Head and neck squamous carcinomas with exophytic and endophytic type of growth have the same prognosis after surgery and adjuvant radiotherapy

Abstract

Head and neck squamous cell carcinomas (HNSCC) are characterized by exophytic or endophytic growth. We hypothesized that the growth pattern predicts outcome and associates with distinct clinical and immunological profiles. Tumors obtained from 60 HNSCC patients treated with surgery and adjuvant radiotherapy were identified as exophytic or endophytic. Recurrence-free survival (RFS) at 42 months was determined. In a subsets of 30 patients (22 exophytic and 8 endophytic) tumor stroma and parenchyma were evaluated for infiltrating CD4(+) and CD8(+) T, dendritic, myeloid and FOXP3(+) regulatory T cells (Treg) and expression of immunosuppressive cytokines by immunohistochemistry. The localization and frequency of positive cells were determined microscopically and analyzed by hierarchical clustering to distinguish exophytic versus endophytic tumors. 34/60 patients had exophytic and 26/60 endophytic tumors. No differences in clinicopathologic data, disease progression or RFS were seen between the two cohorts. Infiltrates of CD3(+)CD8(+) T cells were larger in endophytic than exophytic tumors, while FOXP3(+) Treg, TGF-β(+), IL-10(+), Arg-1(+), CD11b(+) cells were equally prominent in both. FOXP3(+) Treg accumulated in endophytic tumor nests, while the exophytic tumor stroma was enriched in IL-10(+) cells (both at p < 0.05). Hierarchical clustering based on immunophenotyping failed to identify different clusters in these two tumor types. However, CD68(+) macrophages and FOXP3(+) Treg showed a distinct distribution. The HNSCC growth pattern did not predict RFS. Although higher numbers and differences in localization of immunosuppressive cells in endophytic versus exophytic tumors were observed, no significant relationship was established between the growth pattern and the immune profile of infiltrating lymphocytes.

Fig. 1

Comparison between exophytic and endophytic type of tumor growth in microlaryngoscopy modo Kleinsasser. Exophytic tumors grow out from the mucosal surface in cauliflower-like clusters, while endophytic tumors crawl under the mucosa, carpeting the inferior of the distinct local structure.

Fig. 2

Recurrence-free survival by the tumor growth pattern. Kaplan-Meier curves were used to determine recurrence-free survival (RFS) of HNSCC patients (n=60) with exophytic vs. endophytic tumors.

Fig. 3

Recurrence-free survival by nodal involvement and the tumor growth pattern. (a) Kaplan-Meier curves for RFS of HNSCC patients (n=60) by their nodal status. (b) Kaplan-Meier curves for recurrence-free survival by the nodal status and the tumor growth pattern.

Fig. 4

Immune infiltrates in exophytic vs. endophytic HNSCC. H&E staining and immunohistochemistry of paraffin embedded tissue sections of representative tumors at × 400 mag. In addition to inflammatory cells (CD3⁺, CD4⁺, CD8⁺), staining for CD34⁺ cells indicates localization of blood vessels.

Fig. 5

Inflammatory cells in the tumor stroma vs. tumor nests. (a) Numbers of immune cells positive for various surface markers in the tumor stroma and tumor nests (n=30). Asterisks indicate significant differences at p<0.05 to p<0.001. (b) Absolute numbers of immune cells positively stained for different markers in the stroma and nests in tumors with exophytic (n=22) and endophytic (n=8) tumors. On the right, representative tumor sections showing the presence of IL-10⁺ and FOXP3⁺ cells in the tumor stroma or tumor nests. Mag ×200 for IL-10⁺ and ×400 for FOXP3 staining.

Fig. 6

Hierarchal clustering for numbers of selected infiltrating immune cells found in the tumor stroma or tumor nests in exophytic and endophytic tumors.

Fig. 7

Localization of inflammatory cells in tumor tissues in exophytic and endophytic tumors. Representative immunohistochemistry staining of paraffin embedded tissue sections for CD68⁺ (mag ×200) and FOXP3⁺ cells (mag ×400).