A new class of antibody-functionalized, semi-flexible and filamentous polymers (diameter 5-10 nm, length $200 nm) with a controlled persistence length, a high degree of stereoregularity and the potential for multiple simultaneous receptor interactions has been developed. We have decorated these highly controlled, semi-stiff polymers with T cell activating anti-CD3 antibodies and analyzed their application potential as simple synthetic mimics of dendritic cells (sDCs). Our sDCs do not only activate T cells at significantly lower concentrations than free antibodies or rigid sphere-like counterparts (PLGA particles) but also induce a more robust T cell response. Our novel design further yields sDCs that are biocompatible and non-toxic. The observed increased efficacy highlights the importance of architectural flexibility and multivalency for modulating T cell response and cellular function in general.

Key Points• Cross-presentation of both soluble and cell-associated tumor antigens by human DC subsets is enhanced by addition of adjuvant TLR agonists. • Ability to cross-present exogenous antigen with high IFN␣ secretion puts human pDCs as activators of CD8 ϩ T cells in antitumor responses. IntroductionDendritic cells (DCs) are the professional antigen presenting cells (APCs) of the immune system with the unique capacity to attract and activate naive CD4 ϩ and CD8 ϩ T cells. 1 After infection or inflammation, DCs undergo a complex maturation process and migrate into lymph nodes where they present antigens (Ags) to T cells. The DC family is very heterogeneous and consists of different DC subsets, each with distinct functional characteristics. In human peripheral blood, at least 2 main populations of DCs can be distinguished: CD11c positive myeloid DCs (mDCs) and CD11c negative plasmacytoid DCs (pDCs). Myeloid DCs can be further subdivided based on the expression of CD16, CD1c, and BDCA3. 2 Transcriptional profiling revealed significant differences between the human blood DC subsets, 3 probably reflecting differences in their Ag-presenting capacities. Furthermore, mDCs and pDCs show clearly different responses to products derived from pathogens, as a result of their distinct Toll-like receptor (TLR) expression profiles. 4 Myeloid DCs have the capacity to produce IL-12 in response to microbial stimuli through TLRs, and thereby, induce Th1 responses. 5,6 Plasmacytoid DCs (pDCs), in contrast, are the key effectors in innate immunity because of their capacity to produce large amounts of type I IFNs in response to bacterial or viral infections. 7 Similar to mDC-derived IL-12, pDC-derived type I IFNs also participate in T-cell priming as Th1-inducing cytokines. 8 In addition to directing CD4 ϩ Th responses, DCs are also important for the generation of CD8 ϩ cytotoxic T-cell responses against viruses and tumors. As professional APCs, DCs have the unique capacity to take up, process, and present exogenously encountered Ags for cross-presentation via MHC class I molecules to CD8 ϩ T cells. Numerous studies have been performed to comprehend this cross-presentation process, and these have revealed 2 major pathways: (1) the "canonical" proteasome dependent cytosolic pathway, and (2) the TAP and proteasome independent pathway. [9][10][11][12] Many studies however, made use of murine DCs to study cross-presentation capacities and mechanisms used by different DC subsets. There is ample evidence that identified the CD8␣ ϩ DC as the superior cross-presenting DC subset in mice. 13,14 Recently, a lot of effort has been put toward finding the human counterpart of the murine cross-presenting CD8␣ ϩ DC subset. Despite basic similarities between human and mouse DCs, direct comparison is difficult because of large differences in cell-surface markers and TLR expression, in particular also for pDCs, which in contrast to mice are the sole TLR9-expressing subtype of DCs in Submitted June 7, 2012; accepted November 9, 2012. Prepublished onl...

Effective vaccines consist of 2 components: immunodominant antigens and effective adjuvants. Whereas it has been demonstrated that targeted delivery of antigens to dendritic cells (DCs) improves vaccine efficacy, we report here that co-targeting of TLR ligands (TLRLs) to DCs strongly enhances adjuvanticity and immunity. We encapsulated ligands for intracellular TLRs within biodegradable nanoparticles coated with Abs recognizing DC-specific receptors. Targeted delivery of TLRLs to human DCs enhanced the maturation and production of immune stimulatory cytokines and the Ag-specific activation of naive CD8 ؉ T cells. In vivo studies demonstrated that nanoparticles carrying Ag induced cytotoxic T-lymphocyte responses at 100-fold lower adjuvant dose when TLRLs were co-encapsulated instead of administered in soluble form. Moreover, the efficacy of these targeted TLRLs reduced the serum cytokine storm and related toxicity that is associated with administration of soluble TLRLs. We conclude that the targeted delivery of adjuvants may improve the efficacy and safety of DC-based vaccines. (Blood. 2011;118(26):6836-6844) IntroductionMost vaccines currently on the market are based on the induction of long-lived Ab responses. A major challenge is the generation of vaccines that, next to Abs, also induce Ag-specific killing of pathogen-infected cells or tumor cells by cytotoxic T lymphocytes (CTLs). This might open new opportunities for the treatment of cancer or persistent viral infections, for which induction of strong cellular immunity seems to be essential. 1 Dendritic cells (DCs) are professional APCs that play a key role in regulating adaptive immunity, and both preclinical and clinical studies have exploited DCs in an attempt to induce antiviral or antitumor CTL responses. Most of these studies explored ex vivo Ag loading of autologous monocyte-derived DCs that were readministered to the patient, a laborious and costly procedure. The discovery of pattern recognition receptors, such as C-type lectin receptors (CLRs) that mediate Ag uptake, allow for a more direct strategy by targeting Ags to DCs in vivo. Some of these CLRs are restricted to APCs, such as macrophages or DCs. Therefore, targeted delivery exploiting CLRs has already been demonstrated to enhance Ag presentation and results in immunity when DC maturation stimuli such as TLR ligands (TLRLs) are co-administered. 2 TLRs constitute a family of pattern recognition receptors that recognize pathogen-associated molecular patterns and trigger immune cell activation. Consequently, natural TLRLs and their mimetics are increasingly being applied in immunotherapeutic strategies. 3 TLRLs are potent inducers of innate immune responses and have been evaluated as monotherapies to treat infection and cancer. 4,5 In addition, TLRLs have been applied as adjuvants to stimulate adaptive immune responses. 3 We hypothesized that targeted delivery of both Ag and TLRLs directly to DCs may greatly enhance immune responses. This approach would guarantee that the APCs that take up Ag are...

Purpose: It is unknown whether the route of administration influences dendritic cell (DC)-based immunotherapy. We compared the effect of intradermal versus intranodal administration of a DC vaccine on induction of immunologic responses in melanoma patients and examined whether concomitant administration of interleukin (IL)-2 increases the efficacy of the DC vaccine.Experimental Design: HLA-A2