Prognostic implication of histological features associated with EHD2 expression in papillary thyroid carcinoma

Abstract

Papillary thyroid carcinoma (PTC) is a heterogeneous tumor with various histological and molecular subtypes. EHD2 is involved in endocytosis and endosomal recycling. This study aimed to investigate the prognostic significance of EHD2 expression in PTC and develop a new model for predicting persistent/recurrent disease after thyroidectomy. Pathologic slides of 512 consecutive patients with PTC ≥ 1 cm were retrospectively reviewed. BRAF mutation analysis and immunohistochemistry for EHD2 were performed. Clinical significance of EHD2 mRNA expression was analyzed in 388 PTC patients using The Cancer Genome Atlas dataset. The presence of dyscohesive cells and psammoma bodies were found have significant association with persistent/recurrent disease (p = 0.049 and p = 0.038, respectively). The best discrimination of disease-free survival was found by dividing patients into three prognostic groups based on the following two risk factors according to the size category: psammoma bodies ≥ 4 and dyscohesive cells (≥ 1% and ≥ 20% in PTCs of < 2.0 cm and ≥ 2.0 cm, respectively). In PTCs of ≥ 2.0 cm, patients with the two risk factors had a hazard ratio of 13.303 (p = 0.005) compared to those without risk factors. High expression level of EHD2 was associated with BRAF V600E (p < 0.001), presence of dyscohesive cells (p = 0.010), and absence of psammoma bodies (p = 0.001). Increased EHD2 mRNA expression level was associated with extrathyroidal extension (p < 0.001), pT3-4 (p < 0.001), lymph node metastasis (p < 0.001), higher risk of recurrence (p < 0.001), and BRAF V600E (p < 0.001). Our prognostic model is useful for predicting persistent/recurrent disease after surgery of PTC. EHD2 mRNA expression could be a novel prognostic marker for PTC patients.

Fig 1. Histologic features of dyscohesive cells and psammoma bodies.

(A) Dyscohesive cells predominantly seen at the invasive front of the papillary carcinoma. (B) High magnification of the invasive front showing isolated single and loosely cohesive clusters of polygonal shaped tumor cells with abundant cytoplasm. (C) Dyscohesive cells often accompanied by micropapillary pattern. (D) Higher magnification revealing dyscohesive cells and micropapillary structure. (E) Psammoma bodies found at the cores of papillae and stroma of tumor. (F) A case of papillary carcinoma with numerous psammoma bodies.

Fig 2. Relationship between grade of dyscohesive cells and clinicopathologic features in all 512 patients with papillary thyroid carcinoma.

Fig 3. Relationship between grade of psammoma bodies and clinicopathologic features in all 512 patients with papillary thyroid carcinoma.

Fig 4. Kaplan-Meiyer analyses of disease-free survival of all 512 patients with papillary thyroid carcinoma (PTC) according to dyscohesive cells and psammoma bodies.

Dyscohesive cells (A) and psammoma body (B) predicted disease-free survival following surgical resection of PTC. (A) p = 0.033 between 0% and 1%-10% subgroups; p = 0.329 between 0% and 11%-19% subgroups; p = 0.036 between 0% and ≥ 20% subgroups; p = 0.449 between 1%-10% and 11%-19% subgroups; p = 0.463 between 1%-10% and ≥ 20% subgroups; p = 0.307 between 11%-19% and ≥ 20% subgroups; (B) p = 0.090 between 0 and 1–3 subgroups; p = 0.005 between 0 and ≥ 4 subgroups; p = 0.381 between 1–3 and ≥ 4 subgroups. (C) Results of disease-free survival of patients who were subdivided into three groups based on risk factors of dyscohesive cells ≥ 1% and psammoma body ≥ 4. p = 0.069 between 0 risk factors and 1 risk factor; p = 0.004 between 0 risk factors and 2 risk factors; p = 0.064 between 1 risk factor and 2 risk factors.

Fig 5. Subgroup analyses of disease-free survival of patients with papillary thyroid carcinoma (PTC) according to tumor size category and different cutoff of dyscohesive cells.

(A) Results of disease-free survival of 402 patients with PTC between 1.0 cm and 2.0 cm. Patients were subdivided into three groups based on risk factors of dyscohesive cells ≥ 1% and psammoma body ≥ 4. p = 0.204 between 0 risk factors and 1 risk factor; p = 0.011 between 0 risk factors and 2 risk factors; p = 0.091 between 1 risk factor and 2 risk factors. (B) Results on disease-free survival of 110 patients with PTC ≥ 2.0 cm. Patients were subdivided into three groups based on risk factors of dyscohesive cells ≥ 20% and psammoma body ≥ 4. p = 0.081 between 0 risk factors and 1 risk factor; p = 0.010 between 0 risk factors and 2 risk factors; p = 0.345 between 1 risk factor and 2 risk factors.

Fig 6. Immunohistochemical staining for EHD2 protein.

The immunohistochemical results showing no expression (A), weak expression (B), moderate expression (C), and strong expression (D) in tumor cells. Endothelial cells (internal control) are positive in all staining.

Fig 7. Correlation between EHD2 expression levels and histological parameters.

(A) Positive correlation of expression levels of EHD2 in all tumor cells with grades of dyscohesive cells in tumor periphery and invasive front. (B) Negative correlation of EHD2 expression in all tumor cells with the number of psammoma bodies in the entire tumor tissue.

Fig 8. Relative expression levels of EHD2 mRNA in TCGA dataset.

Scatter dot plots (median with interquartile range) showed that the EHD2 mRNA expression was associated with histologic subtypes (A), extrathyroidal extension (B), pT stage (C), lymph node metastasis (D), risk of recurrence (E), and molecular subtype (F). EDH2 mRNA expression level was negatively correlated with thyroid differentiation score (G) but positively correlated with ERK score (H).