Three dimensional cultivation increases chemo- and radioresistance of colorectal cancer cell lines.

Jana Koch,Christian Gromoll,Tobias Leibold,Hans J Schlitt,Marc-H Dahlke,Philipp Renner,Dina Mönch,Annika Maaß,Thomas Hehr,Arun Rishi

doi:10.1371/journal.pone.0244513

Abstract

Although 2D cell cultures are commonly used to predict therapy response, it has become clear that 3D cultures may better mimic the in vivo situation and offer the possibility of tailoring translational clinical approaches. Here, we compared the response of 2D and 3D colorectal cancer (CRC) cell lines to irradiation and chemotherapy. Classic 2D cultures and 3D spheroids of CRC cell lines (CaCo2, Colo205, HCT116, SW480) were thoroughly established, then irradiated with doses of 1, 4, or 10 Gy, using a clinical-grade linear accelerator. The response was assessed by immunohistochemistry, flow cytometry, and TUNEL assays. Upon irradiation, CRC 3D spheroids were morphologically altered. After irradiation with 10 Gy, annexin V/PI staining revealed a 1.8- to 4-fold increase in the apoptosis rate in the 2D cell cultures (95% CI 3.24±0.96), and a 1.5- to 2.4-fold increase in the 3D spheroids (95% CI 1.56±0.41). Irradiation with 1 Gy caused 3- and 4-fold increases in TUNEL positive cells in the CaCo2 and HCT116 (p = 0.01) 2D cultures, respectively, compared with a 2-fold increase in the 3D spheroids. Furthermore, the 2D and 3D cultures responded differently to chemotherapy; the 3D cultures were more resistant to 5-FU and cisplatin, but not to doxorubicin and mitomycin C, than the 2D cultures. Taken together, CRC cells cultured as 3D spheroids displayed markedly higher resistance to irradiation therapy and selected chemotherapeutic drugs than 2D cultures. This in vitro difference must be considered in future approaches for determining the ideal in vitro systems that mimic human disease.

Highlights

Colorectal cancer is the third most common cancer in the world, with about 30% of the estimated new cases arising in the rectum [1]
We compared the response of 2D colorectal cancer (CRC) cell lines and 3D CRC cell-line-derived spheroids to irradiation and chemotherapy
2D and 3D cultures were treated with standard chemotherapeutic drugs and the response to treatment was analysed by a RealTime-GloTM MT Cell Viability Assay

Summary

Introduction

Colorectal cancer is the third most common cancer in the world, with about 30% of the estimated new cases arising in the rectum [1]. The anatomical localization in the pelvis, as well as its particular blood and lymphatic supply, characterises rectal cancer as a distinct entity with regard to its invasive growth patterns, surgical approach and treatment outcomes [2]. According to current international guidelines, patients with advanced rectal cancer receive either long-course neoadjuvant radiochemotherapy or short-course radiation therapy before complete tumour removal [3]. Neoadjuvant radiochemotherapy or neoadjuvant radiotherapy before surgery is associated with a reduced incidence of local recurrence compared with surgery alone [4,5,6]. The effect of neoadjuvant therapy on overall survival has been thoroughly debated over the years, but its effectiveness has never been proven. In addition to the improvement of pretreatment regimens, there has been a major breakthrough in surgical treatment with the introduction of total mesorectal excision (TME) [7]

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