Cancer Biology

Heterogeneous prostatic carcinoma-associated fibroblasts (CAF) contribute to tumor progression. This was established using transgenic mouse models. Paracrine interactions between fibroblasts and epithelial cells were further interrogated using isolated 2D cell culture systems, but 3D culture systems currently being developed can better mimic reciprocal interactions potentially found in the native tissue. To understand paracrine and juxtacrine signaling among fibroblasts and epithelia, 3D co-cultures with species differences allows for further subsequent analysis of the cultures. The use of mouse and human cells, for example, in one system allows for species-specific FACS or quantitative PCR analysis. This protocol describes the use of a 3D Co-culture System of Mouse Prostatic Wild-type Fibroblasts with Human Prostate Cancer Epithelial Cells.

In 1999, Hahnfeldt et al. proposed a mathematical model for tumor growth as dictated by reciprocal communications between tumor and its associated vasculature, introducing the idea that a tumor is supported by a dynamic, rather than a static, carrying capacity. In this original paper, the carrying capacity was equated with the variable tumor vascular support resulting from the net effect of tumor-derived angiogenesis stimulators and inhibitors. This dynamic carrying capacity model was further abstracted and developed in our recent publication to depict the more general situation where there is an interaction between the tumor and its supportive host tissue; in that case, as a function of host aging (Benzekry et al., 2014). This allowed us to predict a range of host changes that may be occurring with age that impact tumor dynamics. More generally, the basic formalism described here can be (and has been), extended to the therapeutic context using additional optimization criteria (Hahnfeldt et al., 1999). The model depends on three parameters: One for the tumor cell proliferation kinetics, one for the stimulation of the stromal support, and one for its inhibition, as well as two initial conditions. We describe here the numerical method to estimate these parameters from longitudinal tumor volume measurements.

A tumorsphere is a solid, spherical formation developed from the proliferation of one cancer stem/progenitor cell. These tumorspheres (Figure 1a) are easily distinguishable from single or aggregated cells (Figure 1b) as the cells appear to become fused together and individual cells cannot be identified. Cells are grown in serum-free, non-adherent conditions in order to enrich the cancer stem/progenitor cell population as only cancer stem/progenitor cells can survive and proliferate in this environment. This assay can be used to estimate the percentage of cancer stem/progenitor cells present in a population of tumor cells. The size, which can vary from less than 50 micrometers to 250 micrometers, and number of tumorspheres formed can be used to characterize the cancer stem/progenitor cell population within a population of in vitro cultured cancer cells and within in vivo tumors (Lo et al., 2012; Liu et al., 2009). While several cell lines can be used for tumorsphere formation assay (e.g. primary mammary tumor cells from Her2/neu-transgenic mice, MCF7, BT474 and HCC1954), some cell lines may not form typical tumorsphere structures and may be difficult to count or classify definitively as tumorspheres.