Testing: Molecular tests
Manufacturer Test Methods Linear range Abbott Molecular Realtime HIV-1 RT-PCR 40-10,000,000 copies/mL Roche Molecular Cobas Ampliprep/Cobas RT-PCR 20-10,000,000 copies/mL Taqman quantitative HIV-1 v.2.0 COBAS® TaqMan® HIV Test RT-PCR 34- 10,000,000 copies/mL v2.0 For Use With The High Pure System Cobas Amplicor HIV Monitor RT-PCR 50- 75,000 copies/ml v1.5 400- 750,000 copies/ml Siemens VERSANT HIV 3.0 Assay bDNA signal 75 -500,000 copies/mL (bDNA) amplification
Accessed 4/2015 http://www.fda.gov/downloads/BiologicsBloodVaccines/BloodBloodProducts/ApprovedProducts/PremarketApprovalsPMAs/UCM149971.pdf
Memorial Sloan-Kettering Cancer Center Laboratory Testing: Molecular Tests
Highly specific and excellent sensitivity
8
7 ULOQ
6
5
4
3
2 LLOQ
1 LOD
0
Expected viral loads log IU/mL viralExpected loads log 012345678 Measured viral loads log IU/mL
Memorial Sloan-Kettering Cancer Center Laboratory Testing: Studies
Comparison of WB to a rapid antigen test to on specimens reactive by a 4th generation IA Philadelphia, PA Accuracy Rapid antigen test vs WB: 98.2% vs. 57.9% Specificity Rapid antigen test vs WB: HIV-2 misidentified as HIV-1 by WB.
Cardenas, A.M. et al., Journal of Clinical Virology 58S1 (2013), e97–e103
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Memorial Sloan-Kettering Cancer Center Laboratory Testing: Studies
Comparison of a rapid antigen test to WB, 3rd and 4th generation immunoassays. Commercial panels (HIV-1 and HIV-2) Seroconversion Rapid antigen test: 7-15 days earlier than WB. Accuracy Rapid antigen test was reactive in 99.0% and 92.5% of 3rd and 4th generation
Masciotra, S. et al., Journal of Clinical Virology 58S (2013) e54– e58
Memorial Sloan-Kettering Cancer Center Laboratory Testing: Studies
4th generation HIV Ab/Ag, WB and HIV-1 RNA HIV treatment center: Amsterdam, Netherlands ~5,000/years over 10 years: 75 cases Ab/Ag positive, WB negative, HIV-1 RNA not detected 7 cases Ab/Ag positive, WB negative, HIV-1 RNA detected 6 cases Ab/Ag negative, WB negative, HIV-1 RNA detected Sensitivity/specificity: 99.6%/99.8%.
Jurriaans, S. et al., Journal of Clinical Virology 52S (2011), S67-S69
Memorial Sloan-Kettering Cancer Center Laboratory Testing: Studies
Comparison of 3rd and 4th generation IA Multi sites study: CA and WA Seroconversion 4th generation vs 3rd generation: 7-11 days earlier Specificity 4th generation vs 3rd generation: 99.7-100%
Mitchell, E.O. et al., Journal of Clinical Virology 58S1 (2013), e79-e84
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Memorial Sloan-Kettering Cancer Center Laboratory Testing: Studies
Comparison of WB to RAT and NAAT. 11 U.S. public health laboratories 570 remnant 3rd gen IA reactive, HIV WB neg or indeterminate. ~90% confirmed HIV negative ~10% HIV positive (~3% acute HIV and 0.2% HIV-2).
Linley, L. et al., Journal of Clinical Virology 58S1 (2013), e108-e112
Memorial Sloan-Kettering Cancer Center Laboratory testing: Genotyping
Utility: Monitoring HIV-1 resistance Baseline before start of antiretroviral therapy (ART) Guide alternative ART selection in cases of drug therapy failure
Memorial Sloan-Kettering Cancer Center Laboratory Testing: Genotyping
Examples of Genotyping assays
Manufacturer Test Methods LOD Target Abbott Molecular Viro Seq HIV-1 Sequencing 2000 copies /mL Protease sequencing system RT Siemens Molecular TRUGENE HIV -1 Sequencing 1000 copies/mL Diagnostics Genotyping Assay
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Memorial Sloan-Kettering Cancer Center U.S. Preventive Services Task Force
http://www.uspreventiveservicestaskforce.org/Page/Topic/recommendation-summary/human-immunodeficiency-virus-hiv-infection-screening
Memorial Sloan-Kettering Cancer Center CDC HIV Testing Guidelines
1989: Testing for HIV-1 immunoassays and confirmation with HIV-1 WB or IFA (MMWR v38, 1989) 1992: Testing for HIV-2 antibodies (MMWR v41, 1992) 2004: Use of HIV-1 WB/IFA to confirm reactive rapid test results (MMWR v53, 2004)
Memorial Sloan-Kettering Cancer Center CDC HIV Screening Guidelines
2014: Recommended algorithm Initial testing for HIV should be done using 4th generation immunoassay Reactive screen should be confirmed with an Ab test that differentiates HIV-1 and HIV-2 If reactive samples do not confirm, additional testing with a HIV-1 NAT Algorithm should be applied to reactive rapid tests
CDC and APHL. Laboratory Testing for the Diagnosis of HIV Infection: Updated Recommendations. June 27, 2014. Accessed [04/2015]
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Memorial Sloan-Kettering Cancer Center CDC HIV Screening Guidelines
2014: Alternative algorithms Reactive 3rd generation immunoassay followed by recommended algorithm Reactive WB/IFA followed by HIV-1 NAT and HIV-2 ab test if necessary Reactive 4th generation IA followed by HIV-1 NAT Nonreactive 3rd and 4th generation IA followed by HIV-1 NAT
CDC and APHL. Laboratory Testing for the Diagnosis of HIV Infection: Updated Recommendations. June 27, 2014. Accessed [04/2015]
Memorial Sloan-Kettering Cancer Center New Testing algorithm
CDC and APHL. Laboratory Testing for the Diagnosis of HIV Infection: Updated Recommendations. June 27, 2014. Accessed [04/2015]
Memorial Sloan-Kettering Cancer Center Reporting results
CDC guidelines, 2014
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Memorial Sloan-Kettering Cancer Center Conclusions
Several, new, sensitive and user-friendly assays are now available for accurate diagnosis of HIV
Recent CDC/APHL recommendations use combinations of these new methods to identify more cases of acute HIV infections
Memorial Sloan-Kettering Cancer Center Self-Assessment Questions
1. The new CDC/APHL recommends that a reactive 4th generation immunoassay be followed by testing with which of the following tests? A. HIV-1/HIV-2 ab C. Western Blot B. HIV-1 RNA D. HIV-2 RNA
2. Which of the following HIV antigens is detected by antibody/antigen combo assays? A. p24 C. gp120 B. p55 D. gp160
Memorial Sloan-Kettering Cancer Center Self-Assessment Questions
3. Use of the new CDC/APHL HIV testing algorithm improves detection of acute HIV? A. True B. False
4. Which of the following test is reactive during the eclipse period of HIV infection? A. qualitative HIV-1 RNA C. Both of the above B. 4th generation IA D. None of the above
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