Prophylactic treatment with PEGylated bovine IFNλ3 effectively bridges the gap in vaccine-induced immunity against FMD in cattle.

Foot-and-mouth disease (FMD) is a vesicular disease of cloven-hoofed animals with devastating economic implications. The current FMD vaccine, routinely used in enzootic countries, requires at least 7 days to induce protection. However, FMD vaccination is typically not recommended for use in non-enzootic areas, underscoring the need to develop new fast-acting therapies for FMD control during outbreaks. Interferons (IFNs) are among the immune system's first line of defense against viral infections. Bovine type III IFN delivered by a replication defective adenovirus (Ad) vector has effectively blocked FMD in cattle. However, the limited duration of protection-usually only 1-3 days post-treatment (dpt)-diminishes its utility as a field therapeutic. Here, we test whether polyethylene glycosylation (PEGylation) of recombinant bovine IFNλ3 (PEGboIFNλ3) can extend the duration of IFN-induced prevention of FMDV infection in both vaccinated and unvaccinated cattle. We treated groups of heifers with PEGboIFNλ3 alone or in combination with an adenovirus-based FMD O1Manisa vaccine (Adt-O1M) at either 3 or 5 days prior to challenge with homologous wild type FMDV. We found that pre-treatment with PEGboIFNλ3 was highly effective at preventing clinical FMD when administered at either time point, with or without co-administration of Adt-O1M vaccine. PEGboIFNλ3 protein was detectable systemically for >10 days and antiviral activity for 4 days following administration. Furthermore, in combination with Adt-O1M vaccine, we observed a strong induction of FMDV-specific IFNγ+ T cell response, demonstrating its adjuvanticity when co-administered with a vaccine. Our results demonstrate the promise of this modified IFN as a pre-exposure prophylactic therapy for use in emergency outbreak scenarios.

Analytical comparability to evaluate impact of manufacturing changes of ARX788, an Anti-HER2 ADC in late-stage clinical development.

ARX788 is an anti-HER2 antibody drug conjugate (ADC) developed using Ambrx proprietary Engineered Precision Biologics technology. The manufacturing process of ARX788 has been optimized during the course of early to late-phase clinical development. A comprehensive evaluation of side-by-side comparability between pre- and post-change process for ARX788 drug substance and drug product from a quality perspective was conducted based on ICH Q5E guidelines consisting of batch release assays, physicochemical and biophysical characterization, biological characterization, and forced degradation studies. All results have substantiated a high degree of similarity between the pre- and post-change ARX788 drug substance batches and drug product lots, demonstrating that the process manufacturing changes did not impact product quality.

ARX788, a Site-specific Anti-HER2 Antibody-Drug Conjugate, Demonstrates Potent and Selective Activity in HER2-low and T-DM1-resistant Breast and Gastric Cancers.

First-generation antibody-drug conjugates (ADC) are heterogeneous mixtures that have shown clinical benefit, but generally exhibited safety issues and a narrow therapeutic window due, in part, to off-target toxicity caused by ADC instability. ARX788 is a next-generation, site-specific anti-HER2 ADC that utilizes a unique nonnatural amino acid-enabled conjugation technology and a noncleavable Amberstatin (AS269) drug-linker to generate a homogeneous ADC with a drug-to-antibody ratio of 1.9. ARX788 exhibits high serum stability in mice and a relatively long ADC half-life of 12.5 days. When compared in vitro against T-DM1 across a panel of cancer cell lines, ARX788 showed superior activity in the lower HER2-expressing cell lines and no activity in normal cardiomyocyte cells. Similarly, ARX788 significantly inhibited tumor growth, and generally outperformed T-DM1 in HER2-high and HER2-low expression xenograft models. Breast and gastric cancer patient-derived xenograft studies confirmed strong antitumor activity of ARX788 in HER2-positive and HER2-low expression tumors, as well as in a T-DM1-resistant model. The encouraging preclinical data support the further development of ARX788 for treatment of patients with HER2-positive breast and gastric cancer, including those who have developed T-DM1 resistance, and patients with HER2-low expression tumors who are currently ineligible to receive HER2-targeted therapy.

Nonclinical Development of Next-generation Site-specific HER2-targeting Antibody-drug Conjugate (ARX788) for Breast Cancer Treatment.

Conventional antibody-drug conjugates (ADC) utilize native surface-exposed lysines or cysteines on the antibody of interest to conjugate cytotoxic payload. The nonspecific conjugation results in a mixture with variable drug-to-antibody ratios (DAR), conjugation sites, and ADCs that are often unstable in systemic circulation. ARX788 is an ADC consisting of a HER2-targeting antibody site-specifically conjugated with a potent antitubulin cytotoxic drug-linker, AS269. The site-specific conjugation is achieved by first incorporating the nonnatural amino acid, para-acetyl phenylalanine (pAF), into the antibody, followed by covalent conjugation of AS269 to the pAF to form a highly stable oxime bond resulting in a DAR 2 ADC. ARX788 exhibits significant, dose-dependent antitumor activity against HER2- expressing breast and gastric xenograft tumors. Pharmacokinetic (PK) studies in multiple species showed the highly stable nature of ARX788 with overlapping PK profiles for the intact ADC and total antibody. Metabolism studies demonstrated that pAF-AS269 was the sole major metabolite of ARX788, with no evidence for the release of free drug often observed in conventional ADCs and responsible for adverse side effects. Furthermore, ARX788 demonstrated a favorable safety profile in monkeys with a highest nonseverely toxic dose of 10 mg/kg, which was well above the efficacious dose level observed in preclinical tumor models, thus supporting clinical development of ARX788.

FGF21 analogs of sustained action enabled by orthogonal biosynthesis demonstrate enhanced antidiabetic pharmacology in rodents.

Fibroblast growth factor 21 (FGF21) mitigates many of the pathogenic features of type 2 diabetes, despite a short circulating half-life. PEGylation is a proven approach to prolonging the duration of action while enhancing biophysical solubility and stability. However, in the absence of a specific protein PEGylation site, chemical conjugation is inherently heterogeneous and commonly leads to dramatic loss in bioactivity. This work illustrates a novel means of specific PEGylation, producing FGF21 analogs with high specific activity and salutary biological activities. Using homology modeling and structure-based design, specific sites were chosen in human FGF21 for site-specific PEGylation to ensure that receptor binding regions were preserved. The in vitro activity of the PEGylated FGF21 ana-logs corresponded with the site of PEG placement within the binding model. Site-specific PEGylated analogs demonstrated dramatically increased circulating half-life and enhanced efficacy in db/db mice. Twice-weekly dosing of an optimal FGF21 analog reduced blood glucose, plasma lipids, liver triglycerides, and plasma glucagon and enhanced pancreatic insulin content, islet number, and glucose-dependent insulin secretion. Restoration of insulin sensitivity was demonstrated by the enhanced ability of insulin to induce Akt/protein kinase B phosphorylation in liver, muscle, and adipose tissues. PEGylation of human FGF21 at a specific and preferred site confers superior metabolic pharmacology.