apoptosis – Drug Discovery Opinion

Dicing with Death

Unlike necrosis, apoptosis (programmed cell death) is generally beneficial to an organism and loss of apoptotic pathways can lead to uncontrolled cell proliferation. Apoptosis takes place, for example, when a cell is irreparably damaged or infected by a virus and the signal for apoptosis can come from the cell itself, from surrounding tissue, or from immune cells. Although a variety of signals and pathways can trigger apoptosis, it is ultimately the action of caspases that degrades cellular organelles and proteins. Chromosome fragmentation is a hallmark of apoptosis and, in mammals, caspases activate apoptotic chromosome fragmentation by cleaving and inactivating an apoptotic nuclease inhibitor.

Working with C. elegans, a team led by researchers at the University of Colorado at Boulder have now shown that the Dicer ribonuclease (DCR-1), instead of being inactivated by caspases, undergoes a change of function. DCR-1 normally processes small RNAs but cleavage by the CED-3 caspase produces a C-terminal fragment which shows deoxyribonuclease activity and produces 3′ hydroxyl DNA breaks on chromosomes and promotes apoptosis. Although there are many enzymes that cleave either RNA or DNA, this is the first demonstration that proteolysis of a ribonuclease can generate a deoxyribonuclease.

The researchers are now investigating whether the function of DCR-1 can be altered in the same way in human cells – if so, the authors hope that their work may lead to new ways to treat diseases caused by abnormal apoptosis such as cancer and autoimmune diseases.

The study is published in the journal Science.

Blazing a TRAIL

Tumour necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) is one of several members of the TNF gene superfamily that induce apoptosis through engagement of death receptors. The protein is a potentially attractive treatment for cancer since it induces apoptosis in a variety of cancer cells. However, not all cells of a particular type react uniformly to anti-cancer treatments and, although a promising drug candidate, TRAIL is not 100% successful. Genetic explanations have been put forward to explain the differences in cell responses to treatment but a team of scientists at Harvard University wanted to investigate the mechanisms involved in more detail. They exposed both cancer cells and normal cells to varying concentrations of TRAIL and found that a proportion of them always survived. When the surviving cells were isolated, they and their immediate progeny remained highly resistant to the apoptotic effects of TRAIL for a short time. After reproducing for several days, however, the sensitivity of the cells to TRAIL reverted to that of the original colonies with around 90% of the cells dying and 10% surviving. Using a variety of imaging techniques, the team showed that levels of proteins involved in TRAIL-induced apoptosis were different in the sensitive and resistant cells, despite the fact that the cells were genetically identical. They found that the altered protein levels were initially inherited by progeny cells but that inheritance was transient. The initial differences in protein expression between the cells were completely random – cells do not produce proteins uniformly but rather in bursts, with the timing and level of production varying from cell to cell.

The findings offer an alternative explanation to the cancer stem-cell hypothesis about why some cells are more resistant to chemotherapy or radiation treatment and the team hope that the new insight may contribute to the design of more effective treatments.

The study was published online on April 12^th in Nature.

Chemical Biology used to Probe Apoptotic Pathways

The journal Nature Chemical Biology defines chemical biology as ‘both the use of chemistry to advance a molecular understanding of biology and the harnessing of biology to advance chemistry’.

Scientists at the Karolinska Institute, Stockholm, have exploited the principles of chemical biology in the study of apoptotic pathways. In a recent publication, the scientists identified a set of 40 chemical agents (‘bioprobes’) that induce apoptosis from screening of a chemical library.

helenalin Using a variety of reporter cell lines, they were able to establish that the ‘bioprobes’ induced different patterns of signalling. Experiments using a calcium chelator, BAPTAAM, showed that Ca²⁺ was involved in induction of apoptosis by the majority of the ‘bioprobes’ and that Ca²⁺ was in general required several hours into the apoptosis process. Further studies showed that the calmodulin pathway was an important mediator of the apoptotic response. Inhibition of calmodulin kinase II (CaMKII) resulted in more effective inhibition of apoptosis compared to inhibition of calpain, calcineurin/PP2B or DAP kinase. One of the ‘bioprobes’, the plant alkaloid helenalin, was used to study the role of CaMKII in apoptosis. Helenalin induced CaMKII, ASK1 and Jun-N-terminal kinase (JNK) activity, and inhibition of these kinases inhibited apoptosis.

The study shows that calcium signalling is generally not an early event during the apoptosis process and suggests that a CaMKII/ASK1 signalling mechanism is important for sustained JNK activation and apoptosis by some types of stimuli.