Objective: An altered function of lymphocytes, DC and
immature myeloid cells appears to be an hallmark
of tumor-mediated immune suppression and the two
inhibitory co-stimulatory receptors PDL-1 and CTLA4
might have a role in this context. The aim of the
present in vitro study was to assess whether PaCa
cells cross-talk with normal mononuclear circulating cells
(PBMC) causing them to acquire an immunesuppressive
phenotype and to evaluate whether PDL1 and CTLA4 are
involved.
Methods: PBMC from blood donors were cultured for 4
days in Control (CTL) and in the PaCa cancer cell line
Capan1 conditioned media (CM). Lymphocytes subsets
(CD4+, CD8+, CD4+CD25+) and CD33+ immature
myeloid cells subsets (CD14+/-; HLA-DR+/-) expressing
or not PDL1 and/or CTLA4 were analysed by flow
cytometry. To assess immunesuppressive function,
myeloid cells were FACS sorted and co-coltured with
allogenic total T lymphocytes in 1:20 and 1:40 ratio.
Total T lymphocytes proliferation was determined by 3HThymidine
uptake.
Results: Capan1 CM caused an expansion of
CD4+CD25+ (p=0.01) and a reduction of CD33+CD14-
HLA-DR+ (p=0.03) cells. In this latter cellular subset,
CM caused also an increase of PDL1 (p=0.046) and
a decrease of CTLA4 (p=0.05) positive cells. FACS
sorted CTL and CM CD33+CD14-HLA-DR+ cells did
not significantly affect the proliferation of allogenic
total T lymphocytes at 1:20 (p=0,54) or at 1:40 ratio
(p=0,81). The CD33+CD14-HLA-DR+ PDL-1+ cells did
not significantly modify allogenic T cells proliferation with
respect to PDL- cells (p=0,11), while those cells which
were CTLA4 negative caused a significant inhibition of
T cell proliferation in comparison of CTLA4 positive cells
(p=0,008).
Conclusions: PaCa-derived soluble factors induce
the expansion of the inhibitory lymphocytes subset
CD4+CD25+ and a reduction of the immature
CD33+CD14-DR+ dendritic cells. The tumor associated
reduced expression of the inhibitory molecule CTLA4 in
this cell population was demonstrated to characterize an
immunosuppressive phenotype and this study suggests to
take care in the use of anti-CTLA4 therapies