Discussion comments on "Exponentiated Teissier distribution with increasing, decreasing and bathtub hazard functions".

In this note, we present some discussion comments on a note entitled 'A note on the unimodality and log-concavity of the exponentiated Teissier distribution' submitted in J. Appl. Stat.

A new class of distributions as a finite functional mixture using functional weights.

In this paper, we introduce a new family of distributions whose probability density function is defined as a weighted sum of two probability density functions; one is defined as a warped version of the other. We focus our attention on a special case based on the exponential distribution with three parameters, a dilation transformation and a weight with polynomial decay, leading to a new life-time distribution. The explicit expressions of the moments generating function, moments and quantile function of the proposed distribution are provided.

Exponentiated Teissier distribution with increasing, decreasing and bathtub hazard functions.

This article introduces a two-parameter exponentiated Teissier distribution. It is the main advantage of the distribution to have increasing, decreasing and bathtub shapes for its hazard rate function. The expressions of the ordinary moments, identifiability, quantiles, moments of order statistics, mean residual life function and entropy measure are derived.

Mitigation of Oxidation in Therapeutic Antibody Formulations: a Biochemical Efficacy and Safety Evaluation of N-Acetyl-Tryptophan and L-Methionine.

Purpose: Biotherapeutics can be susceptible to oxidation during manufacturing and storage. Free L-methionine is known to protect methionine residues in proteins from oxidation. Similarly, free tryptophan and other indole derivatives have been shown to protect tryptophan residues from oxidation.

Evaluating the Use of Antibody Variable Region (Fv) Charge as a Risk Assessment Tool for Predicting Typical Cynomolgus Monkey Pharmacokinetics.

The pharmacokinetic (PK) behavior of monoclonal antibodies in cynomolgus monkeys (cynos) is generally translatable to that in humans. Unfortunately, about 39% of the antibodies evaluated for PKs in cynos have fast nonspecific (or non-target-mediated) clearance (in-house data). An empirical model relating variable region (Fv) charge and hydrophobicity to cyno nonspecific clearance was developed to gauge the risk an antibody would have for fast nonspecific clearance in the monkey.

In silico selection of therapeutic antibodies for development: viscosity, clearance, and chemical stability.

For mAbs to be viable therapeutics, they must be formulated to have low viscosity, be chemically stable, and have normal in vivo clearance rates. We explored these properties by observing correlations of up to 60 different antibodies of the IgG1 isotype. Unexpectedly, we observe significant correlations with simple physical properties obtainable from antibody sequences and by molecular dynamics simulations of individual antibody molecules.

Specific catalysis of asparaginyl deamidation by carboxylic acids: kinetic, thermodynamic, and quantitative structure-property relationship analyses.

Asparaginyl (Asn) deamidation could lead to altered potency, safety, and/or pharmacokinetics of therapeutic protein drugs. In this study, we investigated the effects of several different carboxylic acids on Asn deamidation rates using an IgG1 monoclonal antibody (mAb1*) and a model hexapeptide (peptide1) with the sequence YGKNGG. Thermodynamic analyses of the kinetics data revealed that higher deamidation rates are associated with predominantly more negative ΔS and, to a lesser extent, more positive ΔH.

Investigation into temperature-induced aggregation of an antibody drug conjugate.

Conjugation of an antibody to a drug can produce heterogeneous species that may have different physical stabilities and safety profiles. We explored the effect of thermal stress on the physical stability, specifically aggregation, of an antibody drug conjugate (ADC), ADC 1, wherein the antibody was linked to the val-cit-Monomethyl Auristatin E (vc-MMAE) linker drug through the reduction of interchain disulfides. We also explored the effects of conjugation on the secondary and tertiary structures of ADC 1.

Characterization of antibody-polyol interactions by static light scattering: implications for physical stability of protein formulations.

In this study, the nature of interactions between monoclonal antibodies and polyols was studied using static light scattering. Solutions of mAb-U and mAb-P (4-12 mg/mL) were analyzed using static light scattering in buffer, 10% w/v trehalose and ethylene glycol solutions at pH 5.0, 7.

Isomerization of Asp-Asp motif in model peptides and a monoclonal antibody Fab fragment.

Isomerization of aspartyl (Asp or D) residues is a critical degradation route to consider for stable monoclonal antibody formulations. Among the known hotspot sequences, the DD motif is relatively understudied. To gain mechanistic insights, we used model hexapeptides, YADXFK, YADDXK, and DIDDDM, as surrogates for the hotspots in a Fab protein (YADDFK and DIDDDM), to characterize the rate-pH profile of Asp isomerization.

Application of second-derivative fluorescence spectroscopy to monitor subtle changes in a monoclonal antibody structure.

In this study, the tertiary structure of a monoclonal antibody was analyzed under thermal and chemical stresses using second-derivative fluorescence spectroscopy. The effect of polyols, sucrose, and ethylene glycol on the tertiary structure of monoclonal antibody-U (mAb-U) (pH 7.0) was studied under thermal stress (25°C-75°C).

Culture temperature modulates aggregation of recombinant antibody in cho cells.

During production of therapeutic monoclonal antibodies (mAb), it is highly desirable to remove and control antibody aggregates in the manufacturing process to minimize the potential risk of immunogenicity to patients. During process development for the production of a recombinant IgG in a CHO cell line, we observed atypical high variability from 1 to 20% mAb aggregates formed during cell culture that negatively impacted antibody purification. Analytical characterization revealed the IgG aggregates were mediated by hydrophobic interactions likely caused by misfolded antibody during intracellular processing.

Opposite effects of polyols on antibody aggregation: thermal versus mechanical stresses.

Purpose: To investigate the physical stability of antibody-polyol formulations under thermal and mechanical stresses.

Methods: mAb-U was analyzed in buffer, trehalose, sucrose, glycerol and ethylene glycol solutions at pH 7.0.

Solubilities and transfer free energies of hydrophobic amino acids in polyol solutions: importance of the hydrophobicity of polyols.

The purpose of this work was to investigate the difference in the hydrophobicities of various polyols and the nature of interactions between hydrophobic amino acid side chains and polyols. The interactions were explored by conducting solubility studies of three amino acid derivatives, N-acetyl tryptophanamide (NATA), N-acetyl leucinamide (NALA), and N-acetyl glycinamide (NAGA), in the solutions of sorbitol, sucrose, trehalose, glycerol, ethylene glycol, ribose, and deoxyribose. Hydrophobicity index of polyols was calculated using molecular modeling.

Modulation of the thermodynamic stability of proteins by polyols: significance of polyol hydrophobicity and impact on the chemical potential of water.

The exact mechanism of the modulation of chemical potential of proteins by polyols is not yet well understood. Present study investigates the role of hydrophobicity of polyols, and their impact on water activity and/or surface tension, in determining their stabilization/destabilization potential. Results with ribose and methyl-glucose show that the enhanced stability of proteins is not mediated via the effect on interfacial tension, a hypothesis that has so far been restricted to glycerol.

Physicochemical stability of the antibody-drug conjugate Trastuzumab-DM1: changes due to modification and conjugation processes.

In the manufacture of the antibody-drug conjugate Trastuzumab-DM1 (T-DM1), the lysine residues on the antibody trastuzumab (Tmab) are modified to form the intermediate Tmab-MCC (T-MCC) and then conjugated with the drug DM1. Our goal is to understand the effects of modification and conjugation steps on the physicochemical stability of the antibody. The structural stability of Tmab relative to its modified and conjugated forms was assessed, employing thermally induced stress conditions to formulations containing Tmab, T-MCC, and T-DM1.

Cold denaturation of monoclonal antibodies.

The susceptibility of monoclonal antibodies (mAbs) to undergo cold denaturation remains unexplored. In this study, the phenomenon of cold denaturation was investigated for a mAb, mAb1, through thermodynamic and spectroscopic analyses. Tryptophan fluorescence and circular dichroism (CD) spectra were recorded for the guanidine hydrochloride (GuHCl)-induced unfolding of mAb1 at pH 6.

Triple light chain antibodies: factors that influence its formation in cell culture.

THIOMABs are recombinant antibodies engineered with reactive cysteines, which can be covalently conjugated to drugs of interest to generate targeted therapeutics. During the analysis of THIOMABs secreted by stably transfected Chinese Hamster Ovary (CHO) cells, we discovered the existence of a new species--Triple Light Chain Antibody (3LC). This 3LC species is the product of a disulfide bond formed between an extra light chain and one of the engineered cysteines on the THIOMAB.

Role of benzyl alcohol in the prevention of heat-induced aggregation and inactivation of hen egg white lysozyme.

The aim of the study was to investigate the stability of a model protein, lysozyme, in the presence of the commonly used preservative benzyl alcohol. Techniques including lytic assay, size exclusion chromatography, circular dichroism, differential scanning calorimetry, native polyacrylamide gel electrophoresis and dynamic light scattering were used to study the overall stability of lysozyme in the presence of benzyl alcohol. The stability of lysozyme against thermal stress was higher in the presence of benzyl alcohol.

In situ precipitation and vacuum drying of interferon alpha-2a: development of a single-step process for obtaining dry, stable protein formulation.

Feasibility studies were performed to develop a process for obtaining stable dry protein formulations based on in situ polyethylene glycol (PEG)-induced precipitation and vacuum drying of interferon alpha-2a (IFNalpha2a) solution in a vial. Using a laboratory scale freeze dryer, the process was carried out in two phases: first, protein solution containing PEG was concentrated to achieve protein precipitation, and second, remaining water was removed by further reducing the chamber pressure. Drying conditions, i.

Effect of polyols on polyethylene glycol (PEG)-induced precipitation of proteins: Impact on solubility, stability and conformation.

Effect of polyols on the solubility of bovine serum albumin (BSA) in the presence of polyethylene glycols (PEGs) was investigated in order to strengthen the understanding of the observed effects of polyols and PEGs on protein properties in solution. Effect of polyols and/or PEGs on the thermodynamic (conformational) stability of BSA was measured using DSC and circular dichroism (CD). Glucose, sucrose, raffinose, glycerol and sorbitol, all reduced the extent of protein precipitation.

A method to investigate non-thermal effects of radio frequency radiation on pharmaceuticals with relevance to RFID technology.

A method is reported to accurately and precisely control temperature of a solution sample to investigate non-thermal effects of radio frequency radiation (RFR) on pharmaceuticals. This method utilizes a transverse electromagnetic (TEM) cell connected in series with a radiation source. The temperature of a sample under study, within the TEM cell, is regulated using a combination of a fiber-optic thermometer and thermo-electric cooler.

A high-throughput method for detection of protein self-association and second virial coefficient using size-exclusion chromatography through simultaneous measurement of concentration and scattered light intensity.

Purpose: To characterize protein self-association along with second virial coefficient (a measure of solution nonideality) using size-exclusion chromatography (SEC) utilizing a novel flow cell that is capable of simultaneously measuring protein concentration and scattered light intensity.

Methods: beta-lactoglobulin A (beta Lg), known to exhibit NaCl-dependent monomer-dimer equilibrium at pH 3.0, was used as the model protein.

Moisture-induced aggregation of lyophilized DNA and its prevention.

Purpose: To investigate the moisture-induced aggregation (i.e., a loss of solubility in water) of DNA in a solid state and to develop rational strategies for its prevention.

Protein structural conformation and not second virial coefficient relates to long-term irreversible aggregation of a monoclonal antibody and ovalbumin in solution.

Purpose: The purpose of the study was to investigate the relationship of the second virial coefficient, B22, to the extent of irreversible protein aggregation upon storage.

Methods: A monoclonal antibody and ovalbumin were incubated at 37 degrees C (3 months) under various solution conditions to monitor the extent of aggregation. The B22 values of these proteins were determined under similar solution conditions by a modified method of flow-mode static light scattering.