Caracterización del Inmunofenotipo para el EGFR, TBXA2R Y SMAD3 En Arreglos de Tejido para el Diagnóstico Diferencial Entre Adenoma e Hiperplasia Paratiroidea en una Muestra de Pacientes Intervenidos Quirúrgicamente por HPTP no Familiar

Abstract

Backround: Primary hyperparathyroidism (pHPT) is a relatively common endocrinophathy. Its real incidence resides among women and people older than 60 years of age. It occurs as a consequence of an excessive secretion of parathyroid hormone (PTH) by one or more of the parathyroid glands. It can either be asymptomatic, or be associated with a variety of symptoms. Around 80% of the patients with pHPT have unique glandular enlargement, also called adenoma; between 10-15% of the patients have multiglandular disease caused by sporadic (nonfamilial) parathyroid hyperplasia (SPH). The standard treatment is surgery, with consists in the removal of the affected gland (s). In a considerable percentage of these patients, the diagnosis changes during surgery or follow-up, which in some cases, the patient has to have a second surgery, thus increasing the morbidity and costs. For this reason to identify and confirm the diagnosis of these two entities has both medical value (prognostic and diagnostic), as well as economic impact for patients with pHPT. Objective.- To determine if there are differences in the immunophenotype for the biomarkers EGFR, TBXA2R, and SMAD3, in order to allow us to differentiate categorically between an adenoma and parathyroid hyperplasia in a sample of Mexican subjects which had surgery secondary to sporadic pHPT at our institution. Methods.- A total of 49 samples were included: 37 adenomas and 12 parathyroid hyperplasias. Consent form was signed by the subjects. All the patients had surgery due top HPT at the INCMNSZ between 1991 and 2009. We carried out microarrays (7x8) consisting of tissue samples obtained of the paraffin block of the patients organized in arrays of 2 mm in diameter. The data was reviewed in a retrospective manner initially, and prospectively later on to evaluate clinical behavior. Immunohistochemisty was performed with antigenic retrieval by protease and antigen-antibody reaction using peroxidase-DAB blockade. We used specific policlonal antibodies for EGFR, TBXA2R, and SMAD3. Immunophenotypes were evaluated by an expert pathologist, which was blinded to the final phenotypes. The readings were expressed in a qualitative way. A descriptive and bivariate analysis of all the variables and their association with the different immunophenotypes was carried out using SPSS versión 13.0 and Excell Office. A p≤0.05 was considered statistically significant. Results.- 82% were female with a mean age of 52 (30-77 years). Adenomas were localized preoperatively in 75.7% of the cases. All the hyperplasias were bilateral. Preoperative serum calcium was 11.5+1.2 and 10.6+1.2 mg/dl in adenomas and hyperplasias respectively, while preoperative PTH was 334.7+425.7 vs 326.9+209.8 pg/ml respectively. PTH was slightly superior after surgery in patients with hyperplasia, but was not statistically significant. Preoperative urinary calcium and surgical time were variables that differed statistically between adenomas and hyperplasias (p≤0.05). 32.4% of the adenomas were positive for EGFR and TBXA2R, while in the hyperplasias only 8.7% and 16.7% were positive to both markers respectively. Opposite to the previous markers, 90% of the adenomas and hyperplasias were negative for SMAD3. Conclusions.-. We demonstrated a fast and efficient way to evaluate tissue arrays in up to 56 patients simultaneously. Our results also showed a tendency towards a phenotypical differentiation between the two pathologies for our 3 studied biomarkers. Nevertheless, none of our results was statistically significant probably due to the size of the sample. With all these data we conclude that a single marker use is limited and insufficient. Therefore we recommend to identify a “signature” composed of a combination of different biomarkers. We need prospective evaluation in a bigger sample, as well as the use of quantitative evaluation such as qRT-PCR for mRNA.