Abstracts for the 25th Annual Scientific Meeting of the International ...

J Immunother Volume 33, Number 8, October 2010
Abstracts
its presentation to DC is critical in overcoming self antigen
tolerance and avoiding autoimmune response. We are developing a
DC based vaccine therapy protocol targeting HAAH with a novel
approach of using nano-particles to introduce HAAH to the
professional antigen presenting cell. As the target, HAAH is a
tumor specific antigen which is specifically expressed on the surface
of malignant cells. It is responsible for proliferation, motility and
invasive phenotype of cancer cells. HAAH internalizes when bound
to the antibodies. Neutralizing or inhibiting expression of the cellsurface
HAAH, reverses cancer cell to normal phenotype. Anti-
HAAH antibodies have proven efficiency in inhibiting tumor
growth in animal studies (passive immunotherapy). Aside from the
target, the loading of DC is accomplished by utilizing nanoparticles
with HAAH fragments on their surface. The technique is
novel and consists of engineering lambda-bacteriophage to express
HAAH-constructs on their surfaces. We have been successful in
purifying this baceriophage and neutralizing them to become nonreplicating
particles. These particles have 200 to 300 copies of
HAAH fragments on their surface and their CpG motifs in DNA
sequence act as an excellent adjuvant and stimulant. Delivering
HAAH antigens on lambda head will help to induce co-stimulatory
activity in antigen presenting cell such as dendritic cells, during
processing of foreign antigens on those cells. Thus in this case such
added lambda protein and its CpG moiety act as an adjuvant. Also
due to particulate nature, phage-displayed HAAH peptides can
access maturation through both the major histocompatibility
complex (MHC) I and MHC II pathway, and activates both
T cell and B cell mediated immunity against HAAH specific cancer
antigens. The added cellular response mediated by CD8+ T cells
helps to eliminate cancer cells by active lysis process. The nontypical
presentation of the antigen and its specificity to cancer cells
results in overcoming the self antigen tolerance an un-wanted autoimmune
response. The first indication is lung cancer for which we
have an HAAH-based companion diagnostic test for enrolment
and monitoring.
Global Gene Expression Analysis of 15 kD Granulysin
Stimulated Monocytes Compared to GM-CSF
Luciano Castiello*, Michael W. Finnw, Francesco M. Marincola*,
Alan M. Krenskyw, Carol Claybergerw, David F. Stroncek*,
Marianna Sabatino*. *Department of Transfusion Medicine;
w Laboratory of Cellular and Molecular Biology, NIH/NCI,
Washington, DC.
Granulysin is an immunoregulatory protein existing in several
isoforms. The 9 kD isoform is proinflammatory, chemotactic and
cytotoxic against microbes and tumors. The 15 kD isoform is not
well characterized functionally although its expression plays a key
role in some excessive immune reactions. Unpublished data from
our laboratory indicates that the 15 kD form induces the
differentiation of monocytes into immature dendritic cells (DCs).
The aim of this study was to characterize at the transcriptome level
the effects of 15 kD granulysin on monocytes. RNA of elutriated
monocytes from three donors cultured for 4, 12, 24 and 48 hours in
presence of GM-CSF or 15 kD granulysin was amplified, labeled
and hybridized on Agilent Oligo Microarrays. Gene expression
analysis revealed a common cytokine-induced signature at each
time point, with a more specific signature for each cytokine at later
time point. GM-CSF specifically downregulated genes related to
cell cycle arrest and the immune response. In particular, cytokine
production, lymphocyte mediated immunity and humoral immune
responses were the Gene Ontology families overrepresented among
the downregulated genes at late time points. On the other hand,
15 kD granulysin induced immune response, chemotaxis and cell
adhesion genes. Moreover, only 15 kD granulsyin induced the
activation of pathways related to fundamental DC functions, such
as co-stimulation of T cell activation and Th1 development. This
study provides important insights on the effects of a novel agent,
15 kD granulysin, that will likely be useful for future immunotherapies
aimed at activation of the immune response (Fig. 1).
FIGURE 1. A, Heat map of 6103 genes statistically differentially
expressed (P