The Microwell-mesh: A high-throughput 3D prostate cancer spheroid and drug-testing platform

E. O. Mosaad,J. A. Clements,M. R. Doran,K. Futrega,K. F. Chambers

doi:10.1038/s41598-017-18050-1

Abstract

Treatment following early diagnosis of Prostate cancer (PCa) is increasingly successful, whilst the treatment of advanced and metastatic PCa remains challenging. A major limitation in the development of new therapies is the prediction of drug efficacy using in vitro models. Classic in vitro 2-dimensional (2D) cell monolayer cultures are hypersensitive to anti-cancer drugs. As a result, there has been a surge in the development of platforms that enable three dimensional (3D) cultures thought to better replicate natural physiology and better predict drug efficacy. A deficiency associated with most 3D culture systems is that their complexity reduces the number of replicates and combination therapies that can be feasibly evaluated. Herein, we describe the use of a microwell platform that utilises a nylon mesh to retain 3D micro-tumours in discrete microwells; termed the Microwell-mesh. The Microwell-mesh enables the manufacture of ~150 micro-tumours per well in a 48-well plate, and response to anti-tumour drugs can be readily quantified. Our results demonstrate that 3D micro-tumours, unlike 2D monolayers, are not hypersensitive to Docetaxel or Abiraterone Acetate, providing a superior platform for the evaluation of sequential drug treatment. In summary, the Microwell-mesh provides an efficient 3D micro-tumour platform for single and sequential drug screening.

Highlights

Common 2-dimensional (2D) monolayer cell culture generally fails to mimic the complex behaviour of native tissue. 2D culture limits cell-cell interaction, modifies gene expression and limits tissue-like matrix accumulation[1]
The use of 2D culture in cancer drug screening remains common, as standard 2D tissue culture plates are inexpensive, high throughput fluidics systems are compatible with such plates, and a range of platforms are available to facilitate imaging or characterisation of cultures maintained in standard 2D tissue culture plates
Transition to the widespread use of 3D cultures in cancer research is dependent on the development of efficient culture systems that enable similar high throughput capacities

Summary

Introduction

Common 2-dimensional (2D) monolayer cell culture generally fails to mimic the complex behaviour of native tissue. 2D culture limits cell-cell interaction, modifies gene expression and limits tissue-like matrix accumulation[1]. A major limitation of microwell platforms, such as the one used in our previous study, is that the addition of drug(s) or the exchange of medium can displace micro-tumours from their discrete microwells. Displaced micro-tumours can be lost through medium exchange or fall into adjacent microwells where they can amalgamate with other micro-tumours, resulting in a culture of heterogeneous micro-tumour sizes and numbers To overcome this limitation our group developed the “Microwell-mesh”[11]. The openings in the mesh are large enough to allow a single cell suspension to pass through and aggregate at the bottom of each microwell, while small enough to prevent aggregated micro-tumours from escaping individual microwells This feature enables the simultaneous and efficient manufacture of hundreds of uniform micro-tumours, in a format that facilitates the multiple medium exchanges required for complex and/or sequential drug treatment

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Results