Progression through the cell cycle and programmed cell death involve the complex interaction of several families of proteins in a timely coordinated fashion. Cells divide during mitosis and cytokinesis separates the mother cell into 2, while the chromosomes are segregated by the mitotic spindle to ensure propagation through future generations. The cell cycle is regulated by cyclins, cyclin-dependent kinases (CDKs), and cyclin-dependent kinase inhibitors (CDKIs). The cell cycle is divided into 4 distinct phases (G1, S, G2, and M). The progression of a cell through the cell cycle is promoted by CDKs, which are positively and negatively regulated by cyclins and CDKIs, respectively. These mechanisms play a central role in the sensitivity of malignant cells to chemotherapy. Drugs interfering with the spindle formation or targeting to damage the DNA content of the cell alter the capacity to propagate an intact genome during replication and eventually kill the cells. However, cancer cells have the capacity to arrest, repair the damage, and resume the cycle. This cell cycle-mediated resistance may be overcome by the understanding of chemotherapeutic cell cycle effects and by appropriate sequencing and scheduling of agents in combination chemotherapy. In particular, it is critical to identify and abrogate cell cycle checkpoint events to enhance the therapeutic index of our treatments.
Cell Cycle Regulation
For the cell to move from one stage of its life cycle to the next, certain proteins must be activated while others must be inhibited. The induction of apoptosis (programmed cell death) and the cell cycle are intimately related. The molecular cascade of apoptosis is characterized by the early release of mitochondrial cytochrome c, activation of Apaf-1, activation of caspase 9, and subsequent cleavage of downstream caspases in a self-amplifying cascade. The biochemical cascade of apoptosis is subject to regulation at several levels. Members of the Bcl-2 family of proteins may be either antiapoptotic (Bcl-2, Bcl-xL, and Mcl-1) or proapoptotic (BAD, Bax, Bak, and others). Several drugs promote the loss of the anti-death protein and the activation of the pro-death protein, altering this delicate balance. Identification and characterization of the genes responsible for protecting the cell from death is also important to develop targeted therapies and turn an arrested cell into a dead cell.
Our laboratory program depends on the identification of cell cycle-regulated targets either through the molecular analysis of proliferative and apoptotic pathways or through the study of gene microarrays that compare normal with malignant tissue. As many of the drugs we study have cell cycle effects, we explore the machinery of cell cycle regulation and apoptosis with target validation in the laboratory that also can be carried into clinic. Once these targets are identified, we then begin the process of drug targeting with testing both in vitro and in vivo novel, small molecules that inhibit these critical pathways. For the agents showing the most promise, we will then translate these laboratory observations into clinical trials. The clinical trials are designed to determine the safety of new drug combinations; to examine pharmacokinetic interactions between drugs, to examine tumor tissue for target validation; and to obtain preliminary data on activity for eventual Phase II clinical trials in STS.
Drugs used for chemotherapy today as well as a novel class of cell cycle-specific modulators are currently in investigational use in the laboratory and in early clinical trials. These targeted agents include flavopiridol, bryostatin-1, UCN-01, SAHA, the Nutlins, inhibitors of aurora kinase B, inhibitors of Chk1, and inhibitors of MAPK. With each of these new drugs, we have observed effects that clearly link them to the cell cycle. Understanding the physiology of cell division and cell death allows us to develop a mechanistic approach to drug development with rapid translation of preclinical discoveries into clinical cancer therapy. Some of our current laboratory projects include:
- Examination of the mechanisms of flavopiridol induced apoptosis;
- Determining the functions of p53: cell cycle arrest versus cell death;
- Targeting aurora kinase B: its relationship to polyploidy and the centrosome;
- HDAC inhibitors and their effects on the mitotic spindle;
- Drug-induced autophagy as an alternate mechanism of cell death
- Targeting chk1 to enhance chemotherapy-induced apoptosis: a mechanistic approach;
- The Nutlins: a new class of targeted drugs for cancer therapy;
- Drg1 and its role in regulating apoptosis;
- Development of new targeted therapies for the treatment of soft tissue sarcomas;