Date of Award

May 2026

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Biomedical Sciences

First Advisor

David S. Bradley

Abstract

Immunotherapies have transformed cancer treatment by enabling durable antitumor responses through targeted activation of the immune system. Among the most potent immune-stimulatory molecules are bacterial superantigens, which activate large fractions of the T-cell repertoire through simultaneous binding of major histocompatibility complex (MHC) class II molecules and conserved regions of the T-cell receptor (TCR) β-chain. Because these interactions are strongly human MHC–restricted, HLA-transgenic mouse models are essential for evaluating the therapeutic potential and safety of superantigen-based immunotherapies.This dissertation investigates the antitumor activity and mechanisms of action of two superantigens encoded within the Staphylococcus aureus enterotoxin gene cluster—Staphylococcal enterotoxin I (SEI) and Staphylococcal enterotoxin G (SEG)—using the aggressive B16-F10 melanoma model in HLA-DQ8 transgenic mice. Prior work from our laboratory demonstrated that dual SEI+SEG therapy produced complete regression of established melanoma and elicited durable, tumor-specific memory in HLA-DQ8 mice, capable of rejecting tumor rechallenge for more than 400 days. These findings motivated a focused mechanistic investigation to define how each superantigen independently shapes antitumor immunity. Using four multiparameter flow cytometry panels, we characterized immune activation, myeloid remodeling, checkpoint regulation, proliferative status, and TCR Vβ family usage within the tumor microenvironment. The data reveal that SEI and SEG operate through distinct yet complementary immune pathways. SEI primarily remodeled the innate immune compartment by significantly reducing monocytic and granulocytic myeloid-derived suppressor cells, preserving antigen-presenting cell activation, and broadly expanding Vβ families across multiple lymphocyte subsets, consistent with wide HLA-DQ8–restricted clonal engagement. In contrast, SEG acted predominantly on the adaptive immune axis by reducing PD-1, CTLA-4, and LAG-3 expression on intratumoral T cells and selectively expanding Vβ families associated with cytotoxic effector function, including Vβ17a and Vβ8.1/8.2. Together, these findings demonstrate that SEI and SEG reprogram the tumor microenvironment through mechanistically distinct but convergent pathways: SEI disrupts suppressive myeloid networks and broadens TCR engagement, while SEG relieves inhibitory checkpoint signaling and restores adaptive T-cell functional competence through targeted Vβ activation. This integrated mechanistic framework provides a biological explanation for the synergy previously observed with SEI+SEG combination therapy and establishes a foundation for the rational development of HLA-tailored superantigen-based immunotherapies for melanoma and potentially other solid tumors.

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