Challenges and Strategies for a Thorough Characterization of Antibody Acidic Charge Variants.

Heterogeneity of therapeutic Monoclonal antibody (mAb) drugs are due to protein variants generated during the manufacturing process. These protein variants can be critical quality attributes (CQAs) depending on their potential impact on drug safety and/or efficacy. To identify CQAs and ensure the drug product qualities, a thorough characterization is required but challenging due to the complex structure of biotherapeutics. Past characterization studies for basic and acidic variants revealed that full characterizations were limited to the basic charge variants, while the quantitative measurements of acidic variants left gaps. Consequently, the characterization and quantitation of acidic variants are more challenging. A case study of a therapeutic mAb1 accounted for two-thirds of the enriched acidic variants in the initial characterization study. This led to additional investigations, closing the quantification gaps of mAb1 acidic variants. This work demonstrates that a well-designed study with the right choices of analytical methods can play a key role in characterization studies. Thus, the updated strategies for more complete antibody charge variant characterization are recommended.

Fractional re-distribution among cell motility states during ageing.

Ageing in humans is associated with the decreased capacity to regulate cell physiology. Cellular properties, such as cell morphology and mechanics, encode ageing information, and can therefore be used as robust biomarkers of ageing. Using a panel of dermal fibroblasts derived from healthy donors spanning a wide age range, we observe an age-associated decrease in cell motility. By taking advantage of the single-cell nature of our motility data, we classified cells based on spatial and activity patterns to define age-dependent motility states. We show that the age-dependent decrease in cell motility is not due to the reduced motility of all cells, but results from the fractional re-distribution among motility states. These findings highlight an important feature of ageing cells characterized by a reduction of cellular heterogeneity in older adults relative to post-adolescent/adults. Furthermore, these results point to a mechanistic framework of ageing, with potential applications in deciphering emergent ageing phenotypes and biomarker development.

Evolution of cellular morpho-phenotypes in cancer metastasis.

Intratumoral heterogeneity greatly complicates the study of molecular mechanisms driving cancer progression and our ability to predict patient outcomes. Here we have developed an automated high-throughput cell-imaging platform (htCIP) that allows us to extract high-content information about individual cells, including cell morphology, molecular content and local cell density at single-cell resolution. We further develop a comprehensive visually-aided morpho-phenotyping recognition (VAMPIRE) tool to analyze irregular cellular and nuclear shapes in both 2D and 3D microenvironments. VAMPIRE analysis of ~39,000 cells from 13 previously sequenced patient-derived pancreatic cancer samples indicate that metastasized cells present significantly lower heterogeneity than primary tumor cells. We found the same morphological signature for metastasis for a cohort of 10 breast cancer cell lines. We further decipher the relative contributions to heterogeneity from cell cycle, cell-cell contact, cell stochasticity and heritable morphological variations.

Particle margination and its implications on intravenous anticancer drug delivery.

"Margination" refers to the movement of particles in flow toward the walls of a channel. The term was first coined in physiology for describing the behavior of white blood cells (WBCs) and platelets in blood flow. The margination of particles is desirable for anticancer drug delivery because it results in the close proximity of drug-carrying particles to the endothelium, where they can easily diffuse into cancerous tumors through the leaky vasculature. Understanding the fundamentals of margination may further lead to the rational design of particles and allow for more specific delivery of anticancer drugs into tumors, thereby increasing patient comfort during cancer treatment. This paper reviews existing theoretical and experimental studies that focus on understanding margination. Margination is a complex phenomenon that depends on the interplay between inertial, hydrodynamic, electrostatic, lift, van der Waals, and Brownian forces. Parameters that have been explored thus far include the particle size, shape, density, stiffness, shear rate, and the concentration and aggregation state of red blood cells (RBCs). Many studies suggested that there exists an optimal particle size for margination to occur, and that nonspherical particles tend to marginate better than spherical particles. There are, however, conflicting views on the effects of particle density, stiffness, shear rate, and RBCs. The limitations of using the adhesion of particles to the channel walls in order to quantify margination propensity are explained, and some outstanding questions for future research are highlighted.