Establishment of a Primary Culture of Patient-derived Soft Tissue Sarcoma

Summary

The following protocol focuses on the establishment of a primary culture of patient-derived soft tissue sarcoma (STS). This model could help us to better understand the molecular background and pharmacological profile of these rare malignancies and could represent a starting point for further research aimed at improving STS management.

Abstract

Soft tissue sarcomas (STS) represent a spectrum of heterogeneous malignancies with a difficult diagnosis, classification, and management. To date, more than 50 histological subtypes of these rare solid tumors have been identified. Thus, due to their extraordinary diversity and low incidence, our understanding of the biology of these tumors is still limited. Patient-derived cultures represent the ideal platform to study STS pathophysiology and pharmacology. We thus developed a human preclinical model of STS starting from tumor specimens harvested from patients undergoing surgical resection. Patient-derived STS cell cultures were obtained from the surgical specimens by collagenase digestion and isolated by filtration. Cells were counted, seeded, and left for 14 days in standard monolayer cultures and then processed by downstream analysis. Before performing molecular or pharmaceutical analyses, the establishment of STS primary cultures was confirmed through the evaluation of cytomorphologic features and, when available, immunohistochemical markers. This method represents a useful tool 1) to study the natural history of these poorly explored malignancies and 2) to test the effects of different drugs in an effort to learn more about their mechanisms of action.

Introduction

Soft tissue sarcomas (STS) include a spectrum of heterogeneous lesions of mesenchymal origin accounting for 1% of all solid tumors^1,2, and the new World Health Organization classification recognizes the presence of more than 50 different subtypes³. Among these, the most common histotypes are adipocyte sarcoma or liposarcoma and leiomyosarcoma, accounting for 15% and 11% of all adult STS, respectively^4,5. Although STS can develop in different parts of the body, the extremities and retroperitoneum are the most common sites, occurr....

Protocol

STS cells are isolated from a tumor mass surgically excised from patients with STS. The protocol has been approved by the Local Ethics Committee and is performed according to Good Clinical Practice guidelines and the Declaration of Helsinki. All patients gave written informed consent to take part in the study. The surgical specimens are analyzed by an experienced sarcoma pathologist and processed within 3 hours of surgery.

1. Tumor Specimen Collection and Processing:

1. Collect tumor specimens with the help of a sarcoma pathologist in a 100-mL sterile urine container with 50 mL of DMEM low glucose medium sealed with a paraffin film....

Results

We designed a simple method to obtain the establishment of a primary culture of patient-derived soft tissue sarcoma and report here an example of results obtained on one specific STS histotype¹³. The protocol was used for the establishment of primary cultures of different STS histotypes including well-differentiated liposarcoma, dedifferentiated liposarcoma, myxoid liposarcoma, pleomorphic liposarcoma, myxofibrosarcoma, undifferentiated pleomorphic sarcoma, GIST, a.......

Discussion

Well defined preclinical models are needed to elucidate the molecular background of tumors, predict poor prognosis, and develop new therapeutic strategies for cancer patients. This is especially important for rare tumors such as STS whose high heterogeneity in terms of morphology, aggressive potential, and clinical behavior challenges our understanding of STS biology and patient management²¹. Moreover, the few commercial sarcoma cell lines available limits preclinical investigations into this grou.......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
DMEM High Glucose	EuroClone	ECB7501L
Fetal bovine serum	EuroClone	ECS0180D
Glutamine	Gibco	25030-024
Paraformaldehyde 4% aqueous solution, EM grade	Electron Microscopy Sciences	157-4-100
Penicillin streptomycin	Gibco	15140122
Triazol reagent	Ambion Life-Technologies	15596018
Trypsin	EuroClone	ECB3052D