A Simple Three-Dimensional In Vitro Culture Mimicking the In Vivo-Like Cell Behavior of Bladder Patient-Derived Xenograft Models.

Hongyong Zhang,Ai-Hong Ma,Robson Amaral,Maike Zimmermann,Kamilla Swiech,Chong-Xian Pan

doi:10.3390/cancers12051304

Abstract

Patient-derived xenograft (PDX) models allow for personalized drug selection and the identification of drug resistance mechanisms in cancer cells. However, PDX models present technical disadvantages, such as long engraftment time, low success rate, and high maintenance cost. On the other hand, tumor spheroids are emerging as an in vitro alternative model that can maintain the phenotype of cancer cells long enough to perform all assays and predict a patient’s outcome. The present work aimed to describe a simple, reproducible, and low-cost 3D in vitro culture method to generate bladder tumor spheroids using human cells from PDX mice. Cancer cells from PDX BL0293 and BL0808 models, previously established from advanced bladder cancer, were cultured in 96-well round-bottom ultra-low attachment (ULA) plates with 5% Matrigel and generated regular and round-shaped spheroids (roundness > 0.8) with a diameter larger than 400 μm and a hypoxic core (a feature related to drug resistance in solid tumors). The responses of the tumor spheroids to the antineoplastic drugs cisplatin, gemcitabine, and their combination were similar to tumor responses in in vivo studies with PDX BL0293 and BL0808 mice. Therefore, the in vitro 3D model using PDX tumor spheroids appears as a valuable tool that may predict the outcome of in vivo drug-screening assays and represents a low-cost strategy for such purpose.

Highlights

Bladder cancer is considered a serious public health problem due to its high recurrence rate and mortality, with approximately 150,000 deaths in the world per year [1,2]; the development of new therapeutic approaches is of paramount importance
The culture of single cells isolated from tumor fragments presented a predominance of tumor cells with respect to fibroblasts, as seen in slides stained with a hematoxylin and eosin Y (H&E) solution (Figure 1A,B)
96-well ultra-low attachment (ULA) plates and RPMI culture medium supplemented with 5% Matrigel, Patient-derived xenografts (PDX) tumor spheroids BL0293 and BL0808 were formed within 48 h at all cell seeding densities used (Figure 1C,D)

Summary

Introduction

Bladder cancer is considered a serious public health problem due to its high recurrence rate and mortality, with approximately 150,000 deaths in the world per year [1,2]; the development of new therapeutic approaches is of paramount importance. The current approach of ‘one treatment fits all’ is not considered the best strategy since it does not take into account the patient-specific genetic variability that has a potential influence on the response to drugs [3]. One of the approaches of personalized oncology is based on the prediction of the response of a patient’s tumor to anti-neoplastic drugs to help the oncologist plan the best treatment for that patient [4]. We previously showed that PDXs retained 92%–97% of the genetic alterations present in the parental patient cancers [7]. This model facilitates personalized drug selection and the identification of tumor resistance mechanisms [8]. PDX-based models present some technical disadvantages such as low success rate (30%–40%), long engraftment time (2 to 8 months), high cost, limited statistical power, and low potential for application to high-throughput studies [9]

Methods

Results

Conclusion