**1. Introduction**

Cancer immunotherapy is an emerging antitumor treatment technique, which works via specific antigen-mediated modulation in the patient's immune system [1]. Conventional anticancer therapies, including chemotherapeutic drugs and targeted treatments, have clinical limitations and adverse effects, such as non-specificity, drug resistance, and low efficacy in cancer mutation and metastasis [2–4]. Hence, the precise control and modulation of adaptive immune responses for pre-existing intratumoral therapy is the most important engineering parameter for developing effective cancer immunotherapeutic approaches [5–7].

Based on the interplay between T cell populations and other immune cellular components, engineering modulation in adaptive immunity could effectively eliminate cancer cells and inhibit tumor growth. As depicted in Figure 1, in particular, CD4<sup>+</sup> T cells differentiate into various T helper cell subsets, including T helper (Th)1, Th2, Th9, Th17, and T follicular helper cells, in which Th1 cells react with antigen-presenting cells (APCs) and indirectly assist in the differentiation of CD8<sup>+</sup> T cells into cytotoxic T lymphocytes (CTLs) by secreting a cytokine, such as interferon (INF)-γ. Additionally, interleukin (IL)-2 secreted from Th1 induces the proliferation of CD8<sup>+</sup> T cells [8,9]. Thus, both activated CD4<sup>+</sup> and CD8<sup>+</sup> T cells augment long-lasting and strong antitumor immune responses by generating memory T cells to persist in anamnestic immune responses. APCs such as macrophages and dendritic cells (DCs) are crucial mediators for inducing T cell activation by antigen presentation on their surfaces. After taking up the tumor antigen molecules, these antigens are processed in the proteasome or phagosomes in the cytosol and presented in the form of peptides via major histocompatibility complex (MHC) class I or II molecules on the cellular

**Citation:** Seong, J.; Kim, K. Activation of Cellular Players in Adaptive Immunity via Exogenous Delivery of Tumor Cell Lysates. *Pharmaceutics* **2022**, *14*, 1358. https://doi.org/10.3390/ pharmaceutics14071358

Academic Editors: Lúis Alexandre Muehlmann and João Paulo Figueiró Longo

Received: 29 April 2022 Accepted: 22 June 2022 Published: 27 June 2022

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**Copyright:** © 2022 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/).

surface of APCs. APCs with the MHC–peptide complex travel to secondary lymphoid organs to stimulate T cells. Upon contacting T cells, APCs initiate the priming of naïve T cells by interacting with the MHC–peptide complex and T cell receptor (TCR), and secrete cytokines to activate T cells. II molecules on the cellular surface of APCs. APCs with the MHC–peptide complex travel to secondary lymphoid organs to stimulate T cells. Upon contacting T cells, APCs initiate the priming of naïve T cells by interacting with the MHC–peptide complex and T cell receptor (TCR), and secrete cytokines to activate T cells.

**Figure 1.** Overview of DC and T cell interplay for anticancer immunotherapy. (**I**) Differentiation of stimulated immature DCs (iDCs) by DAMP molecules and TAAs to mature DCs (mDCs), (**Ⅱ**) process of antigen presentation via MHC molecules in DCs, (**Ⅲ**) priming of T cells to effector T cells, (**Ⅳ**) induction of the tumor cell death by various types of T cells. Reproduced with permission from [10]; images used are from Servier Medical Art. **Figure 1.** Overview of DC and T cell interplay for anticancer immunotherapy. (**I**) Differentiation of stimulated immature DCs (iDCs) by DAMP molecules and TAAs to mature DCs (mDCs), (**II**) process of antigen presentation via MHC molecules in DCs, (**III**) priming of T cells to effector T cells, (**IV**) induction of the tumor cell death by various types of T cells. Reproduced with permission from [10]; images used are from Servier Medical Art.

Previous studies have experimentally demonstrated that APC–T cell interaction by three distinct signals effectively induced antigen-specific T cell activation [11,12]: (1) the interaction between antigenic peptides presented by the MHC and the TCR; (2) costimulatory signals induced by the interaction between B7 molecules (e.g., CD80 and CD86) in APCs and CD28 in T cells, which trigger stronger immune responses; and (3) polarizing signals mediated by the production and secretion of multiple cytokines (e.g., IL-12 and tumor necrosis factor (TNF)-α by APCs [13]. Among these signaling interactions, MHC-mediated antigen cross-presentation is the most critical for the initiation of antigen-specific immune responses. Presented MHC I-immunogenic peptide complexes can be recognized by CTLs [14], whereas T helper cells can be activated by MHC II-mediated extracellular (or exogenous), immunogenic, antigenic peptide complex presentation [15]. Finally, activated T cell populations migrate toward the tumor microenvironment to kill specific tumor cells [16]. Cancer cells overexpress universal tumor-associated antigens (TAAs) and individual mutant neo-antigens [17,18]. To incorporate these TAAs for facilitating APC-dependent antigen presentation and subsequent T cell activation, lysed tumor cell bodies containing soluble tumor antigen molecules (e.g., tumor cell lysates (TCLs)) have been investigated for cancer therapy. Previous studies have experimentally demonstrated that APC–T cell interaction by three distinct signals effectively induced antigen-specific T cell activation [11,12]: (1) the interaction between antigenic peptides presented by the MHC and the TCR; (2) co-stimulatory signals induced by the interaction between B7 molecules (e.g., CD80 and CD86) in APCs and CD28 in T cells, which trigger stronger immune responses; and (3) polarizing signals mediated by the production and secretion of multiple cytokines (e.g., IL-12 and tumor necrosis factor (TNF)-α by APCs [13]. Among these signaling interactions, MHC-mediated antigen cross-presentation is the most critical for the initiation of antigen-specific immune responses. Presented MHC I-immunogenic peptide complexes can be recognized by CTLs [14], whereas T helper cells can be activated by MHC II-mediated extracellular (or exogenous), immunogenic, antigenic peptide complex presentation [15]. Finally, activated T cell populations migrate toward the tumor microenvironment to kill specific tumor cells [16]. Cancer cells overexpress universal tumor-associated antigens (TAAs) and individual mutant neo-antigens [17,18]. To incorporate these TAAs for facilitating APC-dependent antigen presentation and subsequent T cell activation, lysed tumor cell bodies containing soluble tumor antigen molecules (e.g., tumor cell lysates (TCLs)) have been investigated for cancer therapy.

Therefore, cell-based engineering techniques to control and stimulate adaptive immune responses and further tumor suppression have recently been developed for designing efficient cancer therapeutics and vaccinations. TCLs containing various epitope sources are utilized for the induction of both CD8+ and CD4+ T cells [19,20] and potential personalized therapy. Endogenous damage-associated molecular patterns (DAMPs) are released by dying or damaged cells (i.e., host biomolecules that can initiate noninflammatory responses to infection), and these specific TAAs interact with the pattern Therefore, cell-based engineering techniques to control and stimulate adaptive immune responses and further tumor suppression have recently been developed for designing efficient cancer therapeutics and vaccinations. TCLs containing various epitope sources are utilized for the induction of both CD8<sup>+</sup> and CD4<sup>+</sup> T cells [19,20] and potential personalized therapy. Endogenous damage-associated molecular patterns (DAMPs) are released by dying or damaged cells (i.e., host biomolecules that can initiate non-inflammatory responses to infection), and these specific TAAs interact with the pattern recognition receptors (PRRs) of DCs. Sequentially, activated antigen presentation on DCs induces proper T

cell priming toward Th1 cells and differentiation of T lymphocytes into CTLs. Moreover, secreted cytokines, such as IL-12, IL-15, and IFN, from DCs are also able to stimulate T cell activation [20]. These T cells primed by APCs with TCL-derived antigens are key effectors of anticancer immunity. Antigen-specific memory T cells, which exert immediate effector functions without the need for further differentiation, sufficiently suppress tumor recurrence [21].

A growing body of research has explored the potential capability of TCLs for cancer vaccination. Because of the high antitumor immunity effects of TCLs, the majority of recent cancer immunotherapies utilize the TCL-mediated activation of APCs and T cells, along with the MHC pathway. However, due to the technical drawbacks of the naked form of TCLs, including a short half-life and the limited availability of various antigens, a lower therapeutic effect in immunity than the treatment of a specific antigen has been frequently observed. To strengthen the clinical efficacy of TCLs, particularly in the case of in vivo administration, precisely designed delivery systems should be utilized for increasing the stability of cargo TCLs and facilitating the co-administration of adjuvants. Therefore, the TAA-mediated activation of TCLs as an immune activator could be applied to induce in vitro necrosis of cancer cells, and stimulate downstream antitumor responses and immunological memory generation.

To this end, this review focuses on the current progress in engineered cancer immunotherapies by exogenous TCL delivery, emphasizing (1) practical applications using TCL-mediated DC activation and sequential stimulation in adaptive immunity in various cancer types, (2) the significance of adaptive immunological functions, and (3) the utilization of a series of delivery platforms for the co-administration of multiple adjuvants for effective cancer vaccination and antitumor treatment.
