الفهرس 
INTRODUCTIONOnly 14 pages are availabe for public view
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Abstract
Immunotherapy has emerged as a promising weapon in the fight against cancer. Toll-like receptors (TLRs) are pattern recognition receptors expressed on immune cells or non-immune cells such as epithelial cells lining the oral and gut mucosa, as well as cancer cells. TLR agonists such as BCG, Imiquimod and MPLA have been approved by the FDA as immunotherapeutic drugs. TLR signaling activation may either achieve anti-or pro-tumor effect. Furthermore, the downstream effect of such activation is still an understudied field. One of the possible outcomes is cellular autophagy. It is a cellular degradation system responsible for cellular homeostasis, it also serves as a cellular response against stress. It has been reported that TLR-signaling and autophagy interact in various complex mechanisms. Further insight into these mechanisms and complexities could present enormous potential in paving the way for the success of immunotherapy.
Study objective: The present work is designed to investigate the therapeutic outcome of TLR agonists (BCG, Imiquimod and MPLA) in OSCC, both in vitro and in vivo in relation to autophagy. The null hypothesis denotes that their therapeutic effect is not related to autophagy.
Methodology: This work includes an in vitro study followed by an in vivo study. The in vitro Study was carried out as duplicate examination. First, the SCC-4 cell line was established. Second, cytotoxicity assay was done to determine the IC50 for TLR agonists BCG, Imiquimod and MPLA. Followed by TLR2, 4 and 7 active expression verification for TLR agonists that displayed a cytotoxic effect; namely, BCG and Imiquimod. Lastly, flow cytometry was used to quantify phenotypes associated with autophagy using Human LC3B Alexa Fluor® 594‑conjugated Antibody, upon treatment with BCG, Imiquimod and Cisplatin, as well as the negative control group. The in-vivo study: Forty-one Syrian golden hamsters were randomly divided into 7 groups of drugs, drug combinations and controls. The seven groups were as follows: BCG, Imiquimod, BCG+IMQ, MPLA+IMQ, MPLA+BCG, positive control Cisplatin and negative control that received no treatment. OSCC was chemically induced in the hamsters’ left buccal pouches. Different drugs were administered according to the proposed treatment plan. Clinical and histopathologic evaluation of the seven groups was conducted. Finally, different groups were assessed for autophagy expression using MDC stain from the autophagy detection kit.
Results: The in-vitro study reveals that BCG and Imiquimod have cytotoxic effects on SCC-4 cells, while MPLA does not. Both TLRs agonists BCG and Imiquimod increase TLRs 2, 4, and 7 expressions, and induce autophagy at various time points with Imiquimod demonstrating the highest rate of autophagy induction. The in-vivo study shows that BCG, Imiquimod, and their combination effectively treat induced hamster buccal pouch carcinoma, positively affecting tumor volume and increasing life span. These TLR agonists exhibit a therapeutic effect superseding Cisplatin, with lower mortality rates. They also trigger autophagy in a significantly higher manner than negative controls. MPLA-IMQ and MPLA-BCG combinations have conflicting clinical results regarding different parameters.
Conclusion: TLR agonists BCG and Imiquimod exhibit a novel off-label anti-carcinogenic effect on oral squamous cell carcinoma cells both in-vitro and in-vivo, and that effect is mediated, at least partially via autophagy occurring downstream of TLR signaling.
Key words: Oral squamous cell carcinoma (OSCC), Toll like receptor (TLR), autophagy, BCG, Imiquimod, MPLA.