Plants metabolize polychlorinated biphenyls (PCBs) into hydroxylated derivatives (OH-PCBs), which are sometimes more toxic than the parent PCBs. The objective of this research was to compare the toxicity of a suite of PCBs and OH-PCBs toward the model plant, Arabidopsis thaliana. While parent PCBs and higher-chlorinated OH-PCBs exhibited a low or nondetectable toxicity, lower-chlorinated OH-PCBs significantly inhibited the germination rate and plant growth, with inhibition concentration 50% (IC50) ranging from 1.6 to 12.0 mg L−1. The transcriptomic response of A. thaliana to 2,5-dichlorobiphenyl (2,5-DCB), and its OH metabolite, 4′-OH-2,5-DCB, was then examined using whole-genome expression microarrays (Affymetrix). Exposure to 2,5-DCB and 4′-OH-2,5-DCB resulted in different expression patterns, with the former leading to enrichment of genes involved in response to toxic stress and detoxification functions. Exposure to 2,5-DCB induced multiple xenobiotic response genes, such as cytochrome P-450 and glutathione S-transferases, potentially involved in the PCB metabolism. On the contrary, exposure to both compounds resulted in the down-regulation of genes involved in stresses not directly related to toxicity. Unlike its OH derivative, 2,5-DCB was shown to induce a transcriptomic profile similar to plant safeners, which are nontoxic chemicals stimulating detoxification pathways in plants. The differentiated induction of detoxification enzymes by 2,5-DCB may explain its lower phytotoxicity compared to 4′-OH-2,5-DCB.

Monoclonal antibodies (mAbs) may be prone to self‐association leading to formation of dimers, trimers, or other high molecular weight species during bio‐processing. In order to implement appropriate manufacturing control strategies during bio‐processing, it is important to understand various real life bio‐processing conditions where such self‐associations may manifest. One such case study is presented here of increase in dimer content for an mAb during scale‐up bio‐processing and the approach taken to understand the under‐lying mechanism. In this example, a therapeutic mAb demonstrated a consistently higher dimer values (~0.5% higher) in the drug product (DP) during release when compared to the same value measured in the corresponding drug substance (DS) lot. This observation was interesting since the DS was supplied frozen, and the DS and DP share the same formulation composition and therefore investigation of this dimer change was the scope of the characterization study. Variable path length spectroscopy and size exclusion chromatography was used for protein quantification and to monitor %dimer respectively during characterization of fill‐finish unit operations. At the start of DP manufacturing process, immediately after thaw of bulk DS, a protein concentration gradient was observed and the concentration ranged from 90 mg/mL (top of container) to 210 mg/mL (bottom of container). The dimerization kinetics in the same DS container was dependent on concentration with higher concentrations demonstrating higher rates of dimerization. After the bulk DS was mixed for further processing, %dimer in purified bulk DS was quantitated to be approximately 1.4% which is identical to levels observed during scale‐up manufacturing of DP. After each unit operation, the in‐process samples tested for %dimer showed a gradual increase in dimer as a function of time over the next 7 days accumulating to 1.8% dimer at the end of DP manufacturing process. Samples subjected to static incubation at 2–8°C and room temperature (RT; 15–25°C) showed a gradual increase in dimer over the same time frame; however, the rate of increase in dimer at RT was higher compared to samples stored at 2–8°C. The results from this demonstrate two important key findings: self‐association kinetics of mAbs could be exacerbated by protein cryoconcentration and temperature conditions during bioprocessing. Since these two parameters are commonly encountered during manufacturing, the proposed mitigation strategy is to ensure homogeneity of the bulk DP during processing. The temperature dependent self‐association kinetics of mAb could be mitigated by processing at lower temperature (e.g., 2–8°C) and by storing the finished DP at lower temperature after manufacturing. The results from this study also highlight the criticality of setting slightly wider specifications for DP compared to DS following ICH Q6B guidelines.

Hydroxylated polychlorinated biphenyls (OH-PCBs) are toxic contaminants produced via biotic or abiotic transformation of PCBs. In this study, we have tested the toxicity of 2,5-dichlorobiphenyl (2,5-DCB) and three of its OH-derivatives, 2'-OH-, 3'-OH-, and 4'-OH-2,5-DCB toward the model plant, Arabidopsis thaliana. Toxicity tests showed that the parent 2,5-DCB (5 mg L −1 ) had little effect on the plants, while all three OH-metabolites (5 mg L −1 ) exhibited a significant toxicity, with 4'-OH-2,5-DCB being the most potent (inhibition concentration 50%-IC 50 in germination tests = 9.8 mg L −1 for 2'-OH-2,5-DCB, 9.5 mg L −1 for 3'-OH-2,5-DCB, and 4.8 mg L −1 for 4'-OH-2,5-DCB). Whole-genome expression microarrays (Affymetrix) showed that exposure to the three OH-PCBs resulted in rather similar expression patterns, which were distinct from the one developing in response to 2,5-DCB. Searching an Arabidopsis microarray database (Genevestigator) revealed that, unlike the parent compound, the three OH-derivatives induced expression profiles similar to inhibitors of brassinosteroid synthesis (i.e., brassinazole, propiconazole, and uniconazole), resulting in severe iron deficiency in exposed plants. Our results suggest that the higher phytotoxicity of OH-derivatives as compared to 2,5-DCB is at least partly explained by the inhibition of the brassinosteroid pathway.

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