(10c) Altering Vaccine Placement of Cytotoxic and Helper T Cell Antigens Influences Immunological Activation

Spherical nucleic acids (SNAs) are nanomaterials that can improve the delivery and potency of vaccine components. Comprised of a nanoparticle core with a dense surface layer of radially-oriented oligonucleotides (e.g. unmethylated cytosine-phosphate-guanine (CpG) motif DNA, which agonizes toll-like receptor 9 (TLR9) in antigen-presenting cells (APCs)), SNAs are modular structures with several advantages over conventional vaccines. They have high affinity target binding, rapid cellular uptake without the need for transfection reagents, high biocompatibility, reduced nuclease degradation, and easy drainage to the lymph nodes upon subcutaneous injection.^1–6 Their ease of synthesis and inherent modularity enable the design of compositionally equivalent structures with varied arrangement of the two key components—oligonucleotide adjuvant (immune system activator) shell and antigen (immune system target). Structural modifications have previously shown that vaccine architecture affects function and produces distinct immune responses; this has thus far been demonstrated in tumor models including model ovalbumin (OVA) lymphoma tumors and HPV, as well as prostate cancer-relevant systems targeting prostate-specific membrane antigen (PSMA).^6,7 However, these results only incorporate antigens activating one class of immune cells: cytotoxic CD8⁺ T cells.

Due to a tumor’s heterogeneity and ability to evolve and evade the immune system, it is important not only to generate an enhanced major histocompatibility complex I (MHC-I) targeting cytotoxic CD8⁺ T cell response, but also engage synergistic interactions. In particular, both CD8⁺ and helper CD4⁺ T cells are necessary for long-lasting tumor rejection.^8,9 Therefore, in this work, I hypothesized that one can enhance vaccine efficacy by simultaneously using MHC-I and -II antigens to prime both CD8⁺ and CD4⁺ T cells. This is especially important in melanoma, where traditional treatments such as chemotherapy or radiation are less effective, because of the tumor’s ability to easily evade the immune system. However, current clinical alternatives are either ineffective at raising broad responses capable of handling this evasion, logistically difficult, or expensive, and none consider the impact of structure on the resulting immune response. SNAs function as robust cancer vaccines by controlling the presentation of immunostimulatory cues and target multiple melanoma-associated antigens in an effort to lower tumor immune evasion. This work utilizes the rational vaccinology approach to improve vaccine potency by presenting multiple epitopes in a specific structural arrangement to stimulate both cytotoxic and helper T cells.

In this work, I have found that the structural placement of these two classes of antigens (one targeting CD8⁺ and another targeting CD4⁺ T cells) dramatically alters the efficacy of a vaccine by inducing changes at the genetic, cellular, and organismal levels. I observed that compositionally equivalent SNAs with different antigen placement direct different immune responses, which are changed at the transcriptome level. One particular SNA structure, with the MHC-I antigen hybridized to the shell and the MHC-II antigen encapsulated in the core, elicited increases in theeffector state of cells and anti-tumor cytokine production compared to the inverse structure with swapped placement of the MHC-I and -II antigens (Figure 1) and a simple mixture of both antigens and adjuvant. Importantly, these immunological trends translate to an in vivo melanoma system, where the different SNAs induce significant differences in tumor growth.

This research highlights how the heterogeneity of tumors can be addressed through the rational design of more complex vaccines. By incorporating this approach and considering antigen placement in vaccine design, I successfully demonstrate how an SNA vaccine’s potency can be altered. This concept is broadly applicable to the field, as it addresses the opportunity to use nanostructures to present and coordinate the processing of multiple immunostimulatory cues to immune cells. Importantly, this knowledge is translatable to other systems and biological knowledge, as it informs the mechanistic understanding of the structural basis for vaccine function.

References:

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(2) Banga, R. J.; Chernyak, N.; Narayan, S. P.; Nguyen, S. T.; Mirkin, C. A. Liposomal Spherical Nucleic Acids. J. Am. Chem. Soc. 2014, 136 (28), 9866–9869.

(3) Radovic-Moreno, A. F.; Chernyak, N.; Mader, C. C.; Nallagatla, S.; Kang, R. S.; Hao, L.; Walker, D. A.; Halo, T. L.; Merkel, T. J.; Rische, C. H.; Anantatmula, S.; Burkhart, M.; Mirkin, C. A.; Gryaznov, S. M. Immunomodulatory Spherical Nucleic Acids. Proc. Natl. Acad. Sci. 2015, 112 (13), 3892–3897.

(4) Choi, C. H. J.; Hao, L.; Narayan, S. P.; Auyeung, E.; Mirkin, C. A. Mechanism for the Endocytosis of Spherical Nucleic Acid Nanoparticle Conjugates. Proc. Natl. Acad. Sci. 2013, 110 (19), 7625–7630.

(5) Rosi, N. L. Oligonucleotide-Modified Gold Nanoparticles for Intracellular Gene Regulation. Science 2006, 312 (5776), 1027–1030.

(6) Wang, S.; Qin, L.; Yamankurt, G.; Skakuj, K.; Huang, Z.; Chen, P.-C.; Dominguez, D.; Lee, A.; Zhang, B.; Mirkin, C. A. Rational Vaccinology with Spherical Nucleic Acids. Proc. Natl. Acad. Sci. 2019, 116 (21), 10473–10481.

(7) Qin, L.; Wang, S.; Dominguez, D.; Long, A.; Chen, S.; Fan, J.; Ahn, J.; Skakuj, K.; Huang, Z.; Lee, A.; Mirkin, C.; Zhang, B. Development of Spherical Nucleic Acids for Prostate Cancer Immunotherapy. Front. Immunol. 2020, 11, 1333.

(8) Ostroumov, D.; Fekete-Drimusz, N.; Saborowski, M.; Kühnel, F.; Woller, N. CD4 and CD8 T Lymphocyte Interplay in Controlling Tumor Growth. Cell. Mol. Life Sci. 2018, 75 (4), 689–713.

(9) Shankaran, V.; Ikeda, H.; Bruce, A. T.; White, J. M.; Swanson, P. E.; Old, L. J.; Schreiber, R. D. IFNγ and Lymphocytes Prevent Primary Tumour Development and Shape Tumour Immunogenicity. Nature 2001, 410 (6832), 1107–1111.