Bcl-2

Overview
Bcl-2 is the prototype for a family of mammalian genes and the proteins they produce. They govern mitochondrial outer membrane permeabilization (MOMP) and can be either pro-apoptotic (Bax, BAD, Bak and Bok among others) or anti-apoptotic (including Bcl-2 proper, Bcl-xL, and Bcl-w, among an assortment of others). There are a total of 25 genes in the Bcl-2 family known to date. Bcl-2 derives its name from B-cell lymphoma 2, as it is the second member of a range of proteins initially described as a reciprocal gene translocation in chromosomes 14 and 18 in follicular lymphomas.

Function of Bcl-2
There are a number of theories concerning how the Bcl-2 gene family exert their pro- or anti-apoptotic effect. An important one states that this is achieved by activation or inactivation of an inner mitochondrial permeability transition pore, which is involved in the regulation of matrix Ca2+, pH, and voltage. It is also thought that some Bcl-2 family proteins can induce (pro-apoptotic members) or inhibit (anti-apoptotic members) the release of cytochrome c in to the cytosol which, once there, activates caspase-9 and caspase-3, leading to apoptosis. Although Zamzami et al. suggest that the release of cytochrome c is indirectly mediated by the PT pore on the inner mitochondrial membrane, strong evidences suggest an earlier implication of the MAC pore on the outer membrane.



The members of the Bcl-2 family share one or more of the four characteristic domains of homology entitled the Bcl-2 homology (BH) domains (named BH1, BH2, BH3 and BH4) (see the figure on your left). The BH domains are known to be crucial for function, as deletion of these domains via molecular cloning affects survival/apoptosis rates. The anti-apoptotic Bcl-2 proteins, such as Bcl-2 and Bcl-xL, conserve all four BH domains. The BH domains also serve to subdivide the pro-apoptotic Bcl-2 proteins into those with several BH domains (e.g. Bax and Bak) or those proteins that have only the BH3 domain (e.g. Bid, Bim and Bad). The Bcl-2 family has a general structure that consists of a hydrophobic helix surrounded by amphipathic helices. Many members of the family have transmembrane domains. The site of action for the Bcl-2 family is mostly on the outer mitochondrial membrane. Within the mitochondria are apoptogenic factors (cytochrome c, Smac/DIABLO, Omi) that if released activate the executioners of apoptosis, the caspases. Depending on their function, once activated, Bcl-2 proteins either promote the release of these factors, or keep them sequestered in the mitochondria. Whereas the activated pro-apoptotic Bak and/or Bax would form MAC and mediate the release of cytochrome c, the anti-apoptotic Bcl-2 would block it, possibly through inhibition of Bax and/or Bak.

Role in disease
The Bcl-2 gene has been implicated in a number of cancers, including melanoma, breast, prostate, and lung carcinomas, as well as schizophrenia and autoimmunity. It is also thought to be involved in resistance to conventional cancer treatment. This supports a role for decreased apoptosis in the pathogenesis of cancer. Schizophrenia is a neurodegenerative chronic illness that affects about 60 million individuals world wide and is characterized by hallucinations, disorderly thought, delusions and changes in sensitivity and emotional state. The mechanism of apoptosis is connected to schizophrenia because it is known to occur in the early development of the nervous system and also eliminates injured or diseased neurons throughout life. (Schizophrenia Research. 2006; 81, 47–63). An increase in apoptosis caused by stresses such as excessive calcium flux, oxidative stress and mitochondrial dysfunction, has been found in schizophrenia. This apoptotic activity is localized in the synapses and neuritis, whose structural components contain substrates for caspases. Researchers have found that increasing apoptotic stimuli, increased caspase-3 activity, thus degenerating the neuritis and synaptic spines. "The stresses, mentioned above, increase the ratio of pro/anti-apoptotic factors, therefore increasing the likelihood of this caspase activity event which leads to schizophrenia." (Schizophrenia Research. 2006; 81, 47–63).

Apoptosis also plays a very active role in regulating the immune system. When it is functional, it can cause immune unresponsiveness to self-antigens via both central and peripheral tolerance. "In the case of defective apoptosis, it may contribute to etiological aspects of autoimmune diseases." (Clinical and Developmental Immunology. 2006. 13(2-4); 273-282). The autoimmune disease, type 1 diabetes can be caused by defective apoptosis, which leads to aberrant T cell AICD and defective peripheral tolerance. Due to the fact that dendritic cells (DCs) are of the most important antigen presenting cells of the immune system, their activity must be tightly regulated by such mechanisms as apoptosis. "Researchers have found that mice containing DCs that are Bim -/-, thus unable to induce effective apoptosis, obtain autoimmune diseases more so than those that have normal DCs."(Clinical and Developmental Immunology. 2006. 13(2-4); 273-282). "Other studies have shown that the lifespan of DCs may be controlled by factors such as a timer dependent on anti-apoptotic Bcl-2." (Clinical and Developmental Immunology. 2006. 13(2-4); 273-282). These investigations illuminate the importance of regulating antigen presentation as mis-regulation can lead to autoimmunity.

Cancer is one of the worlds leading causes of death and occurs when the homeostatic balance between cell growth and death is disturbed. Research in cancer biology has discovered that a variety of aberrations in gene expression of anti-apoptotic, pro-apoptotic and BH3-only proteins can contribute to the many forms of the disease. An interesting example can be seen in lymphomas. The over-expression of the anti-apoptotic Bcl-2 protein in lymphocytes alone did not act in an oncogenic manner. "Its combined expression with the cell cycle mitogen promoting myc gene led to an aggressive malignancy of the B-cell lineage leading to the creation of lymphomas." (Blood. 2007. 1; 16). In follicular B-cell lymphoma, a chromosomal translocation occurs between the fourteenth and the eighteenth chromosomes - t(14;18) - which places the Bcl-2 gene next to the immunoglobulin heavy chain locus. This fusion gene is deregulated, leading to the transcription of excessively high levels of anti-apoptotic bcl-2 protein. This decreases the propensity of these cells for undergoing apoptosis.

Apoptosis plays a very important role in regulating a variety of diseases that have enormous social impacts. Bcl-2 is essential to the process of apoptosis because it suppresses the initiation of the cell-death process. Further research into the family of Bcl-2 proteins will provide us with a more complete picture on how these proteins interact with each other to promote and inhibit apoptosis. An understanding of the mechanisms involved will help us find potential treatments such as inhibitors to target over-expressed proteins that may lead to disabling diseases such as cancer, neurodegenerative diseases and autoimmunity.

Targeted therapies
An antisense oligonucleotide drug Genasense (G3139) has been developed to target Bcl-2. An antisense DNA or RNA strand is non-coding and complementary to the coding strand (which is the template for producing respectively RNA or protein). An antisense drug is a short sequence of RNA which hybridises with and inactivates mRNA, preventing the protein from being formed.

It was shown that the proliferation of human lymphoma cells (with t(14;18) translocation) could be inhibited by antisense RNA targeted at the start codon region of Bcl-2 mRNA. In vitro studies led to the identification of Genasense, which is complementary to the first 6 codons of Bcl-2 mRNA.

These have shown successful results in Phase I/II trials for lymphoma, and a large Phase III trial is currently underway (Mavoromatis and Cheson 2004).

Genasense did not receive FDA approval after disappointing results in a melanoma trial.

Abbott has recently described a novel inhibitor of Bcl-2 and Bcl-xL, known as ABT-737. . ABT-737 is one among many so-called BH3 mimetic small molecule inhibitors (SMI) targeting Bcl-2 and Bcl-2-related proteins such as Bcl-xL and Mcl-1, which may prove valuable in the therapy of lymphoma and other blood cancers. .