Dados do Resumo

Título

MOLECULAR ANALYSIS AND FINE NEEDLE ASPIRATION: A METHOD FOR DETECTING SENTINEL LYMPH NODE METASTASES IN MELANOMA PATIENTS

Introdução

Melanoma is a potentially lethal neoplasm with an increasing incidence in Brazil and worldwide. Lymph node metastases can be diagnosed in low volume through sentinel lymph node (SL) biopsy, a minimally invasive biopsy technique. This diagnosis is crucial as a prognostic factor and an area of intense clinical research with recent advances in systemic therapies in the adjuvant and neoadjuvant setting.

Objetivo

To standardize a new technique for detection of lymph node metastasis in SL using molecular markers and aspiration biopsy in melanoma patients.

Métodos

This study was approved by the Institutional Ethics Committee under CAAE: 56753822.0.0000.5437, number 2327/2022. Participants who signed the informed consent form underwent SL biopsy during lymphoscintigraphic surgery. Before excision, fine needle aspiration biopsy (FNAB) guided by gamma probe and intraoperative ultrasound was performed. The material collected was diluted in PBS and frozen at -80°C until RNA extraction using the RecoverAll Total Nucleic Acid Isolation kit (Invitrogen) and quantification using NanoDrop 2000 (ThermoFisher). Gene expression will be assessed by droplet digital PCR (ddPCR). The excised SL were sent for histopathological diagnosis. Commercially obtained melanocytic cell lines SK-MEL-28, SK-MEL-19, UACC62, WM-35, WM-164 and A375 will be used to standardize gene expression analysis by ddPCR and as a positive control for the reactions. For the negative control, the HFF1 and HACAT lineages were cultivated.

Resultados

From April to September 2024, a total of 3.753 melanoma patients were screened through active searches. Nineteen participants were included in the study, being eleven men and eight women. Eight patients were diagnosed with superficial spreading invasive melanoma, one with acral lentiginous invasive melanoma, six with nodular invasive melanoma, one with invasive lentigo maligno melanoma , one with acral lentiginous, and two with melanoma without other specifications. One to three different aspirations were obtained from each patient, totaling twenty-five samples, which were submitted to RNA extraction, resulting in concentrations between 3 ng/µL to 153 ng/µL. The cell lines had the pellet isolated in the third passage of the cell culture, which subsequently had the RNA extracted and quantified which ranged from 61 ng/µL to 448.7 ng/µL.

Conclusões

The partial results of the pilot project show sufficient RNA concentrations for gene expression analysis by ddPCR. Molecular analysis of SL by FNAB is feasible, accurate and may help detect metastases in melanoma with less invasion.

Palavras Chave

Sentinel lymph node; FNAB; ddPCR