Actin Inhibition: The identification of PTX-2 as a novel anti-mitotic agent that selectively induces apoptosis in p53 deficient cancer cells
SummaryAlthough cytotoxics agents represent a cornerstone of oncology, toxicity and drug resistance has driven the development of targeted anticancer therapies. The current study highlights a novel antimitotic agent, PTX-2. This molecule, like the Taxol and Taxotere, blocks the cell cycle in late mitosis channelling cells into an apoptotic program. In direct contrast to Taxol and Taxotere, PTX-2 targets cells with p53 mutations; the plant alkaloids on the other hand are relatively insensitive in such c
Cytotoxic agents represent a cornerstone of oncology however many established agents are reaching patent expiry and the last cytotoxic genericization is expected in 2011 (Xeloda). One good example of the effects of genericization is the plant alkaloid Taxol (paclitaxel). Taxol, one of a group of alkaloid mitotic inhibitors, once dominated the cytotoxic market but because of its patent expiry it has lost its leading position to another agent from this class, Aventis’ Taxotere (docetaxel) which in 2004 generated worldwide sales of $1.8 billion.
The effectiveness of the mitosis inhibitors like most cytotoxic agents has been impaired by various side effects, notably neurological and hematological toxicities. Mutations also limit the utility of many therapeutics. In particular p53 mutations which occur in the majority of cancer cells introduces resistance to cytotoxic agents. p53 plays an important role in the decision to either arrest cell cycle progression or induce apoptosis in response to a variety of stimuli. At a molecular level, p53 activates the transcription of several downstream target genes, including p21, an inhibitor of cyclin-dependent kinases which causes arrest in the G1 phase of the cell cycle. In addition p53 target genes activate cell cycle checkpoints and induce apoptosis. On the other hand, defects in p53 disrupt cell cycle check points including the G1 and the G2/M checkpoints as well as the spindle checkpoint.
The vinca alkaloids target the cytoskeleton by binding to tubulin. This therapeutic class interferes with microtubule formation and spindle dynamics thereby preventing cells passing the spindle checkpoint and as a result cells instead enter into apoptotic programmed cell death. The ability of p53 mutations to disrupt this checkpoint allows cells to become resistant to agents such as Taxol. Thus strategies that reintroduce p53 function represent attractive approaches to cancer. In our recent proof of concept evaluation of the aurora kinases (see Aurora Kinase inhibitors - The dawn of a new approach to cancer) we discuss how inhibition of aurora kinase-A can stabilizes p53 causing cell cycle arrest at the G2/M checkpoint and increased sensitivity to cytotoxic agents. Alternatively the development of mitosis inhibitors that are able to activate p53-independent apoptosis is also attractive.
In their recent paper published in the journal Oncogene, Korean researchers report the development of a novel cell-based screening system for identifying mitosis inhibitors. This assay which evaluates mitosis in ovulated oocytes is of importance in its own right and could be of benefit to drug discovery programs irrespective of the molecular target under investigations. The Korean group led by Dr Deug Shin employed this assay to screen 1100 natural compounds for antimitotic activity. Of this library 12 compounds inhibited mitosis preventing cells from completing cytokinesis (the final stage of mitosis when genomic material is split between two daughter cells). The cytotoxic effect of these compounds was then determined in cancer cells and one compound, pectenotoxin-2 (PTX-2) had the particularly exciting property of being more sensitive in cells that do not express p53 as compared to those that do. In contrast to the vinca alkyloids PTX-2 has previously been reported to be an actin depolymerising agent. In vivo, PTX-2 reduced the size of established p53-/- tumors to almost half that of controls while tumors with wild-type p53 were unaffected.
Apoptosis, or programmed cell death, consists of a series of characteristic physiological changes that culminate in the phagocytic removal of cells. This process can be activated via either the extrinsic or the intrinsic pathway. The latter is triggered by cellular stress and chemotherapeutic drugs which cause the outer mitochondrial membrane to become permeablized, allowing intermembrane proteins such as cytochrome c and Apaf-1 to be released. The end result is the activation of caspase-9 and down-stream effector caspases including caspase-3 which eventually culminates in cell death. Mitochondrial membrane permeablization is regulated by the BCL2 family of proteins and the mitochondrial permeability transition pore.
The development of apoptosis stimulators is led by Genta’s Genasense, an antisense therapeutic that inhibits the production of Bcl-2 in cancer cells. Like Genasense, PTX-2 was also proapoptotic apparently stimulating the intrinsic pathway as p53-/- cells displayed an increase in cytochrome c release following treatment. As expected both caspase-9 and -3 activity was also increased through a mechanism apparently dependant on the sequential activation of the apoptosis regulators Bim and Bax, the latter being responsible for the mitochondrial release of proapoptotic factors.
Worth an estimated $35 billion in 2003, analysts predict that the sale of oncology therapeutics will grow to $60 billion by 2010. The development of targeted cancer therapies are current receiving considerable interest. This field includes angiogenesis inhibitors and; signal transduction inhibitors apoptosis stimulators (see Innovative Cancer Therapies: Targeted therapy, a clinical and commercial revolution). First generation apoptosis stimulators include Genta's Genasense; more recently we have highlighted a molecule known as pancratistatin which appears to target mitochondria in cancer cells (see our editorial). In the present study we highlight a further promising apoptosis stimulator, PTX-2. This exciting molecule appears to possess potent cytotoxic activity in p53-/- cancer cells which accounts for over half of all cancers while leaving cells with wild-type p53 unharmed suggesting that the therapeutic index of PTX-2 will be highly attractive. PTX-2 seems to activate the apoptotic program as a result of its ability to block the cell cycle during late phases of mitosis although the exact mechanism requires further investigation. The classic antimitotic agents such as Taxol and Texture also induce apoptosis by interfering with mitosis however in stark contrast to these agents that are less sensitive in cells with p53 mutations, PTX-2 is more active in such cells. This is not only likely to confer anticancer activity in typically resistant cells; it is also likely to extend a high therapeutic margin by leaving healthy cells unharmed. PTX-2 also differs from Genesense which induces apoptosis earlier on in the cell cycle suggesting that the two agents could be used together to facilitate the killing of cells which, for whatever reason progress into mitosis even in the presence of Genesense