Most biologists rely on cell culture in the two-dimensional (2D) format for studying tumour context, but cells growing in monolayers on plastic surfaces do not accurately reflect the in vivo state. 3D cell culture techniques more closely mimic tissues and organs as they produce extracellular proteins in their native configuration, allowing biological instructions to take place. The benefits of 3D cell culture are especially useful when mimicking pathological conditions such as cancer. For example, cells grown in 3D will form natural barriers to drugs such as tight junctions that bind cells tightly together and block or slow the diffusion of drugs. Other currently used 3D models involve growing cells with the support of a scaffold, which may manipulate the tumour microenvironment and hence are not a satisfactory representation of the tumour. Our study has addressed this problem by growing cells in 3D without the support of scaffold, thus creating an environment that more closely mimics the realistic tumour state. Our model produces true 3D tumour spheroids rather than discoid structures, as confirmed with both light and scanning electronic microscopy (SEM). The spheroids represent an improved mini-tumour model as indicated by the visualization of cell junctions under transmitted electronic microscopy (TEM), and the presence of a hypoxic gradient as evidenced by HIF1a staining. Cells grown as 3D spheroids were highly drug resistant compared to cells grown in 2D culture. Gene expression levels were confirmed using microRNA microarray and digital droplet PCR and found to be similar to those in MPM tumour samples, including overexpression of miR-21 and miR-210. We then showed that suppression of miR-21, miR-210 or HIF1a expression was able to sensitize spheroids to chemotherapeutic drugs. Our 3D model also showed up-regulation of genes that contribute to drug resistance such as Hif1a, YAP1, ABCG2 and YB1. In summary, our model provides a simple and cost-effective method for analyzing spheroid biology and gene expression and their influence on cancer drug resistance.