CKBM as a candidate anticancer therapeutic: p53 induction, G2/M arrest and apoptosis as mechanisms of action

CKBM as a candidate anticancer therapeutic: p53 induction, G2/M arrest and apoptosis as mechanisms of action

The G2/M cell cycle checkpoint is emerging as an attractive candidate for new cancer therapies. Two currently active fields of research aurora kinases and histone deacetylases have provided strategies to regulate this checkpoint. Researchers at CK Life Sciences now report on a candidate therapeutic, CKBM, which is able to induce p53 expression, arrest cells at G2/M, induce apoptosis and also to reduce tumor size in gastric cancer xenografts.

Although considerable activity surrounds the development of novel targeted therapeutics, classic cytotoxic agents continue to underlie first line treatments in oncology. The alkaloids and antimetabolites are the most commonly used cytotoxics however alkylating agents, antineoplastic antibiotics and a class of other diverse agents such as platinum compounds are also used by oncologists. In general cytotoxics act on the cell cycle with most agents either acting at early stages of the cycle or after entry into mitosis.

The cell cycle can be broadly divided into two phases; interphase and mitotis. During the three phases of interphase G1, S and G2 the cell prepares for division and undergoes DNA replication. Towards the end of interphase the cell forms the spindle apparatus comprising two centrosomes which act as poles for an assembly of microtubules. Chromsomes attach themselves midway between the two centrosomes and during mitosis the duplicated chromosomes move separately towards opposite centrosomes along the microtubules. Mitosis can be subdivided into four phases: prophase; metaphase; anaphase; and telophase/cytokinesis. During the cell cycle three checkpoints have evolved to ensure accurate DNA replication and to prevent cellular abnormalities from appearing.

The G1 checkpoint arrests the cell cycle by inhibiting G1-S transition machinery. The replication of cells with DNA damage are stalled at the G1 checkpoint until DNA damage can be repaired; if the damage is too extensive to repair, the cell commits suicide via apoptosis. The G1 checkpoint usually arrests the cell cycle by inhibiting cyclin-dependent kinases (cdks). Tumor suppressor genes are components of G1 checkpoint, with p53 being the key protein for coordination of a variety of G1 checkpoint functions. Disruption to the G1 checkpoint is related to the development of cancer while on the other hand proteins involved in the G1 checkpoint such as p53 and the cdks have become targets for cancer therapeutics. The G2/M checkpoint prevents cells from attempting to undergo mitosis in an inappropriate state and due to defects in G1 checkpoint mechanisms, cancer cells depend on G2/M checkpoint far more than normal cells suggesting that therapeutic agents acting at this cell cycle phase may confer tumor selectivity. A third checkpoint, the mitotic checkpoint, has evolved to ensure the correct attachment of sister chromatids to the mitotic spindle and mitotic spindle tension.

The G2/M checkpoint is currently receiving growing attention as a target for cancer therapeutics and the inhibition of two molecular targets recently analyzed by LeadDiscovery, the histone deacetylases (see Histone deacetylase inhibitors-Moving from the bench to a promising companion for classic and targeted cancer therapies) and aurora kinases (see Aurora Kinase inhibitors - The dawn of a new approach to cancer) may offer an effective way of preventing G2/M transition and hence a targeted approach to cancer. Defects in the checkpoint lead to cells inappropriately entering mitosis (either with DNA damage or defective spindle assembly) leading to the appearance of mutated cells. As well as being involved in the G1 checkpoint p53 also plays a key role in the G2/M check point. The induction of p53 is able to arrest cells at this checkpoint and further more if repair is unsuccessful p53 is then able to initiate the apoptosis machinery. Thus reconstituting the check point offers one approach. In contrast, the opposite approach has also been targeted therapeutically by the Japanese company CanBas who are developing agents rationally designed to override rather that reconstitute the G2/M checkpoint, G2/M checkpoint abrogators. This may appear a risky approach however it is one based on the assumption that cells with damaged DNA to die in mitosis (or at the G1 phase of the subsequent cycle).

The field of oncology is characterized by the extensive use of naturally occurring substances. The best examples of this are to be found in the plant alkaloid class which includes vinca alkaloids such as vincristine and vinblastine, taxanes, and topoisomerase inhibitors such as Pfizer’s Camptosar and VePesid (etoposide). The Int J Med Sci study highlighted here evaluates the anticancer mechanism of action of CKBM, a product that contains the water extracts of wu wei zi (Schisandra chinensis), ginseng (Panax ginseng), hawthorn (Fructus Crataegi), jujube (Ziziphus jujube), soybean (Glycine Max) and Baker’s yeast (Saccharomyces cerevisiae) which is processed by a proprietary technology developed by Hong Kong company, CK Life Sciences Int’l Inc.

CKBM is capable of improving immune responsiveness through the induction of cytokine mediators, such as TNF-alpha and IL-6. In their present paper Sharon Luk and colleagues from CK Life Sciences demonstrate that CKBM inhibits tumor cell proliferation through the induction of apoptosis and G2/M cell cycle arrest. Both the immunostimulatory effect and the direct effect on tumor cells could contribute to an overall anticancer effect and of note the therapeutic activity of CKBM has previously been demonstrated in mice with gastric cancer xenografts (Shin et al, 2004).

In their recent study the CK Life Sciences group demonstrate that CKBM inhibited cell proliferation in a dose-dependent manner. Cells appeared to arrest in the G2/M phase with 50% of all cells being in this phase following treatment with the highest concentration of CKBM compared to 10% under control conditions. Concomitant dose dependent increases in p53 as well as p21 14-3-3σ (stratifin), two proteins under the transcriptional control of p53 were also all observed. Each of these proteins are involved in regulating the G2/M checkpoint. Cells arrested at G2/M enter apoptosis and paralleling this was a dramatic increase in apoptosis with the the proportion of cell undergoing apoptosis increasing from 2.8% to 16.5%.

This study therefore establishes CKBM as a G2/M checkpoint regulator, arresting cells in this stage of the cell cycle and inducing their apoptotic death. Therapeutic agents acting on the G2/M checkpoint are emerging as a new and potentially targeted approach to cancer and further study of CKBM is eagerly awaited. In particular evaluation of its efficacy in non-cancerous cells should indicate tumor selectivity and in addition studies in cells harboring p53 mutations should be conducted. The current study employed a cell line with functional p53 however p53 mutation is highly common in cancer and for CKBM to be a useful treatment efficacy should ideally be maintained in such cells.