Analysis of Mitosis and Antimitotic Drug Responses in Tumors by In Vivo Microscopy and Single-Cell Pharmacodynamics

Abstract

Cancer relies upon frequent or abnormal cell division but how the tumor microenvironment affects mitotic processes in vivo remains unclear, largely due to the technical challenges of optical access, spatial resolution, and motion. We developed high-resolution in vivo microscopy methods to visualize mitosis in a murine xenograft model of human cancer. Using these methods, we determined whether the single-cell response to the anti-mitotic drug paclitaxel was the same in tumors as in cell culture; observed the impact of paclitaxel (Ptx) on the tumor response as a whole; and evaluated the single-cell pharmacodynamics of paclitaxel (by in vivo pharmacodynamic microscopy [IPDM]). Mitotic initiation was generally less frequent in tumors than in cell culture, but subsequently it proceeded normally. Paclitaxel treatment caused spindle assembly defects and mitotic arrest, followed by slippage from mitotic arrest, multinucleation and apoptosis. Compared to cell culture, the peak mitotic index in tumors exposed to paclitaxel was lower and the tumor cells survived longer after mitotic arrest, becoming multinucleated rather than dying directly from mitotic arrest. Thus, the tumor microenvironment was much less pro-apoptotic than cell culture. The morphologies associated with mitotic arrest were dose- and time-dependent, thereby providing a semi-quantitative, single-cell measure of pharmacodynamics. Although many tumor cells did not progress through Ptx-induced mitotic arrest, tumor significantly regressed in the model. Our findings demonstrate that in vivo microscopy offers a useful tool to visualize mitosis during tumor progression, drug responses, and cell fate at the single cell level.

Label-free DNA imaging in vivo with stimulated Raman scattering microscopy

“…(ii ) For in vivo cell division dynamic imaging experiments, Ncr nude female mice (Taconic) were used to establish a xenograft model (45,46). Synchronized HeLa cells with double thymidine block were injected into the chamber superficially beneath the inner side of the skin.…”

Targeting mitotic exit with hyperthermia or APC/C inhibition to increase paclitaxel efficacy

“…In the future, in vivo studies are necessary to support this concept, especially in light of a recent report that showed decidedly lower mitotic index in response to PTX administration in xenograft settings compared with that achieved in cell culture. 52 Although lower mitotic rates are observed in human specimens as well, 53 the analysis of fine-needle aspiration biopsies of breast cancer patients demonstrated that the tumor mitotic index was increased up to 10-fold after PTX treatment and lasted up to 2-3 days after PTX administration. 54 These results suggest that the combinatorial treatment regimen that we propose is feasible.…”

Implantable microenvironments to attract hematopoietic stem/cancer cells

“…Dorsal skinfold chambers have been used extensively for intravital imaging of tumors and other tissues (43)(44)(45); in our study we used this system for in situ observation of hematopoietic cell trafficking in an implanted scaffold. One advantage of a skinfold window chamber is the opportunity to image the microenvironment noninvasively over time scales that are relevant to cell homing, localization, and interactions of hematopoietic stem and cancer cells; currently, the use of other methods for such imaging is limited (46).…”