Apoptotic fractions as cell-associated antigen sources: ACR vs blebs

7 CHAPTER 7 W ith an increasing number of clinical successes in targeting both solid and leukemic tumors by tumor infiltrating lymphocytes (TILs) or T cells engineered to express chimeric antigen receptors (CARs), our increased knowledge on DC biology, and the recent remarkable breakthroughs with checkpoint blockade, the field of tumor immunotherapy is clearly advancing. With these advances and the realization that personalized treatment holds the key to the future for anti-cancer therapy, the use of patient-derived whole tumor cells as a source of tumor or leukemia associated antigens (TAA or LAA) warrants more intensive clinical exploitation. Acute myeloid leukemia (AML) is particularly attractive in this regard, as tumor cells are often readily accessible in the blood. However, loading DC with e.g. apoptotic tumor cells can interfere with DC function due to their generally immunosuppressive nature 1–6. It was demonstrated in mice that microvesicles released during apoptosis, i.e. blebs, can induce DC maturation, whereas the larger apoptotic cell remnants (ACR) did not 7. We therefore set out to explore the use of apoptotic blebs (0.2 μm to 0.6 μm in diameter) as a source of TAA for DC loading, as compared to ACR (2 to 10 μm). Since the isolation of blebs from apoptotic cells requires a specific isolation protocol involving stringent centrifugation steps, it is safe to assume that most studies conducted in the past were using ACR and discarded apoptotic blebs. We have demonstrated in chapter 2 that blebs are more readily ingested by monocyte-derived dendritic cells (MoDC) as compared to ACR. Next to a larger percentage of MoDC that had taken up the antigen, bleb-loaded MoDC also up-regulated CCR7, gained in chemotactic reactivity towards the lymphoid homing chemokine CCL19, and possibly retained a higher level of motility after ingesting blebs as a consequence of their smaller size as compared to ACR. Next to an increase in migration, bleb-loaded MoDC also induced TH1 skewing in an MLR and a higher antigen-specific CD8 + T cell outgrowth with higher antigen-specific functional avidity, as compared to ACR-loaded MoDC. It remains elusive what the exact underlying mechanism is that drives the observed increase in homing and priming ability of bleb-loaded MoDC. It might be explained by an increase in activation state of these DC, but this was not reflected by a detectable increase in expression levels of co-stimulatory molecules, other activation markers, or T cell driving cytokines. In any case, our …