intro

2. intro • Persons in a population respond to diseases differently due to the phenotypic variations of resistance. • It is proposed that inheritance factors play a major role in mortality rates. • HIV-1 epidemic has evoked the study of genetic variation and susceptibility to infections in different hosts. • Research has shown a specific allele of the HLA locus to be associated with different rates of progression from infection to AIDS.

3. Chemokine Receptors • Chemokine receptors are G-protein-linked serpentine receptors • Also used as co-receptors for the binding of immunodeficiency viruses (eg HIV) to leucocytes. • Chemokines RANTES, MIP-1 , and MIP-1 role as natural HIV-1 suppressors are being studied. • The chemokine co-receptor and receptors associated with the above mentioned are fusin, CD4, CKR2B, CKR3 and CKR5 (principal cellular receptor). • Individuals at high risk for the HIV-1 infection have been observed to have CD4+T cells that have been relatively resistant to infection.

4. The Genetic mapping of CKR5 and Fusin • The locus encoding fusin and the CKR5 are genetically mapped using the polymerase chain reaction (PCR). • PCR screens a panel of 90 radiation hybrid (RH) DNA samples of the human genome. • The allocation of RH results indicates that fusin is positioned on chromosome 2q21 and CKR5 on chromosome 3p21. • These loci are mapped in small clusters along different locations in the human genome (represented in the subsequent graph).

5. Representation of Chemokine Receptor clusters in the Human Genome

6. Determination of Genotype Frequency among HIV-1 Infected versus Non-infected Individuals • Genomic DNA was screened by using 170 mapped polymorphic loci. • Distortion of allele and geneotype frequency among HIV-1 positive vs high risked HIV-1 negative persons were determined.

7. A Graph Showing Genotypic markers and HIV-1 infection [G test ]

8. Analysis of Graph • According to the data displayed in the graph, CKR5 show a significant distortion of genotype frequencies among the infected vs uninfected. • The other loci (CD4, chemokine SCYAL, HLADQAL, TCRA,TCRB) on the other hand did not show such a significance.

9. Further Examination of CKR5 Allele • Distribution of alleles and genotypes with genomic DNA were determined in 1955 patients in order to find important variables in HIV-1 infection and its progression • Subjects used for the experiment were high risk HIV-1 type ndividuals. • The experiment also included HIV-1-exposed seronegative individuals, HIV-1-infected AIDS patients, and HIV-1-infected individuals who have not yet progressed to AIDS. • CKR5 frequency was found to be greatest in high risk HIV-1 individuals and less in those cosidered to be of low risk. • CKR5 frequency was found least in African Americans.

10. HIV-1 infected vs non infected • CKR5 frequencies were found to be relatively the same in HIV-! Infected and non infected individuals. • A significant difference of CKR5 was found in the genotypic distribution between infected and non infected individuals. • High risk HIV-I antibody negative individuals were found to have 17 homozygous CKR5 32 genes which is highly significant (G=35.0, p=2.5*10-8). • Therefore, the CKR5 32 gene seem to have a recessive phenotype associated with HIV-1 infection resistance and antibody production. • Homozygous CKR5 32 allele was non existent in HIV-1 infected patients but the Heterozygote gene was found. • In homosexual HIV-1 infected long term non progressors heterozygotes were twice the percentage compared to rapid progressors. • In Hemophilia individuals the heterozygote frequency differences between rapid progressors and non progressors were insignificant.

11. Hemophiliacs vs Homosexuals • There is a difference in response in hemophiliacs vs homosexuals due to : • 1) transmission • 2)exposure level • 3)viral load • Hemophiliacs consist of large doses of HIV-1 contaminating clotting factors. • Homosexuals sexual transmission involve HIV-1 mucosal epithelium infection.

12. Analysis of CKR5 Heterozygote frequency

13. Homozygotes vs Heterozygotes for CKR5 32 gene • Multiple tests in 1987 and 1992 for AIDS definition show that heterozygotes for CKR5 show a delayed progression to AIDS ic comparison with homozygotes (x2 =8.1, p=0.0045). • Probability is >0.01 therefore difference is significant. • Hence single-gene CKR5 32 may be dominant and due to interaction with other genes/environment it may prolong AIDS in infected persons.

14. Results of Investigation • Persons homozygous (recessive) for CKR5 have greater reduced risk of HIV-1 infection due to absence of functional CKR5 co-receptor. • Heterozygotes can be infected but due to CKR5 HIV-1 co –receptor limiting viral spreading in infected persons ultimately delaying AIDS. • Large differences in frequencies of CKR5 were found amongst Caucasians (0.11) compared to African Americans (0.017) which may be because CKR5 is a recent recessive mutation.

15. Conclu • A Genetic restriction experiment including HIV-1-infected individuals vs. HIV-1-antibody-negative individuals were performed on 1955 patients. • The study includes the chemokine receptor 5 (CKR5) protein. • CKR5 (structural Gene) is a deletion allele found at a frequency of ~0.1 in Caucasian Americans. • Cohort study show 17 deletion homozygotes occurring exclusively in HIV-1-antibody-negative individuals.

16. References • R. May and R. Anderson, Infectious Disease in Humans (Oxford Univ. Press, New York, 1995). • S. S. Morse, Ed., Emerging Viruses (Rockefeller Univ. Press, New York, 1993). • T. I. A. Sorensen, G. G. Nielsen, P. K. Andersen, T. W. Teasdale, N. Engl. J. Med.318, 727 (1988) . • A. G. Motulsky, Human Genetics: Patterns and Approaches (Springer-Verlag, Berlin, 1981). • S. Wain-Hobson, Curr. Opin. Genet. Dev.3, 878 (1993) [Medline]. • J. M. Coffin, Science267, 483 (1995) [Medline]. • E. L. Delwart et al., ibid.262, 1257 (1993) [Medline]. • R. A. Kaslow et al., Nature Med.2, 405 (1996) [Medline]; B. F. Haynes, G. Pantaleo, A. S. Fauci, Science271, 324 (1996) [Medline]. • W. A. Paxton et al., Nature Med.2, 412 (1996) [Medline]; R. Detels et al., AIDS10, 102 (1996) . • F. Cocchi et al., Science270, 1811 (1995) [Medline]; M. Baier, A. Werner, N. Bannert, K. Metzner, R. Kurth, Nature378, 563 (1995) [Medline]. • Y. Feng, C. C. Broder, P. E. Kennedy, E. A. Berger, Science272, 872 (1996) [Medline]. • B. Federsppiel et al., Genomics16, 707 (1993) [Medline]. • H. Herzog, Y. J. Hort, J. Shine, L. A. Selbie, DNA Cell Biol.12, 465 (1993) [Medline]. • H. Nomura, B. W. Nielsen, K. Matsushima, Int. Immunol.5, 1239 (1993) [Medline]. • P. J. Madden et al., Cell47, 333 (1986) . • C. Combadiere, S. K. Ahuja, H. L. Tiffany, P. M. Murphy, J. Leukocyte Biol.60, 147 (1996) [Medline]; M. Samson, O. Labbe, C. Mollereau, G. Vassart, M. Parmentier, Biochemistry35, 3362 (1996) [Medline]. • T. Dragic et al., Nature381, 667 (1996) [Medline]. • G. Alkhatib et al., Science272, 1955 (1996) [Medline]. • H. Choe et al., Cell85, 1135 (1996) [Medline]. • B. J. Doranz et al., ibid., p. 1149. • H. Deng et al., Nature381, 661 (1996) [Medline]. • P. M. Murphy, Annu. Rev. Immunol.12, 593 (1994) [Medline]. • K. Neote, D. DiGregorio, J. Y. Mak, R. Horuk, T. J. Schall, Cell72, 415 (1993) . • C. J. Raport et al., J. Leukocyte Biol.59, 18 (1996) [Medline]. • M. A. Walter, D. J. Spillett, P. Thomas, J. Weissenbach, P. Goodfellow, Nature Genet.7, 22 (1994) [Medline]. • M. White, M. Carvalho, D. Derse, S. J. O'Brien, M. Dean, Genomics12, 301 (1992) [Medline]; M. Ravnik-Glavac, D. Glavac, M. Dean, Hum. Mol. Genet.3, 801 (1994) [Medline]. • M. Orita, Y. Suzuki, T. Sekiua, K. Hayashi, Genomics5, 874 (1989) [Medline]. • M. Samson, Nature382, 722 (1996) [Medline]. • R. Liu et al., Cell86, 367 (1996) [Medline]. • Eight CKR5 variant alleles were found. These include the 32 deletion; a variant that occurred as a heterozygote in 15 individuals of 600 screened by SSCP; four variants found as heterozygous in single individuals that were HIV-1 infected, but had not progressed to AIDS after 7 to 10 years; and two variants in HIV-1-infected individuals that had not been followed for long term as yet. Preliminary sequence analysis revealed missense alterations in codons for conserved amino acids in three of the variants. • M. W. Hilgartner et al., Am. J. Pediatr. Hematol. Oncol.15, 208 (1993) [Medline].