Tag: <span>stability</span>

Due to its antioxidant properties and favorable safety profile, glutathione (GSH) finds use in protein formulations by improving overall protein stability. Once degraded, primarily by oxidation into glutathione disulfide (GSSG), the protecting effect of GSH is lost. A simple, direct method using reversed-phase separation and charged-aerosol detection (RP-CAD) to quantitate GSH is described in this paper. The analytical methodology is also capable of monitoring several by-product degradants of GSH, both oxidative and non-oxidative. For high-concentration protein formulations, the method provides direct analysis of GSH and its degradants in the presence of protein at up to 225 mg/mL simply through a dilution of the sample. Quantitation of many amino acids typically included in pharmaceutical protein formulations is also possible. Use of an online diverting valve in the method prevents interference in the detector from the high protein concentration in formulation. Accuracy and effectiveness of this method is demonstrated through monitoring the stability of GSH in high-concentration protein formulations through confirmation of GSH concentration and mass-balance of its loss over time. Monitoring GSH stability in protein formulations is necessary, as GSH concentration is indicative of protein stability.

Biologics Production

Today, technology has reached a point where organisms (bacteria, plant and animal cells) can be genetically engineered to produce specific macromolecules and perform complex chemical reactions. Hence, they are called ‚Äúcellular factories.‚ÄĚ Cellular factories have applications in: biomedicine (e.g., implanted insulin secreting cells for the management of diabetes); biotechnology (recombinant protein and enzyme production for pharmaceutical and food industries); bioremediation (toxic waste and pollutant clean-up); green chemistry (production of chemicals with minimum toxic bi-product generation); alternative energy generation (electricity and hydrogen production by bacteria); biosensors (e.g., devices housing ‚Äúcanary cells‚ÄĚ, which can signal the presence of pollutants, viral agents, or toxic chemicals); bioreactive devices (that can detect low concentrations of chemicals, etc.)…

Biologics Production Cell & Gene Therapy

Since the first approval for human use of a recombinant protein therapeutic, this sector of the pharmaceutical market has grown rapidly. The first approved protein therapeutics were small, non-glycosylated proteins such as insulin and human growth hormone; they were produced in bacterial systems. With the advent of mammalian cell-based production systems, it became possible to produce more complex, glycosylated proteins for use as recombinant therapeutics…

Biologics Production

The development and commercialization of biopharmaceutical products is paving the way for significant growth in the pharmaceutical industry. Virtual and small biotech companies are researching and developing new biotechnology-based products that help large pharmaceutical companies backfill their pipelines. Many of these products have been in development for years and are finally reaching the point of commercialization. Because of the nature of many of these substances, they are sensitive to their environment, their packaging and their delivery systems…

Manufacturing

In recent years, cell therapy has been suggested as a promising approach for repair and regeneration of damaged tissues. VesCell‚ĄĘ, a blood-derived autologous cell therapy product consisting of ex vivo enriched angiogenic cell precursors (ACPs) was developed by TheraVitae for the treatment of severe heart diseases. A non-mobilized, blood-derived cell population consisting of low density cells, termed synergetic cell population (SCP), was isolated and cultured in the presence of serum-free medium (X-Vivo 15, Lonza, Walkersville, MD, USA) supplemented with growth factors and autologous serum to yield VesCell. Significant cell numbers (>50×106) exhibiting morphological, immunocytochemical, and functional characteristics of the angiogenic cell lineage were obtained from blood samples. The ACPs expressed the hematopoietic stem cell (HSC) markers CD34, CD133 and CD117, as well as specific angiogenic markers such as vascular endothelial growth factor receptor 2 (VEGFR2) (receptor 2 [R2] is also known as kinase domain region [KDR]), CD144, and CD31‚Ķ

Cell & Gene Therapy Manufacturing Uncategorized

Proteins are widely used in research, medicine and industry, but its extraction from their natural sources can be difficult, tedious and expensive. Therefore, a simple and inexpensive system that allows large-scale production of safe recombinant proteins will always be highly desirable. Traditional production systems that use microbial, insect and mammalian cell cultures have drawbacks, in terms of cost, scalability and product safety. Several studies have shown that molecular farming in plants has many practical, economic, and safety advantages as compared to these conventional methods. Thus, the use of plants for recombinant protein synthesis is gaining wide acceptance…

Biologics Production

Biopharmaceutical companies are constantly evaluating new methods for mammalian cell line development that provide benefits such as shorter timelines, improved consistency, higher production, better genetic stability, and increased flexibility. Each of these advantages extends a large cost benefit to companies as their recombinant protein products are moved from development into the clinic and finally to commercial launch. A versatile system has been developed that is capable of transferring genes of interest into a wide variety of mammalian host cells and offers a number of the above advantages over other methods. The system, which is referred to as GPEx‚ĄĘ (an acronym for ‚Äúgene product expression‚ÄĚ), utilizes replication-defective retroviral vectors, derived from Moloney murine leukemia virus (MLV) and pseudotyped with vesicular stomatitis virus G protein (VSV-G), to stably insert single copies of genes into dividing cells‚Ķ

Biologics Production Viral Vectors

Baculovirus, particularly AcMNPV (Autographa californica multiple nucleocapsids polyhedrosis virus), is widely used for heterologous protein expression. There are several shortcomings in the current practice of preserving and scaling up baculovirus: 1) extracellular baculovirus stocks, routinely prepared in large volumes and stored at 4¬ļ C, are often unstable; 2) laborious and time-consuming steps to amplify and titer the baculovirus stocks are often necessary, and generally recommended, for achieving consistent viral infection and protein expression; 3) once prepared, the baculovirus is suspended and stored in conditioned medium. Given the complex, undefined, and unstable nature of the spent media components, including proteases and nucleases, protein expression tends to vary even when steps are taken to titer the virus stock and adjust the amount of stock used for infection. Here, we will report a new method for preserving and scaling up baculoviruses that: 1) provides a new form of viral stock more stable than the traditional, extracellular stock; 2) eliminates the need for virus amplification and retitering; 3) drastically reduces the turn-around time and resources required for scale-up; and 4) improves yield and consistency in protein expression.

Baculovirus Expression Technology

There is an increasing emphasis in clinical and translational research on the discovery and development of biomarkers that are indicative of a disease state. While biomarkers are not exclusively proteins, the emergence of new mass spectrometry platforms combined with the human genome databases has rejuvenated the search for biomarker proteins, especially in readily available body fluids such as blood. There is currently a tremendous need for an improved ability to “mine” the full depth of the proteome in a high throughput manner. To advance clinical proteomics, methodologies are needed that can accommodate higher throughput while facilitating the ability to observe large numbers of protein events…

Biologics Production Research

lation components that stabilize the molecule in order to provide the desired product storage stability. Generally, an aqueous formulation is preferred; however, the instability of proteins, both physical (e.g. aggregation) and chemical (e.g. deamidation and oxidation), often necessitates the development of lyophilized formulations. In these formulations, selection of the appropriate stabilizing cryoprotectants, lyoprotectants, and bulking agents is critical. Accelerated stability studies are typically used to evaluate the effect of a single factor at a time in order to identify the optimum pH, buffer, and stabilizing excipients. This approach is limited in that many independent time-consuming experiments must be run, the results are obtained only at the evaluated set points, and additional experiments are required to assess potential interactions between the evaluated factors…

Biologics Production