Background: Head and neck squamous cell carcinomas (HNSCC) are phenotypically and molecularly heterogeneous and frequently develop therapy resistance. Reliable patient-derived 3D tumor models are urgently needed to further study the complex pathogenesis of these tumors and to overcome treatment failure. Methods: We developed a three-dimensional organotypic co-culture (3D-OTC) model for HNSCC that maintains the architecture and cell composition of the individual tumor. A dermal equivalent (DE), composed of healthy human-derived fibroblasts and viscose fibers, served as a scaffold for the patient sample. DEs were co-cultivated with 13 vital HNSCC explants (non-human papillomavirus (HPV) driven, n = 7; HPV-driven, n = 6). Fractionated irradiation was applied to 5 samples (non-HPV-driven, n = 2; HPV-driven n = 3). To evaluate expression of ki-67, cleaved caspase-3, pan-cytokeratin, p16^INK4a, CD45, ∝smooth muscle actin and vimentin over time, immunohistochemistry and immunofluorescence staining were performed Patient checkup data were collected for up to 32 months after first diagnosis. Results: All non-HPV-driven 3D-OTCs encompassed proliferative cancer cells during cultivation for up to 21 days. Proliferation indices of primaries and 3D-OTCs were comparable and consistent over time. Overall, tumor explants displayed heterogeneous growth patterns (i.e., invasive, expansive, silent). Cancer-associated fibroblasts and leukocytes could be detected for up to 21 days. HPV DNA was detectable in both primary and 3D-OTCs (day 14) of HPV-driven tumors. However, p16^INK4a expression levels were varying. Morphological alterations and radioresistant tumor cells were detected in 3D-OTC after fractionated irradiation in HPV-driven and non-driven samples. Conclusions: Our 3D-OTC model for HNSCC supports cancer cell survival and proliferation in their original microenvironment. The model enables investigation of invasive cancer growth and might, in the future, serve as a platform to perform sensitivity testing upon treatment to predict therapy response.

Advanced tumors of the head and neck are challenging for the treatment specialist due to the need to synergize oncological and functional requirements. Free flap reconstruction has been established as the standard of care for defects following tumor resection. However, depending on the affected anatomic subsite, advanced tumors may impose specific difficulties regarding reconstruction, especially when full-thickness resection is required. This study aimed to evaluate reconstructive strategies and oncological outcomes in patients with full-thickness resection of the oral cavity. A total of 33 patients with extensive defects due to squamous cell carcinoma of the oral cavity were identified. Indications, reconstructive procedures, and clinical outcome were evaluated. Thirty-two patients (97%) presented locally advanced tumors (T3/T4). Complete tumor resection was achieved in 26 patients (78.8%). The anterolateral thigh flap was the most frequently used flap (47.1%), and the primary flap success rate was 84.8%. The cohort demonstrated a good local control rate and moderate overall and progression-free survival rates. Most patients regained full competence regarding oral alimentation and speech. Full-thickness tumor resections of the head and neck area may be necessary due to advanced tumors in critical anatomic areas. In many cases, radical surgical treatment leads to good oncological results. Free flap reconstruction has been shown to be a suitable option for extensive defects in aesthetically challenging regions.

Surgery is generally accepted as standard treatment in oral cancer, but the reconstructive procedures remain a matter of debate. The aim of this study was to evaluate oncological outcome and quality of life following surgical resection and free-flap reconstruction in patients with early oral squamous cell carcinoma. The presented trial was performed as a prospective, single-center observation study. Inclusion criteria were primary surgery in early-stage oral squamous cell carcinoma with free-flap reconstruction. Endpoints were overall and progression-free survival and quality of life up to 24 months after surgery. Twenty-six patients were included. Overall survival was 100% and progression-free survival was 92.3% in a maximum follow-up time of 21 months. Global quality of life showed no significant alteration after surgery. Patients reported a significant reduction in pain (p = 0.048) and a decreasing impairment of speech one year after surgery (p = 0.021). Free-flap reconstruction is a safe procedure that results in excellent oncological outcome and quality of life. Functional outcome is of high relevance in early-stage tumors of the head and neck and may mostly be affected by reconstructive procedures. Therefore, a prospective evaluation to explore success and the effects of surgical therapy is highly warranted.

The introduction of immune checkpoint inhibition for recurrent and metastatic head and neck cancer has brought a new treatment option for patients suffering from advanced oral cancers without a chance for curation using surgery or radiotherapy. The application of immune checkpoint inhibitors in most cases is based on the expression levels of PD-L1 in the tumor tissue. To date, there is a lack of data on the dynamic regulation of PD-L1 during disease progression. Therefore, this study aimed to evaluate the expression levels of PD-L1 in a large cohort of patients (n = 222) with oral squamous cell carcinoma including primary and recurrent tumors. Semiautomatic digital pathology scoring was used for the assessment of PD-L1 expression levels in primary and recurrent oral squamous cell carcinoma. Survival analysis was performed to evaluate the prognostic significance of the protein expression at different stages of the disease. We found a significant up-regulation of PD-L1 expression from primary disease to recurrent tumors (mean PD-L1 H-scores: primary tumors: 47.1 ± 31.4; recurrent tumors: 103.5 ± 62.8, p < 0.001). In several cases, a shift from low PD-L1 expression in primary tumors to high PD-L1 expression in recurrent tumors was identified. Multivariate Cox regression analysis did not reveal a significantly higher risk of death (p = 0.078) or recurrence (p = 0.926) in patients with higher PD-L1 expression. Our findings indicate that the exclusive analysis of primary tumor tissue prior to the application of checkpoint blockade may lead to the misjudgment of PD-L1 expression in recurrent tumors.

Triggering receptor expressed on myeloid cells 2 (TREM2) is suggested to hamper antitumor immune response in multiple cancers. However, the role of TREM2 in oral squamous cell carcinoma (OSCC) and its expression in tumor-associated macrophages (TAMs) are unknown. In this study, TREM2 expression was analyzed in the primary tumors and corresponding lymph-node metastases of OSCC patients via immunohistochemistry on tissue microarrays. Human peripheral blood mononuclear cells (PBMCs) and single-cell suspensions of tumor and healthy adjacent tissues were analyzed for the presence of TREM2⁺ macrophages and TAMs using flow cytometry. The serum levels of soluble TREM2 (sTREM2) were quantified using an enzyme-linked immunosorbent assay. High TREM2 expression was associated with advanced UICC stages (Spearman’s rank correlation (SRC), p = 0.04) and significantly reduced survival rates in primary tumors (multivariate Cox regression, progression-free survival: hazard ratio (HR) of 2.548, 95% confidence interval (CI) of 1.089–5.964, p = 0.028; overall survival: HR of 2.17, 95% CI of 1.021–4.613, p = 0.044). TREM2 expression was significantly increased in the PBMCs of OSCC patients in UICC stage IV compared with healthy controls (ANOVA, p < 0.05). The serum levels of sTREM2 were higher in advanced UICC stages, but they narrowly missed significance (SRC, p = 0.059). We demonstrated that TREM2 was multi-factorially associated with advanced stages and inferior prognosis in OSCC patients and that it could serve as a prognostic biomarker in OSCC patients. Targeting TREM2 has the potential to reshape the local and systemic immune landscape for the potential enhancement of patients’ prognosis.