Transformed BPECs grown in pWIT medium form squamous tumors. Human mammary epithelial cells were infected with vectors expressing TERT, BMI1, MYC and ESR1 (3G-ER cells). A. Western blot showing expression of the transgenes in parental (NT) and transformed cells. B&C. Tumor formation two weeks after subcutaneous injection of 3G-ER cells into NSG mice (n = 2). B. Stereomicrograph showing tdTomato fluorescence from the tumor. C. Glandular regions express keratin 8/18; squamous regions express keratin 14. All of the cells are ERα + because one of the lentiviruses used to transform the cells expresses ESR1. D&E. At 3 weeks (D, n = 2) and 5 weeks (E, n = 5) after injection, the tumors are dominated by stratified cells undergoing terminal squamous differentiation leading to the formation of keratin pearls. Keratin pearls are marked by asterisks. Scale bars: B 1 mm; upper panels in C 500 μm; lower panels in C 200 μm; D&E 200 μm.