The Jechlinger group uses a 3D culture system of primary mouse mammary epithelial cells to study cancer-initiating oncogenes.
Previous and current research
In patients and mouse models, interference with the activity of cancer-initiating oncogenes can result in tumour regression. However, novel therapies that target the products of mutant alleles in human cancers are only partly successful, since maintenance of remission requires long-term treatment and relapse often occurs in the presence of therapeutic agents. Hence, a better understanding of drug resistance and tumour recurrence is needed for the design of more successful anti-cancer strategies.
Transgenic mice carrying regulatable transgenes represent tractable systems for studying the mechanisms of oncogene dependence, the response and resistance to targeted drugs and tumour recurrence. In a complementary approach, we have developed a 3D culture system of primary mouse mammary epithelial cells to study detailed responses to the induction and de-induction of oncogenes (mimicking treatment with an ideally targeted drug). This 3D system produced phenotypic changes similar to those observed in the mammary glands of the transgenic mice from which the cultures were derived. In addition, this new approach identified and isolated cells that had survived oncogene withdrawal, which represent a pure population of residual ‘dormant’ tumour cells. We recently verified that these residual cells progress to form recurrent tumours similar to relapses observed in the animals and reminiscent of the patient situation. Molecular profiling revealed unique hallmarks of surviving residual cells, which we are currently analysing to better understand mechanisms of breast tumour recurrence.
(A) One cell in anaphase divides with the sets of chromosomes perpendicular to the apical surface, while keeping ZO1 localised to the luminal membrane.
(B) Confocal microscopy (5mm projection through the middle) shows epithelial cell polarity. E-Cadherin, (adherens junctions, lateral); ZO1, (tight junctions, apical); Integrin a6 (basolateral).
(C) Doxycycline exposure causes loss of epithelial polarity and filling of the lumen; Removal of doxycycline results in survival of a re-polarised cell layer, that acquires the ability to exclude Hoechst 33342. Left panels: Confocal microscopy (5mm projection through the middle) shows Dapi, GM130 (apical), ZO1 (tight junction), Integrina6 (basal) at indicated times. Middle panels: Bright-field pictures show: (top) small, hollow acini; (middle) filled, irregular shaped spheres (bottom) hollow, irregular shaped spheres that show debris of internal cells. Right panels: Exclusion of Hoechst 33342 (1 hour incubation) at indicated times.
Future projects and goals
We are employing genomic methods, a range of molecular biology techniques, histopathological analysis as well as live cell imaging to investigate our organotypic 3D cell culture systems and the corresponding mouse models. The main aims include:
- Determine at which point during tumourigenesis cells acquire the ability to escape targeted therapy.
- Identify the molecular properties that distinguish surviving-residual cells, from naive cells.
- Interfere with the mechanisms important for survival of residual ‘dormant’ cells.