1. Virsuses: Human Immunodeficiency Syndrome & Acquired Immunodeficiency Syndrome

2. Simulated Anti-HIV Outline Clinical Detection and Diagnosis of HIV and HIV exposure –ELISA –Western Blot –PCR

3. HIV Diagnostic Tests ELISA Indirect evidence of HIV exposure measures

4. HIV Diagnostic Tests Western Blot PCR Directly measures HIV

5. Polymerase Chain Reaction Measures proviral DNA within the host DNA

6. Western Blot Identifies HIV proteins Protein ladder

7. A diagnostic test for Antibodies to HIV Enzyme Linked Immunosorbent Assay (ELISA) Antibodies

8. Antibodies are proteins produced by our immune system that are directed against specific antigens.

9. Antibodies Antigens Immune system Non-self

10. Now apply these concepts to the diagnostic test known as ELISA to detect antibodies against HIV from a biologic fluid.

11. Do any of these individuals carry antibodies to HIV?

12. HIV Diagnostic Tests ELISA Indirect evidence of HIV exposure measures

13. HIV Diagnostic Tests Western Blot PCR Directly measures HIV

14. Polymerase Chain Reaction Measures proviral DNA within the host DNA

15. Do any of these individuals carry antibodies to HIV?

18. FDA Approves Saliva OraQuick Rapid Test for HIV-1, HIV-2 Antibodies [March 29, 2004] (similar test is also available for blood samples, see next slides).

19. OraQuick Rapid Anti-HIV Blood Test 12 3 20 minute test Cost app. $15.00

20. Onto the ELISA lab

21. For Your Information and Files

22. Acquired Immunodeficiency Syndrome Normal CD4+ count Normal CD4 + (%) AIDS 500-1600/mm 3 20-40% <350/mm 3 begin anti-viral treatment <14% serious immune damage

23. Table 1. For Your personal Information: not for lecture. Antiretroviral Agents AgentTypeDoseMajor Toxicities AZTNRTI300 mg bidnausea, headache, low blood counts ddINRTI125-200 mg bid or 250-400 mg qd (tablet form) diarrhea, pancreatitis, peripheral neuropathy ddCNRTI0.750 mg tiddiarrhea, peripheral neuropathy d4TNRTI30-40 mg bidabnormal liver function tests, peripheral neuropathy 3TCNRTI150 mg bidminor abacavirNRTI300 mg bidhypersensitivity reaction tenofovir NRTI (nucleotide) 300 mg qd nausea, diarrhea, vomiting, flatulence

24. AZT/3TC (Combivir) NRTIone pill bid (300 mg AZT/150 mg 3TC) see above AZT/3TC/a bacavir (Trizivir) NRTIone pill bid (300 mg AZT/150 mg 3TC/ 300 mg abacavir) see above nevirapineNNRTI200 mg bidrash delavirdineNNRTI400 mg tidrash efavirenzNNRTI600 mg qhsrash, dizziness, impaired concentration, insomnia, abnormal dreams saquinavir (Fortovase) PI1200 mg tiddiarrhea ritonavirPI600 mg bidnausea/vomiting, drug interactions indinavirPI800 mg tidkidney stones nelfinavirPI750 mg tid or 1250 mg bid diarrhea amprenavirPI1200 mg bid nausea/vomiting, diarrhea, rash lopinavir/ri tonavir PIthree capsules bid (133.3 mg lopinavir/33.3 mg ritonavir) diarrhea, nausea, weakness, headache

25. Experimental drugs are italicized, and approved drugs are in regular, non-italicized type) Brand Name Generic NameAbbreviationExperimental CodePharmaceutical Company FuzeonFuzeon™enfuvirtideENFT-20Trimeris and Hoffmann-La Roche BMS-488043Bristol-Myers Squibb GSK-873,140GlaxoSmithKline PRO-542Progenics Pharmaceuticals SCH-DSchering-Plough Corporation TNX-355Tanox and Biogen Idec UK-427,857Pfizer

26. Interesting links on HIV http://www.niaid.nih.gov/factsheets/aidsstat.htm –Links to global and US HIV/AIDS statistics http://www.avert.org/pregnanc.htm –Links to HIV and pregnancy as well as numerous other links including statistics on global epidemic; HIV/AIDS quizzes and treatment. http://www.cdc.gov/hiv/pubs/facts/transmission.htm –Links to CDC and a comprehensive fact sheet on HIV transmission

27. Experimental drugs are italicized, and approved drugs are in regular, non-italicized type) Brand NameGeneric NameAbbreviationExperimental Code Pharmaceutical Company Fuzeon™enfuvirtideENFT-20Trimeris and Hoffmann-La Roche BMS-488043Bristol-Myers Squibb GSK-873,140GlaxoSmithKline PRO-542Progenics Pharmaceuticals SCH-DSchering-Plough Corporation TNX-355Tanox and Biogen Idec UK-427,857Pfizer What are Entry Inhibitors (including Fusion Inhibitors)? Entry inhibitors work by preventing HIV from entering healthy T-cells in the body. They work differently than many of the approved anti-HIV drugs – the protease inhibitors (PIs), the nucleoside reverse transcriptase inhibitors (NRTIs), and the non-nucleoside reverse transcriptase inhibitors (NNRTIs) – which are active against HIV after it has infected a T-cell. Entry inhibitors work by attaching themselves to proteins on the surface of T-cells or proteins on the surface of HIV. In order for HIV to bind to T-cells, the proteins on HIV's outer coat must bind to the proteins on the surface of T-cells. Entry inhibitors prevent this from happening. Some entry inhibitors target the gp120 or gp41 proteins on HIV's surface. Some entry inhibitors target the CD4 protein or the CCR5 or CXCR4 receptors on a T-cell's surface. If entry inhibitors are successful in blocking these proteins, HIV is unable to bind to the surface of T-cells and gain entry into the cells. Only one entry inhibitor has been approved by the U.S. Food and Drug Administration (FDA): Fuzeon™ (T-20). This drug targets the gp41 protein on HIV's surface. Some experimental drugs target proteins on T-cells: BMS-488043 targets the gp120 protein, PRO-542 and TNX-355 target the CD4 protein, and SCH-D, GSK-873,140 and UK-427,857 target the CCR5 protein. HIV-positive people who have become resistant to PIs, NRTIs, and NNRTIs will likely benefit from the entry inhibitors because they are a different class of drugs. This is good news for HIV-positive people who have tried and failed many of the currently approved anti-HIV medications. To learn more on how HIV infects a T-cell and begins to create more viruses, and where each class of anti-HIV drugs blocks this process, click on the following lesson link: The HIV Life Cycle (and the targets of each class of anti-HIV drugs) FuzeonBMS-488043GSK-873,140PRO-542SCH-DTNX-355UK-427,857protease inhibitors (PIs)nucleoside reverse transcriptase inhibitors (NRTIs)non-nucleoside reverse transcriptase inhibitors (NNRTIs)FuzeonBMS-488043PRO-542TNX-355 SCH-DGSK-873,140UK-427,857 The HIV Life Cycle (and the targets of each class of anti-HIV drugs)

28. ELISA MICROTITER PLATES Microtiter plate

30. The ELISA protocol 1 2 3 4 antigen sample Labelled 2 nd Ab Color inducing substrate

31. Results POSITIVE ANTI-HIV COLOR CHANGE NEGATIVE ANTI-HIV NO COLOR CHANGE

34. FDA Approves Saliva OraQuick Rapid Test for HIV-1, HIV-2 Antibodies [March 29, 2004] (similar test is also available for blood samples, see next slides).

35. OraQuick Rapid Anti-HIV Blood Test 12 3 20 minute test Cost app. $15.00

36. Onto the ELISA lab

37. For Your Information and Files

38. Acquired Immunodeficiency Syndrome Normal CD4+ count Normal CD4 + (%) AIDS 500-1600/mm 3 20-40% <350/mm 3 begin anti-viral treatment <14% serious immune damage

39. Table 1. For Your personal Information: not for lecture. Antiretroviral Agents AgentTypeDoseMajor Toxicities AZTNRTI300 mg bidnausea, headache, low blood counts ddINRTI125-200 mg bid or 250-400 mg qd (tablet form) diarrhea, pancreatitis, peripheral neuropathy ddCNRTI0.750 mg tiddiarrhea, peripheral neuropathy d4TNRTI30-40 mg bidabnormal liver function tests, peripheral neuropathy 3TCNRTI150 mg bidminor abacavirNRTI300 mg bidhypersensitivity reaction tenofovir NRTI (nucleotide) 300 mg qd nausea, diarrhea, vomiting, flatulence

40. AZT/3TC (Combivir) NRTIone pill bid (300 mg AZT/150 mg 3TC) see above AZT/3TC/a bacavir (Trizivir) NRTIone pill bid (300 mg AZT/150 mg 3TC/ 300 mg abacavir) see above nevirapineNNRTI200 mg bidrash delavirdineNNRTI400 mg tidrash efavirenzNNRTI600 mg qhsrash, dizziness, impaired concentration, insomnia, abnormal dreams saquinavir (Fortovase) PI1200 mg tiddiarrhea ritonavirPI600 mg bidnausea/vomiting, drug interactions indinavirPI800 mg tidkidney stones nelfinavirPI750 mg tid or 1250 mg bid diarrhea amprenavirPI1200 mg bid nausea/vomiting, diarrhea, rash lopinavir/ri tonavir PIthree capsules bid (133.3 mg lopinavir/33.3 mg ritonavir) diarrhea, nausea, weakness, headache

41. Experimental drugs are italicized, and approved drugs are in regular, non-italicized type) Brand Name Generic NameAbbreviationExperimental CodePharmaceutical Company FuzeonFuzeon™enfuvirtideENFT-20Trimeris and Hoffmann-La Roche BMS-488043Bristol-Myers Squibb GSK-873,140GlaxoSmithKline PRO-542Progenics Pharmaceuticals SCH-DSchering-Plough Corporation TNX-355Tanox and Biogen Idec UK-427,857Pfizer

42. Interesting links on HIV http://www.niaid.nih.gov/factsheets/aidsstat.htm –Links to global and US HIV/AIDS statistics http://www.avert.org/pregnanc.htm –Links to HIV and pregnancy as well as numerous other links including statistics on global epidemic; HIV/AIDS quizzes and treatment. http://www.cdc.gov/hiv/pubs/facts/transmission.htm –Links to CDC and a comprehensive fact sheet on HIV transmission

43. Experimental drugs are italicized, and approved drugs are in regular, non-italicized type) Brand NameGeneric NameAbbreviationExperimental Code Pharmaceutical Company Fuzeon™enfuvirtideENFT-20Trimeris and Hoffmann-La Roche BMS-488043Bristol-Myers Squibb GSK-873,140GlaxoSmithKline PRO-542Progenics Pharmaceuticals SCH-DSchering-Plough Corporation TNX-355Tanox and Biogen Idec UK-427,857Pfizer What are Entry Inhibitors (including Fusion Inhibitors)? Entry inhibitors work by preventing HIV from entering healthy T-cells in the body. They work differently than many of the approved anti-HIV drugs – the protease inhibitors (PIs), the nucleoside reverse transcriptase inhibitors (NRTIs), and the non-nucleoside reverse transcriptase inhibitors (NNRTIs) – which are active against HIV after it has infected a T-cell. Entry inhibitors work by attaching themselves to proteins on the surface of T-cells or proteins on the surface of HIV. In order for HIV to bind to T-cells, the proteins on HIV's outer coat must bind to the proteins on the surface of T-cells. Entry inhibitors prevent this from happening. Some entry inhibitors target the gp120 or gp41 proteins on HIV's surface. Some entry inhibitors target the CD4 protein or the CCR5 or CXCR4 receptors on a T-cell's surface. If entry inhibitors are successful in blocking these proteins, HIV is unable to bind to the surface of T-cells and gain entry into the cells. Only one entry inhibitor has been approved by the U.S. Food and Drug Administration (FDA): Fuzeon™ (T-20). This drug targets the gp41 protein on HIV's surface. Some experimental drugs target proteins on T-cells: BMS-488043 targets the gp120 protein, PRO-542 and TNX-355 target the CD4 protein, and SCH-D, GSK-873,140 and UK-427,857 target the CCR5 protein. HIV-positive people who have become resistant to PIs, NRTIs, and NNRTIs will likely benefit from the entry inhibitors because they are a different class of drugs. This is good news for HIV-positive people who have tried and failed many of the currently approved anti-HIV medications. To learn more on how HIV infects a T-cell and begins to create more viruses, and where each class of anti-HIV drugs blocks this process, click on the following lesson link: The HIV Life Cycle (and the targets of each class of anti-HIV drugs) FuzeonBMS-488043GSK-873,140PRO-542SCH-DTNX-355UK-427,857protease inhibitors (PIs)nucleoside reverse transcriptase inhibitors (NRTIs)non-nucleoside reverse transcriptase inhibitors (NNRTIs)FuzeonBMS-488043PRO-542TNX-355 SCH-DGSK-873,140UK-427,857 The HIV Life Cycle (and the targets of each class of anti-HIV drugs)