PROJECT ABSTRACT/SUMMARY Substantial pre-clinical evidence has demonstrated stress-induced tumor progression in animal models of head and neck cancer (HNC). Clinically, patients with locoregionally advanced Human Papillomavirus (HPV)-negative HNC are exposed to a broad range of chronic stressors. The link between cancer progression and chronic stress may be one contributing explanation for the persistently poor oncologic outcomes of the HPV-negative HNC population. To evaluate mechanism and develop therapeutics, animal models of chronic stress are needed. However, existing models use either immunodeficient mice or syngeneic murine tumors, neither of which recapitulate the human tumor microenvironment (TME). To overcome these limitations, we developed a novel mouse model of chronic stress engineered to have a human immune system and carry human HNC xenografts. In preliminary studies using this model, chronic stress led to increased tumor growth and metastases associated with higher levels of circulating stromal-derived factor 1 (SDF-1) and immune alterations in the TME. Taken together, we hypothesize that chronic stress leads to SDF-1/CXCR4 pathway upregulation which induces an immunotolerant TME and increased tumor growth and metastasis in HNC. To test this hypothesis and determine clinical relevance, we will investigate the mechanisms underlying stress-induced tumor growth in both cell line-derived xenograft (CDX) and patient-derived xenograft (PDX) humanized models of HNC. We will further evaluate clinical relevance through a prospective observational study in patients undergoing HNC surgery. In Aim 1, humanized CDX mice will be randomized to chronic social isolation stress or control conditions. Three arms will be designed including (1) animals treated with a CXCR4 antagonist, (2) HNC xenografts with a CXCR4 deletion, and (3) a control arm with wild-type HNC xenografts. Tumor growth and lung metastases will be measured. Intra-tumoral immunophenotyping and RNA sequencing will be performed. In Aim 2, we will recruit patients with locoregionally advanced HPV-negative HNC planned for surgical resection, a population with a poor prognosis and significant baseline exposure to chronic stressors. Patients will undergo plasma cytokine assessment and tumor resection specimens will undergo immunophenotyping and RNA sequencing. In a subset of these patients, autologous humanized patient-derived xenograft (PDX) models will be generated, in which the human immune system and tumor are derived from the same patient. This is uniquely feasible for HNC surgery with fibula reconstruction, as a segment of fibula bone is routinely discarded from which hematopoietic stem cells can be harvested for autologous humanization. These autologous PDX models will be randomized to chronic stress and control conditions and treated with a CXCR4 antagonist as compared with control injection. This study will be the first to investigate the mechanism of stress-induced HNC growth i...