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 1 4/20/2015 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] 2 4/20/2015 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 3 4/20/2015 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 4 4/20/2015 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. 5 4/20/2015 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] 6 4/20/2015 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] 7 4/20/2015 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 8 4/20/2015 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 9 4/20/2015 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 10 4/20/2015 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: