Cell cycle modulation studies

The ability of phytochemicals to modulate cell cycle can be useful in evaluating and optimizing cancer prevention by these compounds. Studies reported here demonstrate that phytochemicals may differentially impact cell cycle and cell number depending upon the mutations in the cancer cell. In addition, cell cycle regulatory proteins are altered by phytochemicals in ways that may explain the cell cycle modulation. The fact that structural features of flavonoids impact cell cycle modulation suggests that cell cycle modulation may be useful in identifying the most active compounds for colon cancer prevention. Finally, cell cycle modulation studies can inform us on plant modifications that may be useful for nutritional enhancement. 

Flow cytometry (FCM) is widely used for exploring mechanism of action of compounds that compromise proliferation since it is rapid, accurate and usable for any cellular context [5], In this chapter we want to point out technical and strategic aspects of use of FCM for cell cycle studies of a putative anticancer agent. As an example we used Edotecarin, a topi inhibitor, firstly evaluating proliferation outcome and classical DNA content analysis by propidium iodide, and then since the compound treatment produced cell cycle perturbation difficult to interprete, a two-parametric analysis by 5-bromo-deoxyuridine (BrdU) was applied for separating cell cycle phases. Moreover we put our efforts into identifing specific cell cycle arrest not easily demonstrable by previously described methods, through the use of in vitro kinetics ( pulse and chase ). Finally, in vivo assessment of efficacy and biomarkers modulation after treatment was analyzed. 

In this chapter we show that topi inhibitors with apparently similar cytotoxic profile could have a specific effect of cell cycle, distinguishing the activity of classical camptothecin, such as irinotecan and SN-38, active specifically on the S phase by edotecarin active as tight binder on topi and without cell cycle specificity. The use of cytometry of solid tumors from xenograft made it possible to provide support to immunohistochemical analysis during the study of biomarkers, comparing cell cycle effects to topi modulation after treatment. 

Modulation of ubiquitin ligases by phosphorylation Regulation of ubiquitin ligase activity by phosphorylation has been shown in the studies on a multisubunit ligase, APC. As the name implies, this complex is critical for cell cycle progression into anaphase. A recent study shows that a form of APC is also expressed in postmitotic neurons. 

The interplay between oscillations and bistability has been addressed in detailed molecular models for the cell cycles of amphibian embryos, yeast and somatic cells [138-141]. The predictions of a detailed model for the cell cycle in yeast were successfully compared with observations of more than a hundred mutants [142]. Other theoretical studies focus on the dynamical properties of particular modules of the cell cycle machinery such as that controlhng the Gl/S transition [143]. 

---