• The BCRL at Fox Chase Cancer Center from 1991 to the present
• Seminal work performed in the BCRL from 1972 to 1991
• Accomplishments
• Translational Research at the BCRL
• Recent Accomplishments
• Conferences and Lectures

The BCRL at the Fox Chase Cancer Center from 1991 to the present

Transformation of HBEC with chemical carcinogens

These studies have been performed on MCF-10F cells using two carcinogenic compounds that require metabolic activation, DMBA and B[a]P, as well as two direct-acting carcinogens, NMU and MNNG. (Figure 10: Click to see).

Figure 10: Flow chart showing the evolution of MCF-10F cells after treatment with the carcinogens NMU, BP, DMBA, or MNNG. Days in culture indicate the number of days in vitro that it took each cell line to emerge. (From: Calaf, G., and Russo, J. Transformation of human breast epithelial cells by chemical carcinogens. Carcinogenesis 14:483-492, 1993).

The acquisition of anchorage independence was manifested at 157 days in culture, and the efficiency varied from carcinogen to carcinogen, being the highest for DMBA and BP. NMU was the least efficient in inducing colony formation and did not originate clones; MNNG originated four colonies from which only one clone, M4, emerged at 248 days post-treatment. M4 exhibited greater colony efficiency (CE) than the parental cells, and generated other subclones that emerged by 446 days in culture. We observed that the apparent refractory nature of human breast epithelial cells to express the phenotypes of neoplastic transformation upon treatment with carcinogens in vitro could be overcome by isolating and reseeding the colonies formed in agar - methocel, which are further selected by reseeding in semisolid medium. Whereas this selective pressure in vitro may not represent the same forces that are operational in vivo, our observations indicate that the emergence of neoplastic phenotypes are a continuum expressed only in those cells that exhibit further growth advantage. The clones derived from DMBA and B[a]P-treated cells, (i.e. D3-1 and BPI-E respectively), exhibited greater invasive and chemotactic capabilities than MCF-10F control cells, and both parameters were similar to those of the tumorigenic T24 cells. The clones BP5, BP7 and BP10 presented greater values for invasion and chemotaxis than MCF- 10F control cells, but less than BPI-E, D3-1 and T24 cells. The higher chemoinvasive and chemotactic capacity of BP1-E and D3-1 cells was observed in late passages and correlated with their higher CE in agar - methocel. BP1-E, cell line expressing tumorigenesis in SCID (Figure 11: Click to see).

Figure 11: BP treated MCF-10F cells (phase contrast) form colonies that isolated and expanded were reseeded in agar methocel originating the cell lines, BP1,BP2, BP5, BP6, BP7 and BP10. From BP1 and BP2 originates the clones BP1-E and BP2B. BP1-E is the clone that will induce tumors in heterologous host. (From: Calaf, G., and Russo, J. Transformation of human breast epithelial cells by chemical carcinogens. Carcinogenesis 14:483-492, 1993).

As indicated above, the levels and localization of ras expression in normal and malignant breast tissues have been examined and quantitated by analyzing breast tissue samples for expression of ras related mRNA and p21 ras protein which has been found to be expressed in biopsies of both normal and malignant breast tissues. However, whether the ras oncogene is a causative agent of human breast cancer has not been proved as yet.

Therefore, one way to evaluate the contribution of ras genes in the development of the tumorigenic phenotype is to introduce this gene into suitable acceptor cells. Transfection of the non tumorigenic cell lines, clones D3-1 and BP1, derived from the carcinogen-treated MCF-10F cell lines, and the MCF-10F cell line with the c-Ha-ras oncogene not only enhanced colony formation in agar-methocel as well as invasiveness, but induced tumorigenicity with a short latency period in SCID mice (Figure 12: Click to see and Figure 13: Click to see).

Figure 12: Flow chart showing the efficiency of c-has-ras in increasing the tumorigenic phenotype of BP1-Ecells.

Figure 13: (a) Nude mice inoculated with MCF10F cells. (b) Nude mice inoculated with BP1-T-ras cells. (From: Russo, J. and Russo, I.H. “Biological and Molecular Basis of Breast Cancer”, Springer –Verlag, Heidelberg, Germany, 2004. pp271)