Stefanie+Schwemlein

Journal Article Summary:
 * Immunotherapeutic applications of CpG oligodeoxynucleotide TLR 9 agonists**

Jörg Vollmer, Arthur M. Krieg Innate immune detection of and response to foreign bodies like microbes and viruses are a result of the immune system’s using receptors that can sense infectious molecules. The specific family of receptors studied here, the Toll-like receptors (TLRs), are highly specific and detect different microbes depending on the receptor. They evolve in order that they might detect new infections and trigger immune responses to protect the body from those foreign invaders. The specific TLR9 detects DNA, and elicits immune response in when pathogenic DNA is detected. This study examined the therapeutic applications of stimulating this receptor which in the body is activated by unmethylated CpG dinucleotides, which are seen most often in bacterial and viral DNA, but which are suppressed in vertebrate DNA. TLR9 can be activated by synthetically-produced CpG oligodeoxynucleotides, and clinical trials have shown that these compounds can act as effective vaccine adjuvants. The observed stimulatory effects of CpG are a result of differences inherent to genomic DNA of vertebrates and pathogens (methylation and frequency-suppression in vertebrate CpG dinucleotides). By activating innate immunity, TLR9-stimulation also expands antigen-specific humoral and cellular immune responses which can act against a number of antigens. Agonists of TLR 9 receptors have potential to act as vaccine adjuvants as well as therapies for the treatment of cancer and infectious and allergic diseases, both alone and in combination with other drugs. The CpG oligodeoxynucleotides promote the activation and maturation of cells of the immune system as well as produce more antibody-secreting cells. The use of these TLR 9 agonists has undergone phase I and II clinical trials which show their ability to enhance the development of T-cell responses and display antitumor activity when used as supplement to vaccines. The agonists have even been shown to act against asthma and other allergic diseases, suggesting that they may elicit responses against a number of diseases. Questions remain in how safe and effective these TLR 9 agonists are in humans.

Vollmer, J�rg, and Arthur M. Krieg. “Immunotherapeutic applications of CpG oligodeoxynucleotide TLR9 agonists.” Advanced Drug Delivery Reviews 61, no. 3 (March 28, 2009): 195-204.

Journal Article Summary: Danielle R. Mercatante, Carl D. Bortner, John A. Cidlowski, and Ryszard Kole Studies have shown that deregulation of apoptosis, or programmed cell death, can facilitate the development and progression of cancers. Humans have a number of apoptotic regulatory genes that such as the one studied here at UNC-Chapel Hill, Bcl-x. This gene codes for protein variants as a result of alternative splicing, and they may have different, and in some cases opposing, functions. Dr. Kole and the other researchers mentioned are interested in the utilization of antisense oligonucleotides in order to manipulate alternative splicing. In this particular case, the oligonucleotides were used to control the alternative splicing of the Bcl-x gene in a way that could better regulate apoptosis in cancerous cells. The researchers used oligonucleotides antisense to the 5’ splice site of Bcl-x, shifting the splicing pattern of the pre-mRNA of Bcl-x from Bcl-xL, an anti-apoptotic gene overexpressed in a number of cancers, to Bcl-xS, a splicing variant of the gene which is pro-apoptotic. It was found that this method was highly successful in PC3 prostate cancer cells, significantly increasing apoptosis in those cells. The same methods were used on MCF7 breast cancer cells, but in this second case, the treatment brought about a considerably weaker apoptotic response. Additionally, the induced shift in pre-mRNA splicing resulted in a hindering of colony formation in both the prostate cancer and breast cancer cells, but again, the results were not as significant in the breast cancer cells. These discrepancies seen between the treatments with the antisense oligonucleotides were analyzed in the context of the different isoforms of the protein coded by the Bcl-x gene, and it was concluded that the effects of the modification of the alternative splicing of the Bcl-x apoptotic regulatory gene depend on the specific characteristics of the cell whose gene is being modified through oligonucleotide treatment. Future studies should examine how this method can be applied to different types of genes and how it affects the expression of those genes.
 * Modification of alternative splicing of Bcl-x pre-mRNA induces apoptosis in prostate and breast cancer cells: Analysis of apoptosis and cell death**

Mercatante, Danielle R., Carl D. Bortner, John A. Cidlowski, and Ryszard Kole. “Modification of alternative splicing of Bcl-x pre-mRNA induces apoptosis in prostate and breast cancer cells: Analysis of apoptosis and cell death.” J. Biol. Chem. (February 7, 2001): M009256200.

Journal Article Summary: Currently we do not have a marketed antiviral molecule to treat poxvirus infection, and because smallpox was declared by the World Health Organization to have been eradicated, vaccinations for the disease are no longer being administered. Because the use of biological weapons is becoming more common, as seen in the use of anthrax and other biological substances, and the outbreaks that follow can be fatal to a large number of the population as the disease continues to spread, it is necessary that we have treatments available post-infection. A20 protein, important to the vaccinia virus replication complex, binds to proteins that are necessary for viral DNA synthesis. A new idea for the design of anti-orthopoxvirus drugs to combat smallpox is to identify and use molecules that are able to inhibit the activity of this virus replication complex and to inhibit it. In this particular study peptide aptamers, which are proteins which inhibit target protein functioning, that interact with this A20 protein were examined. A specific aptamer (aptamer 72) was determined to bind to a region on the A20 protein that is important in DNA replication, therefore inhibiting viral replication and presenting a potential treatment to poxvirus infection. In order to determine whether DNA synthesis was actually inhibited, the researchers tested the inhibitory effect of two of the aptamers. Synthesis was quantified by rtPCR (real-time Polymerase Chain Reaction), and the control showed a higher production of DNA than those expressing the aptamers, therefore the data suggests that these aptamers do affect the replication of DNA. The study showed that vaccinia virus DNA synthesis was hindered and virus production was slowed down in cells that expressed aptamer 72, and therefore the aptamer could be a useful tool in the development of new compounds targeting poxvirus replication.
 * Inhibition of Vaccinia Virus Replication by Peptide Aptamers**

Saccucci, Laurent, Jean-Marc Crance, Pierre Colas, Marc Bickle, Daniel Garin, and Frédéric Iseni. “Inhibition of vaccinia virus replication by peptide aptamers.” Antiviral Research 82, no. 3 (June 2009): 134-140.

Journal Article:

Simvastatin strongly reduces levels of Alzheimer’s disease in beta amyloid peptides amyloid beta 42 and amyloid beta 40 in vitro and in vivo

Statin drugs have recently been shown to decrease the occurrence of AD and dementia in studies that were not related to neurodegenerative disorders. Statins and the APOE isoform apoE4 are believed to affect AD patients in similar ways. Normally, apoE’s function is to carry and deliver lipids to the liver, but the isoform apoE4 seems to carry out this function improperly, as its presence results in an increased risk for atherosclerosis and amyloid plaque development. Because cellular cholesterol levels affect amyloid beta production, it is believed that apoE4’s inability to remove lipids as efficiently from the bloodstream to contribute to the accumulation of beta amyloid plaques in the brain and therefore the disease’s progression. The overproduction minor amyloid beta isoform, amyloid beta 42, produced by the different APP cleaving sites of beta and gamma secretases, has also been identified as an important risk factor for AD. In this study, simvastatin and lovastatin were used (along with methyl-beta-cyclodextrin which normally removes cholesterol from the plasma membrane) to determine whether they could potentially be used as AD treatments to reduce intracellular and secretary amyloid beta (and especially AB42) levels. The study showed that the use of simvastatin and lovastatin reduce intra- and extracellular levels of amyloid beta 42 and amyloid beta 40 levels in vitro and simvastatin also reduced cerebral AB levels in vivo. The cell cultures used in this study were obtained from hippocampan neurons of fetal rats. The neurons were exposed to lovastatin and simvastatin for different amounts of time. A recombinant virus encoding APP65 was prepared and the neurons were infected with the virus. The cells were then placed in maintenance medium and the cells were incubated; the amyloid beta levels were reduced more significantly than expected.

Fassbender, K., M. Simons, C. Bergmann, M. Stroick, D. Lütjohann, P. Keller, H. Runz, et al. “Simvastatin Strongly Reduces Levels of Alzheimer's Disease β-Amyloid Peptides Aβ42 and Aβ40 in vitro and in vivo.” Proceedings of the National Academy of Sciences of the United States of America 98, no. 10 (May 8, 2001): 5856-5861.