Fig. 1.

Reversible phenotypic equilibrium in response to paclitaxel in cancer cells. (A) Regimen for expansion of paclitaxel (Ptx) stress-tolerant cells. Highly metastatic MDA-MB-231 naïve (yellow) cells were treated with Ptx (100 nM) on day 1 and day 3. After 5 d, Ptx was removed, and cells were left in a drug-free culture. Most stressed cells arrested (red) and ultimately died, whereas rare drug-tolerant cells (orange) resumed proliferation after 10–15 d, and clones were expanded. Five single cells per group were analyzed either before treatment, 1 d after Ptx removal, as well as from recently established (n < 64) or long-term expanded, drug-tolerant clones. Populations were analyzed from long-term expanded clones. Frequencies of surviving stressed and drug-tolerant cells observed are indicated between parentheses. Cell-to-cell heterogenous RNA content is indicated with varying colors. (B) Bright field images of untreated, stressed, and drug-tolerant cells at the indicated times after drug removal. Total magnification is indicated. (C) Paclitaxel toxicity assays on naïve or drug-tolerant MDA-MB-231 cells (Upper) and MCF10A cells (Lower). Growth inhibitory concentrations 50% (IC₅₀) are indicated. Data shown are the mean ± SEM from a quadruplicated representative experiment. (D) Bright-field image of an MDA-MB-231-Ptx-tolerant clone (n < 64) during single cell collection by micromanipulation. The number of days in drug-free culture is indicated at the top. The opening of the micropipette of roughly 20 μm is shown. Total magnification is indicated.