In an attempt to counteract the immunosuppression associated with CLL [4
] and the deficit of endogenous DCs in CLL patients [6
], we tested the feasibility of generating functional αDC1 from the blood of CLL patients, the feasibility of DC loading with autologous tumor cells, and their ability to induce functional, CLL-specific immune responses.
Compared with sDCs, αDC1 of patients with CLL showed higher expression of several costimulatory molecules and 10–60 times greater secretion of IL-12p70. Although the overall stimulatory capacity of DC types and their ability to take up CLL material were comparable, αDC1 were much more effective in inducing functional, tumor-specific CTL responses, as manifested by their superior induction of IFN-γ-producing, tumor-specific CD8+ T cells and their higher cytolytic activity.
Our previous experiments demonstrated that αDC1 loaded with defined HLA-A2-restricted peptides are effective inducers of CTL responses against such antigenically defined tumors as melanoma [18
]. This led to the design of our currently ongoing clinical trials of vaccination with peptide-loaded αDC1 in HLA-A2+
patients with advanced melanoma and other forms of cancer. The current data pave the way to the application of αDC1-based vaccines against tumors with poorly defined rejection antigens and to the treatment of patients with unique HLA types.
Important for the feasibility of using CLL-loaded αDC1 as cancer vaccines or as ex vivo inducers of CLL-specific CTLs for adoptive immunotherapy, we have observed that the IL-12-producing function and CTL-inducing function are not compromised by exposure to CLL cells at the stage of DC maturation. In light of the previously reported contribution of the dysfunction of endogenous DCs [6
] to the overall immuno-suppression observed in CLL patients [4
], the current data suggest the possibility of restoring the immunosurveillance in this group of patients using ex vivo-generated DCs.
Two factors crucial for successful generation of αDC1 in our study were the purity of the monocyte population and the type of medium. In contrast to nonhematologic malignancies [18
], in case of CLL, it was critical to apply a magnetic selection of CD14+
cells to avoid contamination of CLL cells in the monocytic fraction of the density-based PBMC separation and the resulting reduced yield of DCs (, inset). This negative impact of CLL cells on early stages of DC generation is reminiscent of the previously reported ability of CLL cells to suppress DC differentiation [6
]. Although the mechanism of such suppression has not been elucidated, the previously reported hyperactivation of STAT3 and ERK is possible by the following factors: tumor-derived prostanoids, vascular endothelial growth factor, IL-10, and IL-6 [24
]. It also remains to be tested whether the defective function of DCs in CLL patients can be reversed using STAT3 blockade or inhibition of MAPK activity, previously shown to enhance immune-mediated, anti-tumor effects [26
]. In contrast to the reports showing the functional deficit of DCs differentiating in the presence of CLL cells [6
], we did not observe any significant IL-12-inhibitory impact of CLL cells added during DC maturation, consistent with our observations that the susceptibility of DCs to IL-12-suppressing factors is reduced upon DC maturation [17
Generation of the optimal cell numbers also required the substitution of the originally used [18
] AIM-V medium with Cellgenix DC medium. Although the mechanism of the superiority of the Cellgenix DC medium remains unclear, the consistently observed generation of higher numbers of αDC1 led to our prioritization of this medium for the prospective generation of αDC1 for our prospective clinical trials.
In conclusion, this study demonstrates that high numbers of αDC1 can be generated successfully from the blood of CLL patients and used as effective inducers of autologous, CLL-specific CTLs. High immunologic activity of such cells suggests their possible use as vaccines or as ex vivo inducers of tumor-specific CTLs for adoptive transfer to overcome the CLL-associated immune dysfunction.