HIV Infection and Memory T Cells [Revised]

AlP 2 1 R21 A1058806-01A1
GARZINO-DEMO, A
1 R21A1058806-01A1 GARZINO-DEMO, ALFREDO
RESUME AND SUMMARY OF DISCUSSION: Dr. Garzino-Demo submitted this amended R21
application in response to PA-02-046 (Innovation for Grants in AIDS Research). The Principal
Investigator will determine HIV suppressor chemokines released during antigen specific CD+ T cell
memory responses, and whether memory cells that express high levels of CCR5 are protected from
HIV infection. Prior to the review, consultants were asked to provide written evaluations of the
applications. They were able to read each other's critiques prior to the review meeting. At the meeting,
based on these written critiques, reviewers were asked to rank and identify R21 applications that were
non-competitive. Non-competitive applications, including this one generally belong to the lower half of
all R21 applications this panel customarily reviews. The discussion focused on the weaknesses
outlined in the critiques below.
DESCRIPTION (provided by applicant): CCR5 ligands RANTES, macrophage inflammatory protein
(MIP)-l (z, and MIP-113, are potent and specific inhibitors of strains of HIV that use CCR5 as a
receptor, i.e., the strains most involved in primary infection. Recently, we observed that release of
CCR5 ligands is a consistent and reproducible parameter of response to antigen activation in studies
using peripheral blood mononuclear cells (PBMC) and whole blood. In this study, we show that CCRS
ligands, especially MIP-1 alpha and MIP-1 beta, are released upon antigen (Tetanus toxoid, TTx)
stimulation in 81% (n=1 6) of subjects tested, as detected by.a standard ELISA in tissue culture
supernatants of antigen-activated cells. In contrast, ELISA for other cytokines from the same
supernatants revealed that IFN-,{ release could be detected only in 31 % of subjects, IL-4 could be
detected only in 12% of the subjects, and IL-2 was not detectable in any of the subjects tested.
Similarly, proliferative responses to TTx, as measured by a standard tritiated thymidine incorporation
assay, were detectable in only in 56% of the subjects. Similar observations have been reported in flow
cytometry studies, and resonate with previous findings emphasizing the role of CCR5 in memory T cell
responses. In summary, release of CCR5 ligands MIP-lot and MIP-l[3 is one of the most sensitive and
reproducible correlates of memory response. These ligands, released in memory responses, may play
a role in controlling HIV infection. For example, while most HIV-specific memory cells are susceptible to
HIV infection, some of these cells display resistance to infection, through an unexplained mechanism.
Taken together, these observations suggest that CCR5 ligands, and in particular MIP-1 c_ and MIP-l[_,
released in the course of memory responses may play a role in protecting CD4 + memory T cells from
infection. Accordingly, we propose 1) to determine whether HIV suppressor chemokines are released
during antigen specific CD4 + T cell memory responses, and 2) to determine whether memory cells that
express high levels of the prototypical CCR5 ligand MIP-113 are protected from HIV infection.
CRITIQUE 1:
Significance: This is a resubmission of a R21 for a new investigator who is responding to the PA-02 -046. The focus on this application is to explore the role of CCR5 ligands as released by memory T cells
and the subsequent protection against HIV infection, in an autocrine or paracrine manner. CCR5
1igandshave teenpostulated~tcklatrongrosn-prvnting infection. Dissectingthis--_
mechanism further will enhance effort at therapeutic or preventive intey ntion. Man of the proposed
experiments, however, have already been shown by others. 'y'eS. J5
Approach: The investigator first proposes to determine that antigen-specific 0D4+ T cell release CORS
ligands, specifically in response to tetanus rechallenge. CD4+IRA- cells will be isolated by magnetic
beads and evaluated with surface markers and characterized for their ability to make MIP-1 b and IFNa. CCR5 ligands will also be measured in the supernatant. Infectivity assays will also be performed in
the presence or absence of the supernatant (with and without immunoabsorption of QORS ligands). The
expected finding is that memory CD4 T cells will make CORS ligands and that MIP-IB will suppress
HIV infection. A wealth of data have been generated by others demonstrating the ability of beta

chemokines to suppress R5 infection and it is unclear how this aim will contribute to the current
understanding of this mechanism. Furthermore, it has also been shown that antigen-specific CD4 T