CD4 for monitoring HIV Progression

1. CD4 for monitoring HIV Progression 31st Annual National Conference of Indian Association of Preventive & Social Medicine, 27th – 29th Feb 2004 PGIMER, Chandigarh Dr. Arvinder Singh BD Biosciences

2. China -CDC -MOH -BDB BD Bio-IS -Value Proposition -Education and training -QC/QA protocols -Application protocols -Reference range studies India -ICMR -NACO -Hospitals / Path labs -BDB Africa -WHO -Countries -DOD -NGOs -BDB Thailand -MOH -COE -BDB Other Countries -WHO -NGOs -BDB GOVT -NIH -WHO -HRF -UNAID -DOD -USAID NGOs -AAACP -MSF -IAVI -WJCF -HVTN -AMFAR -JHU -GMHC BD Activities Brazil -COOPEX -MOH -BDB

3. HIV/AIDSWhat is CD4/CD8

6. Flow Cytometry Clinical Applications • HIV immunophenotyping (CD4 absolute counts) • Cytokine analysis • Leukemia and lymphoma diagnosis - minimum residual disease detection • Cell cycle and DNA ploidy analysis • Stem Cell Analysis • HLA – B 27 Typing • Residual white blood cell detection • Reticulocyte enumeration • Flow crossmatching (organ transplantation)

7. What Is Flow Cytometry? A technology that simultaneously measures multiple characteristics of single cells at a rapid rate

9. 1000 800 600 400 200 0 0 200 400 600 800 1000 Lysed Whole Blood Neutrophils Side Scatter Monocytes Lymphocytes Forward Light Scatter

12. Two-Color Cell Analysis 104 10 3 CD8 PE 10 2 101 10 0 100 101 102 103 10 4 CD4 FITC

14. Fourcolor analysis CD8 PE SSC Height CD45 PerCP CD3 FITC CD8 PE CD4 Cy5 CD3 FITC CD4 Cy5

15. 42 Million People Living with AIDS

16. India – Current Situation HIV • HIV prevalence rate is 0.7% • India has had a sharp increase in the estimated number of HIV infections, from a few thousand in the early 1990s to a working estimate of about 3.8 million children and adults living with HIV/AIDS in 2001 • Estimates range from 8 - 15 million HIV infected adults and children living with HIV in 2010 • Given India's large population, a mere 0.1 percent increase in the prevalence rate would increase the number of adults living with HIV/AIDS by over half a million people

17. India – Current Situation HIV • About 90% of the total reported AIDS cases occur in the sexually active and economically productive 15 to 44 age group • Men account for 79% of HIV infections in India • The predominant mode of HIV transmission is through heterosexual contact • In 2001, the HIV infection rate went above one per cent in six states • Maharashtra, Tamil Nadu and Manipur - account for 75% of estimated HIV cases • the burden of AIDS cases is beginning to be felt in states affected early - Mumbai and Manipur have recorded 20 to 49 per cent bed occupancy by HIV positive people in certain hospitals

18. HIV/AIDS Pandemic Disease Management Vaccines, Drugs, Education • Antiretroviral Therapy (ART) Controls disease progression Increasing drug resistance patterns Expensive (Big Pharma – western countries) Trend to affordable (generic) drugs in developing world • CD4 T-Lymphocyte Count Flow cytometry is gold standard Alternate technologies – currently inadequate FacsCount is considered system of choice CD4 count is the single most important objective criteria to guide clinical decision-making on when to use ART

19. HIV Testing paradigm Screen Confirm* CD4 Viral load Est. annual tests 15-20M 4-6M 5-8M 1-2M *(includes genotyping)

20. PCR Viral load It is a technique that imitates a cell’s ability to replicate DNA by generating multiple copies of specific sequences of DNA through amplification. In clinical diagnostics, a specimen of genetic material can be repeatedly copied by PCR to provide sufficient material to detect the presence or absence of a virus as well as to quantitate its levels in the blood. It’s used for: • The development of new drugs • To establish a patient’s disease status and the likelihood of disease progression • To determine when to start treatment • To help the physician decide which drugs to use in a treatment regimen • To assess whether a treatment regimen is working

21. CD4 and viral load

22. HIV/AIDS Pandemic Situational Analysis Unmet Needs • Affordable drugs/ART •   Affordable CD4

23. Flow Cytometry GOLD STANDARD FOR CD4/CD8 Monitoring in HIV/AIDS Patients Conceptually established by BD Worldwide

24. FACSCount - Complete SystemIVD approved ( US FDA )

25. FACSCount - Complete System Premeasured Reagents

26. FACSCount - Complete System FACSCount Reagents and Controls FACSCount Workstation

27. FACSCount Measuring Absolute CD4, CD8 and CD3 Counts In Three Easy Steps

28. FACSCount - Clinical Need • Safety • Minimal Sample Preparation • Fixed Samples • Absolute Counts • Limited Handling of • Biohazardous Samples

29. FACSCount - Clinical Need • Easy to use • Absolute Counts • Unit Dose Reagents • Minimal Hands-On Time • No Formal Training Required

30. FACSCounts in India • 35 Placements • 23 Instruments through NACO • 12 Instruments in Private Path Laboratories

31. MALES CD3+ 617 – 2485 CD19+ 125 – 701 CD4+ 337 – 1090 CD8+ 174 – 1240 Ratio 0.61 – 2.68 HIV Patients-Low CD4 FEMALES CD3+ 898 – 2786 CD19+ 119 – 721 CD4+ 424 – 1050 CD8+ 255 – 1353 Ratio 0.95 – 2.40 Reference Range of Lymphocytes Subsets In Healthy Individuals (India)

32. FACSCount - Analysis • A hard copy results • The control • Patient sample

33. HIV Monitoring BD provides a gold standard system of reliable products and support services for CD4+ T-cell testing at affordable pricing. • Appropriate Technology • Flow Cytometry • Capacity Building • Training, Support, Service • Reliable Laboratory Systems • GMP/GLP (IVDP)/GCP

34. Performance of Competitive Tests

35. Pharmas taking action • Abbott Laboratories • Bristol-Meyers Squibb • GlaxoSmithKline • Hoffmann La-Roche Ltd • Boehringer Ingelheim • Merck & Co., Inc • Pfizer • Cipla • Ranbaxy • Matrix

36. General concensus is that flow cytometry based CD4 count is the most accepted technique to HIV monitoring • Quantitative – more cells counted, more reliable result • Specific – mAb, identify CD4 T-lymphocytes most precisely • QA/QC – internal and external - necessary for accreditation • Throughput – manual operation is acceptable as technician costs are not significant and does not limit test demand • Existing flow cytometry systems (FACSCalibur or similar) are adequate/accepted for Central Reference facilities – usually at the sites of principal government services • Perform broad clinical testing plus Research studies associated with international partners • Affordable lower cost platform required at Provincial and Local clinic sites – dedicated clinical test platform • Capability to perform CD4 count ( - ideal with broader test menu ) • Current FACSCount system should be viewed as maximal technology complexity – sample preparation potential issue – reagent cost limitation • Challenge from Partec / Guava like technology or alternative assays

37. Application Comparison

38. Affordable Testing for HIV/AIDS in the Developing WorldFebruary 5, 2004 On January 14, 2004, the William J. Clinton Presidential Foundation announced that BD, through BD Biosciences, had entered into an agreement with the Foundation to provide affordable CD4 testing, both reagents and instruments, to the poorest and most severely HIV/AIDS afflicted areas in the developing world. The CD4 immunocytometry test has been used to establish decision points for antiretroviral (ARV) therapy, to monitor efficacy of various treatments, as a prognostic indicator, and as one criterion for initiating prophylaxis for opportunistic infections. The viral load test helps measure how effective ARV therapies are in suppressing the virus and can alert clinicians about the need to adjust dosages or change regimens.

39. BD BiosciencesAccomplishing our Mission – HIV/AIDS BD Biosciences is Helping With the Global AIDS Crisis by: • Supporting the Brazilian AIDS prevention program • Example program for reducing mortality and new infections in the developing world • Our flow instruments and reagents are used as the gold standard to monitor disease progression in HIV-infected individuals • Collaborating with the International AIDS Vaccine Initiative (IAVI) • Speed the development and distribution of preventive AIDS vaccines • Our flow instruments and reagents are used to detect immune response to the vaccine

40. HIV/AIDS PANDEMIC 2010 80+ million PLWHA India (15+ million) China (15+ million) Negative growth rates in most of Africa due to AIDS mortality WW economic impact 2002 Brazil Achieves 50% reduction in AIDS deaths over last 10 years Accessible drugs and health care $500,000 million savings on AIDS impact