High-throughput process development for biopharmaceutical drug substances

Quality by Design (QbD) is gaining industry acceptance as an approach towards development and commercialization of biotechnology therapeutic products that are expressed via microbial or mammalian cell lines. In QbD, the process is designed and controlled to deliver specified quality attributes consistently. To acquire the enhanced understanding that is necessary to achieve the above, however, requires more extensive experimentation to establish the design space for the process and the product. With biotechnology companies operating under ever-increasing pressure towards lowering the cost of manufacturing, the use of high-throughput tools has emerged as a necessary enabler of QbD in a time- and resource-constrained environment. We review this topic for those in academia and industry that are engaged in drug substance process development.     

Scroll-wave dynamics in human cardiac tissue: lessons from a mathematical model with inhomogeneities and fiber architecture

Cardiac arrhythmias, such as ventricular tachycardia (VT) and ventricular fibrillation (VF), are among the leading causes of death in the industrialized world. These are associated with the formation of spiral and scroll waves of electrical activation in cardiac tissue; single spiral and scroll waves are believed to be associated with VT whereas their turbulent analogs are associated with VF. Thus, the study of these waves is an important biophysical problem. We present a systematic study of the combined effects of muscle-fiber rotation and inhomogeneities on scroll-wave dynamics in the TNNP (ten Tusscher Noble Noble Panfilov) model for human cardiac tissue. In particular, we use the three-dimensional TNNP model with fiber rotation and consider both conduction and ionic inhomogeneities. We find that, in addition to displaying a sensitive dependence on the positions, sizes, and types of inhomogeneities, scroll-wave dynamics also depends delicately upon the degree of fiber rotation. We find that the tendency of scroll waves to anchor to cylindrical conduction inhomogeneities increases with the radius of the inhomogeneity. Furthermore, the filament of the scroll wave can exhibit drift or meandering, transmural bending, twisting, and break-up. If the scroll-wave filament exhibits weak meandering, then there is a fine balance between the anchoring of this wave at the inhomogeneity and a disruption of wave-pinning by fiber rotation. If this filament displays strong meandering, then again the anchoring is suppressed by fiber rotation; also, the scroll wave can be eliminated from most of the layers only to be regenerated by a seed wave. Ionic inhomogeneities can also lead to an anchoring of the scroll wave; scroll waves can now enter the region inside an ionic inhomogeneity and can display a coexistence of spatiotemporal chaos and quasi-periodic behavior in different parts of the simulation domain. We discuss the experimental implications of our study.     

Resident memory T cells (T(RM)) are abundant in human lung: diversity, function, and antigen specificity

Recent studies have shown that tissue resident memory T cells (T(RM)) are critical to antiviral host defense in peripheral tissues. This new appreciation of T(RM) that reside in epithelial tissues and mediate host defense has been studied most extensively in skin: adult human skin contains large numbers of functional T(RM) that express skin specific markers. Indeed, more than twice as many T cells reside in skin as in peripheral blood. This T cell population has a diverse T cell receptor repertoire, and can produce a broad array of cytokines. More recently, we have begun to examine other epithelial tissues for the presence of resident T cells. In the present study, we asked whether analogous populations of resident T cells could be found in human lung. We were able to demonstrate abundant resident T cells in human lung-more than 10 billion T cells were present. Lung T cells were largely of the effector memory T cell (T(EM)) phenotype, though small numbers of central memory T cells (T(CM)) and T regulatory cells (T(reg)) could be identified. Lung T cells had a diverse T cell receptor repertoire and subsets produced IL-17, IL-4, IFNγ, as well as TNFα. A significant number of lung T(RM) CD4+Th cells produced more than one cytokine, identifying them as "multifunctional" Th1 type cells. Finally, lung T(RM), but not T(RM) resident to skin or T cells from blood, proliferated in response to influenza virus. This work suggests that normal human lung contains large numbers of T(RM) cells, and these cells are poised to respond to recall antigens previously encountered through lung mucosa. This population of T cells may contribute to the pathogenesis of asthma and other T cell mediated lung diseases.     

Introgression of powdery mildew resistance from Aegilops triuncialis into wheat through induced homeologous pairing

Journal of Plant Biochemistry and Biotechnology - Powdery mildew is a serious fungal disease of wheat caused by Blumeria graminis f. sp. tritici. Chromosome 5U of Aegilops triuncialis carrying...     

Enhancing Non-linear Response of Fullerene via Incorporation of Gold Nanoparticles

Plasmonics - Bi-functional nanocomposite thin films of fullerene C60 and C70 containing Au NPs were synthesized using thermal co-evaporation method. Different atomic concentrations of Au metal...     

Chlamydia Induces Anchorage Independence in 3T3 Cells and Detrimental Cytological Defects in an Infection Model

Chlamydia are Gram negative, obligate intracellular bacterial organisms with different species causing a multitude of infections in both humans and animals. Chlamydia trachomatis is the causative agent of the sexually transmitted infection (STI) Chlamydia, the most commonly acquired bacterial STI in the United States. Chlamydial infections have also been epidemiologically linked to cervical cancer in women co-infected with the human papillomavirus (HPV). We have previously shown chlamydial infection results in centrosome amplification and multipolar spindle formation leading to chromosomal instability. Many studies indicate that centrosome abnormalities, spindle defects, and chromosome segregation errors can lead to cell transformation. We hypothesize that the presence of these defects within infected dividing cells identifies a possible mechanism for Chlamydia as a cofactor in cervical cancer formation. Here we demonstrate that infection with Chlamydia trachomatis is able to transform 3T3 cells in soft agar resulting in anchorage independence and increased colony formation. Additionally, we show for the first time Chlamydia infects actively replicating cells in vivo. Infection of mice with Chlamydia results in significantly increased cell proliferation within the cervix, and in evidence of cervical dysplasia. Confocal examination of these infected tissues also revealed elements of chlamydial induced chromosome instability. These results contribute to a growing body of data implicating a role for Chlamydia in cervical cancer development and suggest a possible molecular mechanism for this effect.     

Regulation of the catalytic activity of the EGF receptor

The epidermal growth factor receptor (EGFR) is a receptor tyrosine kinase involved in cell growth that is often misregulated in cancer. Several recent studies highlight the unique structural mechanisms involved in its regulation. Some elucidate the important role that the juxtamembrane segment and the transmembrane helix play in stabilizing the activating asymmetric kinase dimer, and suggest that its activation mechanism is likely to be conserved among the other human EGFR-related receptors. Other studies provide new explanations for two long observed, but poorly understood phenomena, the apparent heterogeneity in ligand binding and the formation of ligand-independent dimers. New insights into the allosteric mechanisms utilized by intracellular regulators of EGFR provide hope that allosteric sites could be used as targets for drug development.