Evaluation of Human Thymic Function during Health and HIV-1 Infection Ping Ye and Denise Kirschner DIMACS Workshop 9/23/2001

Outline Thymic function in healthy people Thymopoiesis Markers to represent recent thymic emigrants (RTE) Thymopoiesis model TREC model Thymic function during pediatric HIV-1 infection HIV-1 infection Clinical and experimental studies Thymic infection model Evaluation of TREC

The Thymus Janeway CA et al. Immunobiology 5th ed. 2001 Haynes BF. Clin Immunol. 92(1):3-5,1999

Thymopoiesis Berkowitz RD et al. J Immunol. 161(7):3702-10, 1998

Recent Thymic Emigrants (RTE) Lack of phenotypic markers for human RTE Human RTE generally refer to T cells that have undergone only a few cellular divisions after leaving the thymus T cell receptor excision circles (TREC) have recently been used as a measure of the number of RTE

T Cell Receptor Excision Circles (TREC) Pieces of DNA fragments that are generated during TCR gene rearrangement in the thymus and then exported from the thymus to periphery within T cells episomally Haynes BF et al. Annu Rev Immunol. 18: 529-60, 2000 Janeway CA et al. Immunobiology 5th ed. 2001

TREC as A Measure of RTE

Parameters Estimating RTE TREC concentration Is affected by both RTE and peripheral T cell proliferation and death Naïve T cells (CD45RA+, CD62L, CD95+) May have long lifespan May proliferate in an antigen-independent manner May rapidly convert to memory cells May be converted from memory cells Thymic volume Assuming thymic volume is proportional to RTE levels

Two Questions to Address 1st Can TREC concentration be properly used as measure of RTE? 2nd Whether thymic infection with different HIV-1 strains contribute to differences in disease progression?

Thymopoiesis Model Cell Source THYMUS TN ITTP DP SP4 SP8 BLOOD/ TES(t) TN ITTP DP SP4 SP8 BLOOD/ LYMPHOID TISSUES CD4+ RTE CD8+ RTE

Simulation Results for Thymopoiesis Model Thymocytes RTE/day

Model Simulation Values Compared with Experimental Data

TREC Model THYMUS SP4 SP8 CD4+ RTE CD8+ RTE CD4+ T cell CD8+ T cell BLOOD/ LYMPHOID TISSUES

Simulation Results for TREC Model Douek DC et al. Nature. 369(6712):690-5, 1998. Zhang L et al. J. Exp. Med. 187(11): 1767-78, 1998

Four Events Affecting TREC Concentration THYMUS SP4 SP8 1 CD4+ RTE CD8+ RTE 2 3 CD4+ T cell CD8+ T cell 4 TREC TREC BLOOD/ LYMPHOID TISSUES 1 Thymic output 2 T cell division 3 T cell death 4 TREC degradation TREC concentration is affected by

TREC Can Be Equally Affected by Thymic Output and T Cell Division (at Any Age) #

TREC Concentration during Aging Healthy Volunteers sjTREC cjTREC Age (years) Douek DC et al. Nature. 396(6712): 690-5, 1998

Thymic Involution Induce TREC Decline (Over Entire Lifespan) TREC concentration is a good marker for RTE in healthy people

Summary Thymic Function in Healthy people Our model quantifies the number of RTE and TREC concentration over an 80-year lifespan TREC concentration can be equally affected by thymic output and T cell division at any age Thymic involution induces TREC concentration decline over the entire lifespan TREC concentration is a good marker for both CD4+ and CD8+ RTE in healthy people

Outline Thymic function in healthy people Thymopoiesis Markers to represent recent thymic emigrants (RTE) Thymopoiesis model TREC model Thymic function during pediatric HIV-1 infection HIV-1 infection Clinical and experimental studies Thymic infection model Evaluation of TREC

HIV-1 Structure Associated with AIDS Retrovirus family Two identical ss RNA Enveloped virus Antigenic variation Infects CD4+ cells Entry requires CD4 and coreceptor

HIV-1 Entry

Classification of HIV-1 Strains R5 strain Uses CCR5 as coreceptor Replicates slowly and is minimally cytopathic Is primarily transmitted Is prevalent in the early stage of disease X4 strain Uses CXCR4 as coreceptor Replicates fast and is highly cytopathic Is rarely transmitted Appears in approximately 50% of patients in the late stage of disease R5X4 strain Uses either CCR5 or CXCR4 as coreceptor Has similar properties as the X4 strain

HIV-1 Disease Progression Hypotheses for the decline of CD4+ T cells in the blood Changes in cell migration patterns Changes in cell life-spans Changes in cell production by the thymus

Clinical Studies of Thymic Infection with HIV-1 TREC levels in CD4+ and CD8+ T cells decline Naïve CD4+ and CD8+ T cell numbers decline Thymic volume decreases The thymus undergoes morphological changes “Thymic dysfunction” (TD+) is described in a subset of vertically-infected infants

“Thymic dysfunction” (TD+) Two distinct modes of pediatric HIV-1 disease progression Normal progressors 10 years to AIDS (85%) Fast progressors 2-3 years to AIDS (15%) lower CD4+ and CD8+ counts (DiGeorge Syndrome)

Experimental Studies of Thymic Infection with HIV-1 Coreceptors CCR5 Expressed in low levels on DP, SP4, SP8 cells CXCR4 Expressed in high levels on ITTP, DP cells, low levels on SP4 and SP8 cells HIV-1 strains R5 strain Infects DP and SP4 cells Minimally cytopathic X4 strain Infects ITTP, DP and SP4 cells Highly cytopathic Thymocyte maturation stages SP4,SP8 Support active viral replication ITTP,DP Support less viral replication

Two Questions to Address 1st Can TREC concentration be properly used as measure of RTE? 2nd Whether thymic infection with different HIV-1 strains contribute to differences in disease progression?

Thymic Model With HIV-1 Infection Cell Source THYMUS TES(t) TN X4 ITTP ITTPX R5 X4 DPR DP DPX R5 X4 SP8R SP4R SP4 SP4X SP8X SP8 R5 X4 R5 X4 CD4+ RTE CD8+ RTE BLOOD/LYMPHOID TISSUES

Virtual Pediatric Thymic Infection

Simulations of CD4+ RTE Predict Bimodal Pattern of Blood CD4+ T Cell Counts

Simulation Values at Year Two of Virtual Infection Compared with Experimental Data

Bifurcation Parameters Implications for Treatment Viral influx from blood into the thymus Virulence of strains (viral infection rate and production rate) HIV-1 induced death of uninfected thymocytes Progenitor cells from bone marrow Carrying capacity of the thymus

TREC Concentration during HIV-1 Infection HIV-1 Infected Subjects sjTREC Age Age (years) Douek DC et al. Nature. 396(6712): 690-5, 1998

Two Questions to Address 1st Can TREC concentration be properly used as measure of RTE? 2nd Whether thymic infection with different HIV-1 strains contribute to differences in disease progression?

T Cell Kinetics during HIV-1 Infection We include these effects in the model to represent HIV-1 infection

T Cell Counts and TREC Concentrations in HIV-1 Infected Subjects (Age 30) T cells TREC Margolick JB et al. J Acquir Immune Defic Syndr. 6(2): 153-61, 1993 Zhang L et al. J. Exp. Med. 187(11): 1767-78, 1998

Model Prediction for Thymic Output during HIV-1 Infection TREC concentration is a good marker for CD4+ RTE TREC concentration is NOT a good marker for CD8+ RTE

Summary Thymic Function during Pediatric HIV-1 Infection Infection with R5 and X4 strains induces different degrees of thymic dysfunction, which is related to CD4+ T cell counts and disease progression as seen in pediatric patients Thymic infection in pediatric patients is more severe than in adult patients, likely due to higher viral loads and a more active thymus Suppressing viral load in blood, high drug efficacy within the thymus, and improving inherent thymic function are necessary for reconstitution of RTE levels Protease inhibitors have high levels of efficacy directly suppressing viral replication within the thymus, while reverse transcriptase inhibitors have low efficacy TREC concentration is a reliable marker for CD4+ RTE, but not for CD8+ RTE in HIV-1 infected subjects

Conclusions Peripheral T cell turnover should be examined together with TREC concentration as a measure of RTE Infection with different HIV-1 strains induces different degrees of thymic dysfunction, contributing to CD4+ T cell depletion and disease progression With adequate suppression of viral replication within both the blood and the thymus during HAART, thymic function recovery can reconstitute immune cell numbers

Publications Ping Ye and Denise Kirschner (2002). Reevaluation of T cell receptor excision circles as a measure of human recent thymic emigrants. Journal of Immunology, 168(10), 4968-4979. Ping Ye, Athena Kourtis, and Denise Kirschner (2002). The effects of different HIV-1 strains on human thymic function. AIDS Research and Human Retroviruses, 18(17) (In press). Ping Ye, Athena Kourtis, and Denise Kirschner . Reconstitution of thymic function in HIV-1 patients treated with highly active anti-retroviral therapy (submitted).

Acknowledgments Dr. Denise Kirschner Collaborator: Dr. Athena Kourtis Dr. Ramit Mehr Kirschner lab members Funding NIH and Whitaker Foundation to DEK Rackham Graduate School and Elizabeth Glaser Pediatric AIDS Foundation to PY

Clinical Studies of Thymic Function during HAART TREC levels recover Naïve CD4+ and CD8+ T cell numbers increase Thymic volume increases Children with “thymic dysfunction” response well to HAART

Model of Thymic Function Reconstitution during HAART Cell Source THYMUS TES(t) TN ITTP X4 ITTPX DPR R5 X4 DPX DP R5 X4 SP8R SP4R SP4 SP4X SP8X SP8 R5 X4 R5 X4 CD4+ RTE CD8+ RTE BLOOD/LYMPHOID TISSUES Reduce viral influx into the thymus Decrease viral infection and production (if thymic drug efficacy > 0%) Decrease HIV-1 induced death of uninfected thymocytes Increase progenitor cells from bone marrow Increase carrying capacity of the thymus

Simulations of pediatric thymocyte dynamics during HAART

Simulations of CD4+ RTE Predict HAART Agent Thymic Efficacy RTI HAART