Reducing the antigen-independent toxicity of antibody-drug conjugates by minimizing their non-specific clearance through PEGylation.

Recently, we described a family of non-targeting monomethylauristatin E (MMAE) antibody-drug conjugates (ADCs) whose pharmacokinetics could be tuned through incorporation of a short polyethylene glycol (PEG) moiety of up to twelve units into a drug-linker to render the ADC surface more hydrophilic. That work demonstrated that more hydrophilic ADCs were simultaneously more effective and better tolerated in mouse models, suggesting an improvement in therapeutic index via this strategy. Here, we describe the biodistribution and toxicology assessments in Sprague-Dawley rats after intravenous dosing with the aim of elucidating the relationships between these biological outcomes and the underlying physicochemical properties of non-targeted ADCs. Dosing a non-PEGylated ADC exhibited rapid nonspecific cellular uptake, leading to ADC catabolism and rapid release of the cytotoxic payload which reached peak plasma and tissue concentrations within the first day. Introduction of a PEG chain of four, eight, or twelve units resulted in increasingly slower uptake and decreases in peak payload concentrations in all tissues. These ADCs with minimal non-specific uptake also exhibited substantially less hematologic toxicity, with reduced histologic depletion of bone marrow and less dramatic decreases and/or more rapid recovery in peripheral hematologic cell counts (neutrophils, platelets, and reticulocytes). These results support a strong correlation between ADC hydrophobicity, rate of non-specific uptake, peak tissue concentration of released payload, and resulting toxicology parameters. Should these correlations be translatable to the clinic, this would provide a more general and highly tractable strategy for reducing the antigen-independent toxicity of ADCs through drug-linker design to modulate non-specific biodistribution.

Targeted Delivery of Cytotoxic NAMPT Inhibitors Using Antibody-Drug Conjugates.

Antibody-drug conjugates (ADCs) are a therapeutic modality that enables the targeted delivery of cytotoxic drugs to cancer cells. Identification of active payloads with unique mechanisms of action is a key aim of research efforts in the field. Herein, we report the development of inhibitors of nicotinamide phosphoribosyltransferase (NAMPT) as a novel payload for ADC technology. NAMPT is a component of a salvage biosynthetic pathway for NAD, and inhibition of this enzyme results in disruption of primary cellular metabolism leading to cell death. Through derivatization of the prototypical NAMPT inhibitor FK-866, we identified potent analogues with chemical functionality that enables the synthesis of hydrophilic enzyme-cleavable drug linkers. The resulting ADCs displayed NAD depletion in both cell-based assays and tumor xenografts. Antitumor efficacy is demonstrated in five mouse xenograft models using ADCs directed to indication-specific antigens. In rat toxicology models, a nonbinding control ADC was tolerated at >10-fold the typical efficacious dose used in xenografts. Moderate, reversible hematologic effects were observed with ADCs in rats, but there was no evidence for the retinal and cardiac toxicities reported for small-molecule inhibitors. These findings introduce NAMPT inhibitors as active and well-tolerated payloads for ADCs with promise to improve the therapeutic window of NAMPT inhibition and enable application in clinical settings.

Parental and larval exposure to nicotine modulate spontaneous activity as well as cholinergic and GABA receptor expression in adult C. elegans.

Early nicotine exposure has been associated with many long-term consequences that include neuroanatomical alterations, as well as behavioral and cognitive deficits. To describe the effects of early nicotine exposure in Caenorhabditis elegans, the current study observed spontaneous locomotor activity (i.e., reversals) either in the presence or absence of nicotine. Expression of acr-16 (a nicotinic receptor subunit) and a ß-like GABA(A) receptor subunit, gab-1, were also examined with RT-PCR. Worms were exposed to nicotine (30 µM) throughout "zygote formation" (period that includes oocyte maturation, ovulation and fertilization), from hatching to adulthood ("larval development") or across both zygote and larval development. Adult larval-exposed worms only showed an increase in spontaneous behavior when tested on nicotine (p<0.001) but levels of activity similar to controls when tested on plain plates (p>0.30). Larval-exposed worms also showed control levels of acr-16 nicotinic receptor expression (p>0.10) but increased gab-1 expression relative to controls (p<0.01). In contrast, zygote-exposed and zygote- plus larval-exposed worms showed a similar increase in spontaneous behavior on plain plates (p<0.001 and p=0.001, respectively) but control levels of responding when tested on nicotine (p>0.90 for each). However, expression of acr-16 and gab-1 was downregulated in zygote-exposed (p<0.01 and p<0.05, respectively) and significantly upregulated in the zygote- plus larval-exposed worms (p<0.000 for each); most surprising was the over five-fold increase in gab-1 expression. These results suggest that spontaneous motor behavior and receptor expression are differentially modulated by nicotine exposure during larval development and/or zygote formation. As well, these findings demonstrate that C. elegans, as a model system, is also sensitive to nicotine exposure during early development and provides the basis for future research to uncover specific mechanisms by which early nicotine exposure modifies neuronal signaling and alters behavior.