Multicellular Spheroids Compared to Conventional Cell Culture in the Evaluation of Synergistic Hyperthermia and Chemotherapy in the Treatment of Cancer

Cancer is the second leading cause of death in the United States with an estimate of 580,000 deaths in 2013[1]. Of the previous estimate, approximately 163,890 of those deaths are caused by lung cancer alone [1].  Along with lung cancer, breast cancer is one of the deadliest forms of cancer in women. The current treatment of any cancer can be filed under three categories including surgery, chemotherapy, and radiation. Because these diseases have such high mortality rates, there is a significant need for high throughput methods that can be easily used as assays to screen potential new cancer treatments. Currently, most potential new treatments are screened by exposing them to cancer cells grown as a 2-dimensional (2D) monolayer. This conventional method of cell culture is extremely accessible and inexpensive,  and it provides knowledge about how the cells will respond to certain treatments. However, it has been shown that cells grown as a 2D monolayer are physiologically different than the 3-dimensional (3D) tumors that grow inside the human body [2]. Because of this, it is possible that the tumors inside the human body will respond to anti-cancer treatment differently than those cancer cells grown as a 2D model [2-3,5]. In order to minimize this possible difference, a high throughput 3D model was introduced in order to better mimic the activity of drugs on tumors in vivo.

It has been previously shown that the efficacy of chemotherapeutic drugs can be increased when combined with hyperthermia [4], the controlled heating of tissue from 42-45°C [6]. It has been shown through previous in vivo studies, that hyperthermia causes changes in physiological aspects in tumor such as blood perfusion, partial pressure of oxygen, vascular permeability, and intra-tumor pressure. Each of these changes causes the tumor to be more receptive to the chemotherapeutic.

The overall objective of this study was to evaluate the response of lung cancer cells, A549, and breast cancer cells, MDA-MB-231, cultured in both 2D and 3D conditions to hyperthermia and chemotherapy. Two groups of cells were exposed to either cisplatin or paclitaxel. One of these groups was placed in an incubator at 37°C with 5%  to serve as the chemotherapeutic only control. The other group was introduced to hyperthermia by being placed on a hot plate in an incubator set at 43°C with 5% , serving as the chemotherapeutic with hyperthermia group. The temperature of the cells in this group was determined using a temperature probe. Once the cells reached 42^oC, they were exposed to both the hyperthermia conditions and the chemotherapeutic agent for either 30 minutes or 2 hours. After the treatment, the cells were placed in fresh media and both groups were incubated at 37 °C and 5%   for 72 hours after which a resazurin assay was used to determined cell viability.   The efficacy of the combined treatments was measured by comparing IC50 values, the concentration of treatment that reduced cell viability by 50%.  Studies thus far on 2D A549 cells showed that when combining cisplatin with hyperthermia, the IC50 value decreased compared to cisplatin alone, whereas when combining paclitaxel with hyperthermia, the IC50 value increased. Future studies will focus on 2D MDA MB 231 cells and both cell lines as 3D models.             

Acknowledgements:

Research supported by the National Science Foundation REU Program on Bioactive Interfaces and Devices Grant #EEC-0851716.

University of Kentucky Bucks for Brains Program

References:

[1] “Lung Cancer Statistics.” Division of Cancer Prevention and Control, National Center for Chronic Disease Prevention and Health Promotion. 2010 <http://www.cdc.gov/cancer/lung/statistics/>.

[2] Lin R-Z, Chang H-Y. "Recent advances in three-dimensional multicellular spheroid culture for biomedical research." Biotechnology Journal 2008, 3: 1172-1184.

[3] Nirmalanandhan VS, Duren A, et al. "Activity of Anticancer Agents in a Three-Dimensional Cell Culture Model." Assay and Drug Development Technologies 2010, 8.5: 581-590.

[4] Issels, R. D. (2008). "Hyperthermia adds to chemotherapy." European Journal of Cancer 44(17): 2546-2554.

[5] Smalley KSM, Lioni M, and Lerlyn M. “Life Isn’t Flat: Taking cancer biology to the next dimension”. In Vitro Cellular & Developmental Biology-Animal 2006, 42:242-47.