Characterizing the Acute Immune Response to Cutaneous Peptide Vaccination

Abstract

Therapeutic vaccination is a promising modality for inducing antigen-specific immune responses against cellular targets. In particular, MHC-restricted target epitopes formulated into synthetic long peptides (SLPs) are an effective format for delivering vaccine antigens to antigen-presenting cells (APCs). Coupled with adjuvants such as poly(I:C), a potent agonist of the pattern-recognition receptors TLR3 and MDA5, SLPs may be administered cutaneously to induce T cell responses in the context of cancer, autoimmunity, and infectious diseases. However, many aspects of the acute cellular response to peptide vaccination remain uninterrogated. Understanding how SLP vaccines mediate a therapeutic response could inform future efforts to develop more efficacious immunotherapies. To characterize the acute cellular response to SLP vaccination in vivo, a mouse model of cutaneous vaccination using C57BL/6J mice and the model antigen ovalbumin (OVA) was employed. At multiple timepoints over 72 hours after vaccination, tissue samples were extracted and a tissue digestion protocol was developed to enable the preparation of single-cell suspensions for further analysis. Initially, flow cytometry was used to characterize APC populations activated and enriched by vaccination. Subsequently, unbiased single-cell transcriptomic technology was used to dissect the heterogeneity and identities of vaccine-responsive cells following exposure to diverse classes of adjuvants (poly-ICLC, STING agonist, and TLR4 agonist). These analyses revealed that multiple cell subsets located at both the cutaneous vaccine site and the skin-draining lymph node are acutely activated by SLP vaccination. Furthermore, kinetics of the vaccine response by various immune cell subsets was determined by comparing samples at different timepoints after vaccination. Finally, the potency of poly(I:C) as a primary vaccine adjuvant was validated, with additional evidence generated to support that additional adjuvants added to the vaccine formulation may augment the immune response. These findings help elucidate the acute timeline of response to peptide vaccination and may support further translational vaccine development efforts, particular in the context of tumor vaccines.