lant should be back online after hydrogen peroxide treatment
Contamination of a bioreactor with a rare virus caused Genzyme Corp., of Cambridge, Mass., to close one of its plants temporarily to sanitize the facility. The cleanup of the plant in Allston, Mass., is “on track,” according to a spokeswoman.
“We’re continuing the vaporous hydrogen peroxide treatment at the facility, and we believe it will all be complete by the end of July,” Lori Gorski, a spokeswoman for Genzyme, told Pharmaceutical Formulation & Quality in an interview.
Vesivirus 2117 was identified in a bioreactor used in the production of Cerezyme (imiglucerase for injection) at the Allston plant. The plant also produces Fabrazyme (agalsidase beta). Cerezyme is used to treat Gaucher disease; Fabrazyme treats Fabry disease.
Vesivirus 2117 impairs the viability of cells used in the manufacturing process and is not known to cause infection in humans, Genzyme said in a June 25 press release. The company confirmed that the same virus strain contaminated production twice in 2008, once at the Allston plant and once at a plant in Belgium.
The virus is highly uncommon, Gorski said. “We’ve found only one publication that references Vesivirus 2117,” she said. A highly specific assay for the virus, developed by Genzyme, will be made “widely available” to the industry, the press release said.
It is likely that the virus was introduced from raw material used in the manufacturing process, said the company, which is working with suppliers to address the problem and prevent recurrence. The company said it is also evaluating adding steps to its raw materials screening and virus removal processes to make them more robust, including using the new assay to test all raw materials for the presence of the virus.
Crystal Rice, a U.S. Food and Drug Administration (FDA) spokeswoman, declined to comment on the contamination because it “continues to be under discussion and investigation.”
Production interruption will mean shortages of the two drugs for a period of time, the company announced. A section of Genzyme’s Web site informs patients and physicians about plans to ration the drugs (www.genzyme.com/supplyupdate).
“Because Cerezyme and Fabrazyme inventories are not sufficient to avoid shortages during the period of suspended production and recovery, Genzyme has begun working with regulatory authorities, physicians, and patient organizations to carefully manage product supply, with the goal of protecting the most vulnerable patients,” the press release said.
Gaucher disease and Fabry disease are both rare genetic disorders, and the company said it has met with representatives of the National Gaucher Foundation and others to devise guidelines to determine who needs the drugs the most. A period of “constrained supply” is expected to begin in August for Cerezyme and in October for Fabrazyme and should last six to eight weeks in each case, the company said.
Although production has been shut down, there were some finished lots of Cerezyme in inventory. Polymerase chain reaction testing found no evidence of Vesivirus 2117 in these lots, the company said, and after conferring with the FDA and its European equivalent, the European Medicines Agency, Genzyme was allowed to ship the product. Shipments of existing lots of Fabrazyme were not put on hold, the company said.
The sanitization process at the Allston plant includes disassembly and removal of equipment, cleaning, disinfection of cell culture and downstream production suites with vaporous hydrogen peroxide, installation of new air filters and insulation, and other measures, according to the press release. Once the sanitization has concluded, equipment will be reassembled, tested, and brought back into operation; production will resume on a rolling basis. Genzyme will not have to perform process validation runs to resume production.
New Therapeutic Targets for Alzheimer’s
Link between soluble amyloid beta proteins and long-term synaptic depression
For the first time, researchers have found a link between soluble amyloid beta (A beta) protein and the synaptic impairment found in Alzheimer’s disease. These findings could offer new targets for treatment of Alzheimer’s disease, the most common neurodegenerative disorder.
“Soluble amyloid beta facilitates long-term synaptic depression (LTD) through a new mechanism of interrupted glutamate uptake,” Shaomin Li, PhD, MD, Neurology, Brigham and Women’s Hospital, Boston, told Pharmaceutical Formulation & Quality. “These findings could lead to a new direction to treat Alzheimer’s disease. If we can develop some compounds or other methods that rescue the A beta-enhanced LTD or reduce the glutamate levels caused by A beta-interrupted glutamate transporters, it should be a new direction for the treatment of Alzheimer’s disease.”
This is the first study to find this link. “This work by the Selkoe Lab is the first to demonstrate a credible mechanistic link between A beta and LTD,” said David E. Kang, PhD, assistant adjunct professor in the School of Medicine at the University of California, San Diego in La Jolla, Calif. (Li S, Hong S, Shepardson NE, et al. Soluble liogomers of amyloid Beta protein facilitate hippocampal long-term depression by disrupting neuronal glutamate uptake. Neuron. 2009;62(6):788-801.)
Alzheimer’s disease is characterized by progressive memory and cognitive impairment. Although the precise cause of the disease remains unknown, the literature suggests that A beta plays a critical pathogenic role, according to the authors. Several studies have shown that soluble A beta proteins inhibit hippocampal long-term potentiation; however, their role in LTD—a cellular communication phenomenon that has been linked to neuronal degeneration—is unclear.
— David E. Kang, PhD, UC San Diego
Soluble A Beta-Facilitated Depression
The results showed that soluble A beta oligomers facilitated LTD in the hippocampus, a brain region associated with memory, by inhibiting uptake of glutamate—a major excitatory neurotransmitter. According to various theories, glutamate plays a role in the progressive neuronal loss associated with Alzheimer’s disease.
“The enhanced LTD due to increased extracellular glutamate levels could explain some previous other findings, such as neurotoxicity, epilepsy, neuronal death, and spine shrinkage,” Dr. Li said.
“LTD is the opposite of long-term potentiation (LTP), which facilitates synaptic connectivity in the form of new memories,” Dr. Kang said. “The claim is that glutamate recycling is perturbed by A beta, which ultimately means that less glutamate can be taken up by the synapse to be reused to elicit more connectivity between synapses. Synapses are communication points between nerve cells. Without them, there can be no communication. More LTD unfortunately means less connectivity between nerve cells and less capacity to store and retrieve information.”
Dr. Li and colleagues also showed that LTD could be prevented by an extracellular glutamate scavenger system.
New Therapeutic Targets
“While more research would be needed to confirm these results and their relevance to Alzheimer’s disease, e.g., do native endogenous oligomers modify LTD in the brain of AD patients, this work provides new therapeutic targets for development, such as the p38/MAPK pathway that appears to be critically involved in A beta oligomer-mediated LTD facilitation,” said Jorge Busciglio, PhD, associate professor of neurobiology and behavior, University of California-Irvine. “The results by Selkoe and colleagues further underscore the importance of soluble forms of A beta as pathogenic agents acting locally and leading to synaptic dysfunction.”
Designing therapies to block A beta’s action could be the key to preventing early neurodegenerative changes. “The important implication here is that simply blocking the action, availability, or activity of A beta to the most vulnerable sites (such as the synapse) may be sufficient to impede the most immediate neurodegenerative action of A beta at the level of the synapse or connectivity before nerve cells begin to die for good,” Dr. Kang said. “In other words, therapeutic agents designed to block the actions of soluble A beta at the synapse could repel some of the early neurodegenerative changes in Alzheimer’s disease before they become irreversible.”
Artificial Liver on Horizon for Testing New Drugs
Model maintained liver-specific functions for three weeks
—Heike Mertsching, PhD, Fraunhofer Institute for Interfacial Engineering and Biotechnology IGB
German researchers have developed an artificial liver that offers a potential alternative to testing new drugs on animals.
“With this model, substances which cause side effects, e.g., by the formation of toxic metabolites, could be detected early and thereby animal experiments could be reduced,” Heike Mertsching, PhD, Fraunhofer Institute for Interfacial Engineering and Biotechnology IGB, Stuttgart, Germany, told Pharmaceutical Formulation & Quality. “Furthermore, the results of animal testing, especially concerning liver toxicity, are not directly transferable to the human organism. Species-specific differences in enzyme activities lead to about 30% false negative or false positive prognoses.”
Dr. Mertsching and colleague Johanna Schanz, PhD, designed an artificial liver model from a 10 cm to 15 cm-long porcine jejunal segment with a vascular system, an arterial flow, and venous reflux, Dr. Mertsching said.
To remove the pig cells, the researchers perfused the intestine with a sodium desoxycholate solution, Dr. Mertsching said. A buffer solution removed any remaining cell debris. The researchers incubated the matrix in a DNase solution to eliminate any leftover pig DNA.
“Before we integrated the DNase step in the protocol, DNA leftovers could sometimes be detected on the mucosa structures, but this problem is solved now,” Dr. Mertsching said. The researchers confirmed decellularization by examining the tissue macroscopically, Dr. Mertsching said. “The matrix turns from flesh-colored to snow white. Furthermore, quantification of isolated DNA, as well as histological analysis of matrix samples, verifies the decellularization.”
Simulates Physiological Conditions
After removing the pig cells, the researchers seeded human hepatocyte and endothelial cells onto the structure. To supply the cells and tissues with nutrients, they put the model into a computer-controlled bioreactor with a flexible pump tube developed at Fraunhofer IGB.
“Our system is cultivated in a bioreactor system simulating physiological conditions,” Dr. Mertsching said. “This means our culture medium is pumped through the vascular system with normal blood pressure of 80 mmHG to 120 mmHG. Furthermore, the mechanical impulse through blood flow shows induction of endothelial cell differentiation or sustainability of their differentiation status.”
The cells were active for an extended period. “We could maintain liver-specific functions like albumin or urea synthesis as well as biotransformation processes over three weeks,” Dr. Schanz told Pharmaceutical Formulation & Quality. The three-week time frame allowed long-term studies and analysis of multiple applications or accumulation of substances in the liver.
This liver model is different from existing models, which are based on isolated liver enzymes, primary cell cultures, or liver slices. These systems rapidly lose function and, as a result, only permit short-term study—a maximum of two days, researchers say. Dr. Schanz is optimistic that a longer cultivation time may be possible.
Mimics Human Cells
Just as they do in the human body, these cells detoxify, break down drugs, and build up proteins; these capabilities are important for drug testing or transplantation, because the effect of a drug can change when it is broken down in the body. After ingestion, drugs enter the liver and are metabolized by the hepatocytes. The resulting metabolites may have different pharmacological properties than the original drug, Dr. Mertsching said. To maintain isolated hepatocyte function, it is critical to imitate their in vivo microenvironment, including the co-culture with non-parenchymal cells, a suitable 3-D scaffold, and a sufficient nutrient and gas supply.
“Our liver model possesses an integrated vascular system, which allows the application of substances over, and the analysis of their metabolites in, the blood circuit,” she said. Over the next two years, the researchers will complete extensive validation studies on the liver model. After the testing is complete, they will consider how to bring their model to market. n