Nonclinical Safety Research Articles

Abstract 5202: AMG 794, a Claudin 6-targeted half-life extended (HLE) bispecific T cell engager (BITE®) molecule for non-small cell lung cancer and epithelial ovarian cancer

Abstract AMG 794 is a half-life extended BiTE® immune therapy targeting the oncofetal antigen Claudin 6 (CLDN6). AMG 794 redirects T cells to kill CLDN6-expressing tumor cells and is being developed for the treatment of non-small cell lung cancer (NSCLC) and epithelial ovarian cancer (EOC). CLDN6 is a compelling tumor antigen that is expressed during embryonic and fetal development, transcriptionally silenced in adult tissues, and re-expressed on the surface of NSCLC and EOC cells. By immunohistochemistry, CLDN6 staining of the cell membrane was observed in 27% of non-squamous NSCLC (n = 63) and 69% of EOC (n = 92) samples, the majority of which were of the high-grade serous ovarian cancer subtype. Expression of CLDN6 protein was not detected in most normal adult tissues, with rare CLDN6 immunostaining limited to individual cells in the pituitary, pancreas, small intestine, kidney, and female reproductive organs. AMG 794 is a fully human BiTE® molecule that binds both human and cynomolgus monkey CLDN6 and CD3. AMG 794 binds human CLDN6 and CD3 with equilibrium dissociation constant (KD) of 13 nM and 36 nM, respectively. In vitro, AMG 794 redirects human T cells to kill CLDN6-expressing cancer cells with a half-maximal lysis concentration (EC50) of 2.6 ± 1.1 pM to 127.4 ± 53.4 pM. Consistent with the mechanism of action of BiTE® immune therapy, AMG 794 induces T cell activation and transient production of cytokines in co-cultures of T cells and CLDN6-expressing tumor cells. Remarkably, AMG 794 binding and cytotoxic activity is selective for CLDN6 over other claudin family proteins, despite high homology in the extracellular loops with CLDN9. Weekly dosing of AMG 794 significantly inhibited the growth of established lung and ovarian xenograft tumors in immunocompromised mice injected with human T cells. Anti-tumor activity was associated with an increase in tumor-infiltrating T cells. AMG 794 was well tolerated in a one-month repeat-dose toxicology study in cynomolgus monkey, with evidence for target engagement. The potent, selective activity of AMG 794 for CLDN6-expressing NSCLC and EOC cells, together with an acceptable nonclinical safety profile, supported the advancement of AMG 794 into clinical development. A first-in-human study to explore the safety, tolerability, pharmacokinetics, and anti-tumor activity of AMG 794 in patients with CLDN6-positive advanced/metastatic non-squamous NSCLC or EOC will be enrolling patients in March 2022. Citation Format: Elizabeth Pham, Anja Henn, Beate Sable, Joachim Wahl, Kip Conner, Katja Matthes, Shivani Gupta, Rodolfo Yabut, Famke Aeffner, Kristin Lewis Wilson, Jonas Anlahr, Christoph Dahlhoff, Vijay Kale, Matthias Friedrich, Tobias Raum, Peter Kufer, Angela Coxon, Sabine Stienen, Julie M. Bailis. AMG 794, a Claudin 6-targeted half-life extended (HLE) bispecific T cell engager (BITE®) molecule for non-small cell lung cancer and epithelial ovarian cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5202.

Society of Toxicologic Pathology Neuropathology Interest Group Article: Neuropathologic Findings in Nonhuman Primates Associated With Administration of Biomolecule-Based Test Articles.

The increasing specificity of novel druggable targets coupled with the complexity of emerging therapeutic modalities for treating human diseases has created a growing need for nonhuman primates (NHPs) as models for translational drug discovery and nonclinical safety assessment. In particular, NHPs are critical for investigating potential unexpected/undesired on-target and off-target liabilities associated with administration of candidate biotherapeutics (nucleic acids, proteins, viral gene therapy vectors, etc.) to treat nervous system disorders. Nervous system findings unique to or overrepresented in NHPs administered biomolecule-based ("biologic") test articles include mononuclear cell infiltration in most neural tissues for all biomolecule classes as well as neuronal necrosis with glial cell proliferation in sensory ganglia for certain viral vectors. Such test article-related findings in NHPs often must be differentiated from procedural effects (e.g., local parenchymal or meningeal reactions associated with an injection site or implanted catheter to administer a test article directly into the central nervous system) or spontaneous background findings (e.g., neuronal autophagy in sensory ganglia).

Changing the drug development and therapeutic paradigm with biologic drug combinations and bispecifics: How to choose between these two approaches?

Biologics are increasingly being co‐developed in combination or as novel constructs like bispecific antibodies (BsAbs) with the goal of targeting multiple, non‐redundant mechanisms of action. Rational design of combinations and dual‐targeting approaches that consider disease complexities have the potential to improve efficacy and safety, to increase duration of clinical benefit, and to minimize clinical resistance mechanisms. Here we summarize examples of BsAbs and biologic combinations that have been approved by health authorities and present drug development considerations when deciding between these two strategies. These include an understanding of target biology, nonclinical safety risks, dose optimization strategies, the regulatory framework, pharmacokinetic, immunogenicity, and bioanalytical assay considerations. The disease biology, target dynamics, and pharmacology objectives were identified as important factors in early drug development to decide between a BsAb versus a combination. Nonclinical safety assessment and dose optimization strategies can also pose challenges for BsAb versus combinations. High unmet medical needs and lack of treatment options are often the common denominators for deciding to develop a BsAb or a combination. Future development of biologic triple combinations and BsAbs combinations with other biologics will further increase drug development complexities and hold promise for more effective treatment options for patients.

Nonclinical Efficacy and Safety of CX-2029, an Anti-CD71 Probody-Drug Conjugate.

Probody therapeutics (Pb-Txs) are conditionally activated antibody-drug conjugates (ADCs) designed to remain inactive until proteolytically activated in the tumor microenvironment, enabling safer targeting of antigens expressed in both tumor and normal tissue. Previous attempts to target CD71, a highly expressed tumor antigen, have failed to establish an acceptable therapeutic window due to widespread normal tissue expression. This study evaluated whether a probody-drug conjugate targeting CD71 can demonstrate a favorable efficacy and tolerability profile in preclinical studies for the treatment of cancer. CX-2029, a Pb-Tx conjugated to maleimido-caproyl-valine-citrulline-p-aminobenzyloxycarbonyl-monomethyl auristatin E, was developed as a novel cancer therapeutic targeting CD71. Preclinical studies were performed to evaluate the efficacy and safety of this anti-CD71 PDC in patient-derived xenograft (PDX) mouse models and cynomolgus monkeys, respectively. CD71 expression was detected at high levels by IHC across a broad range of tumor and normal tissues. In vitro, the masked Pb-Tx form of the anti-CD71 PDC displayed a >50-fold reduced affinity for binding to CD71 on cells compared with protease-activated, unmasked anti-CD71 PDC. Potent in vivo tumor growth inhibition (stasis or regression) was observed in >80% of PDX models (28/34) at 3 or 6 mg/kg. Anti-CD71 PDC remained mostly masked (>80%) in circulation throughout dosing in cynomolgus monkeys at 2, 6, and 12 mg/kg and displayed a 10-fold improvement in tolerability compared with an anti-CD71 ADC, which was lethal. Preclinically, anti-CD71 PDC exhibits a highly efficacious and acceptable safety profile that demonstrates the utility of the Pb-Tx platform to target CD71, an otherwise undruggable target. These data support further clinical development of the anti-CD71 PDC CX-2029 as a novel cancer therapeutic.

A novel next‐generation FVIIIa mimetic, Mim8, has a favorable safety profile and displays potent pharmacodynamic effects: Results from safety studies in cynomolgus monkeys

BackgroundMim8 is a novel, next‐generation factor VIIIa mimetic in development for subcutaneous prophylactic treatment of patients with hemophilia A with and without inhibitors. In vitro and in vivo models indicate that Mim8 has a distinct hemostatic potential. ObjectivesTo test the nonclinical safety and pharmacodynamics of Mim8. MethodsThe Mim8 nonclinical safety program in cynomolgus monkeys consisted of three studies of 4–26 weeks in duration with Mim8 doses ranging from 0.3–60 mg/kg/week intravenously or subcutaneously. After sacrifice, macroscopic and microscopic pathological examinations were performed. ResultsMim8 was well tolerated with no noteworthy clinical observations. No signs of excessive coagulation or pathological macroscopic or microscopic findings were observed at doses 0.3–3 mg/kg/week subcutaneous. Thrombosis‐related findings were detected during histopathological examination in a small proportion of animals (16%) receiving doses ranging 6–20 mg/kg/week. Dose‐dependent increases in factor X (FX) and factor IX (FIX) concentrations were observed. Shortening of activated partial thromboplastin time (APTT) and increased thrombin generation under ex vivo hemophilia A‐like conditions were observed at all Mim8 dose levels. ConclusionsThrombosis‐related findings observed at doses above 6 mg/kg/week Mim8 may have been exaggerated pharmacological reactions to a procoagulant compound in normocoagulant animals. Increases in FX and FIX concentrations could be because of a half‐life prolongation due to binding to Mim8, but were limited at clinically relevant exposure levels. Subcutaneous administration of up to 3 mg/kg/week (several fold greater than expected clinical exposure) for 26 weeks resulted in relevant pharmacodynamic effects, observed in thrombin generation and APTT, with no signs of thrombi or excessive coagulation activation.

A Technical Guide to Sampling the Beagle Dog Nervous System for General Toxicity and Neurotoxicity Studies.

Beagle dogs are a key nonrodent species in nonclinical safety evaluation of new biomedical products. The Society of Toxicologic Pathology (STP) has published "best practices" recommendations for nervous system sampling in nonrodents during general toxicity studies (Toxicol Pathol 41[7]: 1028-1048, 2013), but their adaptation to the Beagle dog has not been defined specifically. Here we provide 2 trimming schemes suitable for evaluating the unique neuroanatomic features of the dog brain in nonclinical toxicity studies. The first scheme is intended for general toxicity studies (Tier 1) to screen test articles with unknown or no anticipated neurotoxic potential; this plan using at least 7 coronal hemisections matches the STP "best practices" recommendations. The second trimming scheme for neurotoxicity studies (Tier 2) uses up to 14 coronal levels to investigate test articles where the brain is a suspected or known target organ. Collection of spinal cord, ganglia (somatic and autonomic), and nerves for dogs during nonclinical studies should follow published STP "best practices" recommendations for sampling the central (Toxicol Pathol 41[7]: 1028-1048, 2013) and peripheral (Toxicol Pathol 46[4]: 372-402, 2018) nervous systems. This technical guide also demonstrates the locations and approaches to collecting uncommonly sampled peripheral nervous system sites.

Does Geographical Origin of Long-Tailed Macaques (Macaca fascicularis) Matter in Drug Safety Assessment?: A Literature Review and Proposed Conclusion.

Long-tailed macaques are the predominant nonhuman primate species for the nonclinical safety testing of biopharmaceuticals. This species comprises 9 subspecies with Macaca fascicularis fascicularis naturally occurring in Southeast Asia. Since the 17th century, M. f. fascicularis also occurs on Mauritius. Cynomolgus macaques do not naturally occur in China, but are bred in many farms across the country. The current shortage in animal supply raises the question whether geographical animal origin matters and if animals from different geographical regions can be combined on a drug development program or even a single experiment. This article reviews geographical animal origin in relation to selected endpoints that are relevant in nonclinical drug safety testing. Animals from different countries within Asia mainland do not appear to show any meaningful difference. Very little data are available for animals from Asia island. Mauritian animals show consistent differences from Asian animals in several clinical and anatomical pathology parameters. For developmental parameters, animals from Mauritius and Asia are comparable with the exception that Mauritian animals mature faster. In the authors' view, differences between the geographical clusters can be accounted for as long as baseline and reference data are available.

Comprehensive Nonclinical Safety Assessment of Nirmatrelvir Supporting Timely Development of the SARS-COV-2 Antiviral Therapeutic, Paxlovid™.

COVID-19 is a potentially fatal infection caused by the SARS-CoV-2 virus. The SARS-CoV-2 3CL protease (Mpro) is a viral enzyme essential for replication and is the target for nirmatrelvir. Paxlovid (nirmatrelvir co-administered with the pharmacokinetic enhancer ritonavir) showed efficacy in COVID-19 patients at high risk of progressing to hospitalization and/or death. Nonclinical safety studies with nirmatrelvir are essential in informing benefit-risk of Paxlovid and were conducted to support clinical development. In vivo safety pharmacology assessments included a nervous system/pulmonary study in rats and a cardiovascular study in telemetered monkeys. Potential toxicities were assessed in repeat dose studies of up to 1 month in rats and monkeys. Nirmatrelvir administration (1,000 mg/kg, p.o.) to male rats produced transient increases in locomotor activity and respiratory rate but did not affect behavioral endpoints in the functional observational battery. Cardiovascular effects in monkeys were limited to transient increases in blood pressure and decreases in heart rate, observed only at the highest dose tested (75 mg/kg per dose b.i.d; p.o.). Nirmatrelvir did not prolong QTc-interval or induce arrhythmias. There were no adverse findings in repeat dose toxicity studies up to 1 month in rats (up to 1,000 mg/kg daily, p.o.) or monkeys (up to 600 mg/kg daily, p.o.). Nonadverse, reversible clinical pathology findings without clinical or microscopic correlates included prolonged coagulation times at ≥60 mg/kg in rats and increases in transaminases at 600 mg/kg in monkeys. The safety pharmacology and nonclinical toxicity profiles of nirmatrelvir support clinical development and use of Paxlovid for treatment of COVID-19.

SDH, a novel diarylheptane compound, is a potential treatment for inflammatory bowel disease by restoring epithelial barrier function

BackgroundThe global prevalence of inflammatory bowel disease (IBD) is increasing, and mucosal healing is the preferred treatment target of IBD. Sodium (aS,9 R)− 3-hydroxy-16,17-dimethoxy-15-oxidotricyclo[12.3.1.12,6]nonadeca-1(18),2,4,6(19),14,16-hexene-9-yl sulfate hydrate (SDH) is a novel diarylheptane compound, which is designed to treat IBD. Hence, we investigated the potent therapeutic activity of SDH against IBD and explored the underlying mechanisms, and determined if SDH is a safe and well-tolerated oral therapeutic for IBD treatment. MethodsWe characterized its therapeutic properties in vitro and in vivo using Caco-2 cell monolayer and dextran sodium sulfate (DSS)- or 2,4,6-trinitro-benzene sulfonic acid (TNBS)-induced colitis models. We conducted nonclinical toxicology and safety pharmacology research, including general toxicity, toxicokinetics, pharmacokinetics, metabolism and plasma protein binding, cardiovascular safety pharmacology, central nervous system safety pharmacology, respiratory safety pharmacology, fertility and early embryonic development toxicity, reverse mutation assay, chromosomal aberration assay and micronucleus test. ResultsThe results showed that SDH promoted expression of tight junction proteins, and protected the integrity and permeability of the epithelial barrier in both cell and animal models. Moreover, lower doses of SDH showed the similar or better efficacy than cyclosporine A (CsA) and mesalazine in DSS- or TNBS-induced colitis animals. Furthermore, our results identified that SDH has satisfactory safety in these studies we tested. In summary, SDH restored the epithelial barrier through tight junction proteins and was expected to be a novel therapeutic agent for the treatment of IBD.

Selected Resources for Pathology Evaluation of Nonhuman Primates in Nonclinical Safety Assessment.

Humans and nonhuman primates (NHPs) share numerous anatomical and physiological characteristics, thereby explaining the importance of NHPs as essential animal models for translational medicine and nonclinical toxicity testing. Researchers, toxicologic pathologists, toxicologists, and regulatory reviewers must be familiar with normal and abnormal NHP biological traits when designing, performing, and interpreting data sets from NHP studies. The current compilation presents a list of essential books, journal articles, and websites that provide context to safety assessment and research scientists working with NHP models. The resources used most frequently by the authors have been briefly annotated to permit readers to rapidly ascertain their applicability to particular research endeavors. The references are aimed primarily for toxicologic pathologists working with cynomolgus and rhesus macaques and common marmosets in efficacy and safety assessment studies.

Distortion Product Otoacoustic Emission Test is Not the Test to Use in Nonclinical Safety Assessment.

Ototoxicity and ocular toxicity screening are but two examples of specialty product lines that are often employed as Tier II or III nonclinical safety/hazard screening assessments. Compared to the regulatory guidelines that govern over standard toxicology or neurotoxicology programs, there is a paucity of regulatory strategies to address these specialized product lines. With respect to ototoxicity testing, we argue for the inclusion of the "least burdensome principles" adopted by the US FDA in providing the most pragmatic, efficient, and directed identification of potential harm to auditory function in the nonclinical safety arena. We argue for the exclusive use of the auditory brainstem response and the exclusion of the distortion product otoacoustic emissions (DPOAEs) in these Tiered II safety assessment programs. The inclusion of both are a burden on operational staff and, due to the extended episodes of anesthesia required to conduct both assays, this strategy poses a health and welfare concern for the selected animal species to be used. The DPOAE does not provide any sufficiently valid or reliable data above and beyond the gold standard ABR data, followed by complete oto-histopathology and cytocochleogram combination designs.

Investigational New Drug Enabling Nonclinical Safety Assessment of the Iminosugar UV-4, a Broad-Spectrum Host-Targeted Antiviral Agent.

The iminosugar UV-4 is a broad-spectrum antiviral drug candidate with activity in vitro and in vivo against multiple, diverse viruses. The toxicological profile of UV-4, dosed as the hydrochloride salt, was evaluated in single-dose and repeat-dose oral toxicity studies in mice, rats, dogs, and non-human primates (NHP). No moribundity or deaths were associated with the drug up to the maximum tolerated dose. No treatment-related adverse effects were observed following single oral doses in dogs, rats, and mice up to 250, 400, 1000mg/kg, respectively, and in NHP up to 180mg/kg administered three times daily for 10days. UV-4-related findings were generally seen at higher doses after 7- or 14-day exposure. The most common clinical pathology findings (increase in aspartate aminotransferase and decreased platelet count) were consistently found across species and each appeared dose related. The kidney, mesenteric lymph nodes, stomach including gastrointestinal tract, and thymus were identified as target organs in mice, rats, and dogs. In 14-day repeat-dose toxicology studies in mice and dogs conducted in compliance with Good Laboratory Practice regulations, the dog was considered to be the most sensitive species to UV-4 exposure based on the treatment-related adverse effects noted in the identified target organs. The results of these studies demonstrate the safety profile of UV-4 hydrochloride and supported the selection of starting and maximal doses for a single ascending dose first-in-human clinical study.

Characteristics of Extracellular Vesicles and Preclinical Testing Considerations Prior to Clinical Applications.

Cell therapy products have significant limitations, such as storage instability, difficulties with transportation, and toxicity issues such as tumorigenicity and immunogenicity. Extracellular vesicles (EVs) secreted from cells show potential for therapeutic agent development. EVs have not been widely examined as investigational drugs, and non-clinical studies for the clinical approval of EV therapeutic agents are challenging. EVs contain various materials, such as DNA, cellular RNA, cytokines, chemokines, and microRNAs, but do not proliferate or divide like cells, thus avoiding safety concerns related to tumorigenicity. However, the constituents of EVs may induce the proliferation of normal cells; therefore, the suitability of vesicles should be verified through non-clinical safety evaluations. In this review, the findings of non-clinical studies on EVs are summarized. We describe non-clinical toxicity studies of EVs, which should be useful for researchers who aim to develop these vesicles into therapeutic agents. A new method for evaluating the immunotoxicity and tumorigenicity of EVs should also be developed.

Nonclinical cardiovascular safety assessment of thioridazine: Impact of autonomic tone, body temperature, and choice of species

Pending updates to ICH S7B/E14 guidelines will enable the substitution of human TQT studies with support from concomitant negative hERG and non-rodent CV studies. This retrospective analysis compared the effects of thioridazine (THD) (5–20 mg/kg) on heart rate (HR), blood pressure (BP), body temperature (Tc), and QT in the dog (n = 6), cynomolgus monkey (n = 4), and Goettingen minipig (n = 4) with data from previously completed studies employing crossover designs. As QT measurements are confounded by HR and Tc changes, QT effects were individually corrected for changes in HR (QTca) and Tc (QTcaT). THD-induced hemodynamic changes seen in humans were most accurately reflected in the monkey and, to a lesser extent, the dog, but not in the minipig. The minipig was most sensitive to THD QTc effects. When QTca was adjusted for THD-associated Tc decreases in minipigs and monkeys, the minipig revealed a lessened but pronounced QTcaT increase (48 ms). In the monkey, a persistent QTca increase was reduced to only a transient (0.5–3 h) QTcaT increase (20 ms). The dog's lack of THD QTca effects triggered co-administration of atenolol (AT) to attenuate THD-induced HR increases in the dog and monkey. THD + AT revealed peak QTcaT increases of 32 ms in the dog and 40 ms in the monkey, suggesting potential autonomic nervous system (ANS) interference in detecting repolarization changes. These results highlight critical species-specific differences in the outcome of parallel safety investigations. Species selection for nonclinical safety studies should consider the potential impact of Tc and ANS effects to avoid false-negative or overly positive outcomes.

Spontaneous Findings in the Reproductive System of Sexually Mature Male Cynomolgus Macaques.

Sexually mature nonhuman primates are often used in nonclinical safety testing when evaluating biopharmaceuticals; however, there is limited information in historical control databases or in the published literature on the spontaneous findings in the male reproductive system. This review evaluated digital slides from the male reproductive tract (testes, epididymides, prostate, and seminal vesicles) in sexually mature cynomolgus macaques (Macaca fascicularis; n = 255) from vehicle control groups in nonclinical toxicology studies and compared the observations with body weight, organ weight, and geographical origin. The most common microscopic findings were hypospermatogenesis and tubular dilatation in the testes; inflammatory cell infiltrate, cellular debris, and decreased sperm in the epididymides; inflammatory cell infiltrate and acinar dilatation in the prostate; and corpora amylacea and atrophy in the seminal vesicles. There were a few correlative observations in animals when grouped by weight or geographical origin: animals with lower terminal body weights (<5 kg) often displayed features of late puberty despite having sperm in the epididymis, while animals originating from Mauritius had a lower incidence of inflammatory cell infiltrates than those from Southeast Asia/China. This review provides incidence, descriptions, and photomicrographs of the common spontaneous microscopic findings in the reproductive system of mature male cynomolgus macaques.

Practical Considerations in Determining Adversity and the No-Observed-Adverse-Effect-Level (NOAEL) in Nonclinical Safety Studies: Challenges, Perspectives and Case Studies.

Determining the adverse nature of findings from nonclinical safety studies often poses a challenge for the key stakeholders responsible for interpreting the results of definitive toxicity studies in support of pharmaceutical product development. Although there are instances in which responses to treatment clearly indicate intolerability or tissue injury associated with dysfunction; in practice, more often there is uncertainty in characterizing an effect of drug treatment as adverse or not. This is due to the inherent variability in responses of biological test systems to toxicological insults, leaving the ultimate analyses of adversity to individual interpretation and subjectivity. This article is a follow-up to the workshop entitled, "Adverse or Not Adverse?: Thinking process behind adversity determination during nonclinical drug development," conducted at the 58th Annual Meeting of the Society of Toxicology, March 2019 in Baltimore, MD. In this paper, we further discuss and incorporate the perspectives of authors representing different roles, such as Study Director, Study Pathologist, Pharmacology/Toxicology Reviewer (U.S. Food and Drug Administration), and Sponsor in the determination and use of adversity. We also present a practical stepwise approach as an aid in this assessment, and further apply these principles to discuss 10 case studies with different therapeutic modalities and unique challenges.

Threshold for Anaphylactoid Reaction to Polysorbate 80 in Canines.

Polysorbate 80 (PS80) functions as a dispersing agent or solubilizer in many pharmaceuticals, and as a stabilizer in biopharmaceuticals. Topical or parenteral administration of low doses of PS80 in biopharmaceuticals has been associated with mild allergic reactions, including local injection site reactions in humans. High doses of PS80, such as levels found in traditional Chinese herbal parenteral medicines, have been linked to systemic effects consistent with anaphylactoid-type reactions, which are characterized by the direct release of histamine from mast cells (degranulation). Nonclinical safety assessments of PS80 in vivo have mainly focused on canine model systems, a species established to be particularly sensitive to PS80. However, there is conflicting data about the dose and route of administration of PS80 required to elicit an anaphylactoid-type reaction in this model system. Therefore, studies using multiple dosing regimens in anesthetized and conscious dogs including a combination of cardiovascular data, clinical signs, and biomarkers of mast cell degranulation were conducted. An intravenous (IV) bolus of 1mg/kg PS80 (0.25% w/v) elicited a positive anaphylactoid reaction including increased heart rate, hypotension, and clinical signs associated with anaphylactoid reactions (e.g., reddened muzzle). However, a full reaction was not observed with a subcutaneous (SC) injection of PS80 (0.25% w/v) up to 20mg/kg and IV bolus or IV infusions up to 0.5mg/kg. These data establish a threshold dose for eliciting an anaphylactoid reaction in canine which varies depending on the route of administration as well as the rate of PS80 infusion.

Investigational New Drug Enabling Nonclinical Safety Pharmacology Assessment of the Iminosugar UV-4, a Broad-Spectrum Host-Targeted Antiviral Agent.

UV-4 (N-(9-methoxynonyl)-1-deoxynojirimycin) is a broad-spectrum antiviral drug candidate with demonstrated activity in vitro and in vivo against multiple, diverse viruses. Nonclinical safety pharmacology studies were conducted to support the filing of an Investigational New Drug (IND) application. Preliminary in vitro pharmacology testing evaluating potential for binding to "off-target" receptors and enzymes indicated no significant liability for advanced development of UV-4. The safety pharmacology of UV-4 was evaluated in the in vitro human ether-à-go-go-related gene (hERG) assay, in a central nervous system (CNS) study in the mouse (modified Irwin test), in a respiratory safety study in conscious mice using whole body plethysmography, and in a cardiovascular safety study in conscious, radiotelemetry-instrumented beagle dogs. There were no observed adverse treatment-related effects following administration of UV-4 as the hydrochloride salt in the hERG potassium channel assay, on respiratory function, in the CNS study, or in the cardiovascular assessment. Treatment-related cardiovascular effect of decreased arterial pulse pressure after 50 or 200mg of UV-4/kg was the only change outside the normal range, and all hemodynamic parameters returned to control levels by the end of the telemetry recording period. These nonclinical safety pharmacology assessments support the evaluation of this host-targeted broad-spectrum antiviral drug candidate in clinical studies.

28-day repeated dose toxicological evaluation of Coronil in Sprague Dawley rats: Behavioral, hematological, biochemical and histopathological assessments under GLP compliance

Ayurvedic medicines are widely employed globally for prophylaxis and treatment of a variety of diseases. Coronil is a tri-herbal medicine, constituted with the traditional herbs, Tinospora cordifolia, Withania somnifera and Ocimum sanctum, with known immunomodulatory activities. Based on its proven in-vitro activity and in-vivo efficacy, Coronil has been approved as a ‘Supporting Measure for COVID-19’ by the Ministry of AYUSH, Government of India. The current study was aimed to assess the non-clinical safety of Coronil in a 28-day repeated dose toxicity study along with a 14-day recovery period in Sprague Dawley rats. This toxicity study was conducted in accordance with OECD test guideline 407, under GLP-compliance. Specific-Pathogen-Free animals of either sex, housed in Individually-Ventilated-Cages were particularly used in the study. The tested Coronil dose levels were 0, 100, 300 and 1000 mg/kg/day, orally administered to 5 males and 5 female rats per test group. In the current study, no mortality was observed in any group and in addition, Coronil did not elicit any finding of toxicological relevance with respect to clinical signs, ocular effects, hematology, urinalysis and clinical chemistry parameters, as well as macro- or microscopical changes in any organs, when compared to the control group. Accordingly, the No-Observed-Adverse-Effect-Level (NOAEL) of Coronil was ascertained to be 1000 mg/kg/day, subsequent to its 28-day oral administration to male and female rats. The acceptable safety profile of Coronil paves the way further toxicity assessments in rodents for a longer duration as well as in higher animals, and towards its clinical investigation.

Gut-Selective Design of Orally Administered Izencitinib (TD-1473) Limits Systemic Exposure and Effects of Janus Kinase Inhibition in Nonclinical Species.

Izencitinib (TD-1473), an oral, gut-selective pan-Janus kinase (JAK) inhibitor under investigation for treatment of inflammatory bowel diseases, was designed for optimal efficacy in the gastrointestinal tract while minimizing systemic exposures and JAK-related safety findings. The nonclinical safety of izencitinib was evaluated in rat and dog repeat-dose and rat and rabbit reproductive and developmental toxicity studies. Systemic exposures were compared with JAK inhibitory potency to determine effects at or above pharmacologic plasma concentrations (≥1× plasma average plasma concentration [Cave]:JAK 50% inhibitory concentration [IC50] ratio). In rats and dogs, 1000 and 30 mg/kg/day izencitinib, respectively, produced minimal systemic findings (ie, red/white cell changes) and low systemic concentrations (approximately 1× plasma Cave:JAK IC50 ratio) with an 8× nonclinical:clinical systemic area under the curve (AUC) margin compared with exposures at the highest clinically tested dose (300 mg, quaque die, once daily, phase 1 study in healthy volunteers). In dogs, it was possible to attain sufficient systemic exposures to result in immunosuppression characteristic of systemic JAK inhibition, but at high AUC margins (43×) compared with systemic exposures observed at the highest tested dose in humans. No adverse findings were observed in the gastrointestinal tract or systemic tissues. Izencitinib did not affect male or female fertility. Izencitinib did not affect embryonic development in rats and rabbits as commonly reported with systemic JAK inhibition, consistent with low maternal systemic concentrations (2–6× plasma Cave:JAK IC50 ratio, 10–33× nonclinical:clinical AUC margin) and negligible fetal exposures. In conclusion, the izencitinib gut-selective approach resulted in minimal systemic findings in nonclinical species at pharmacologic, clinically relevant systemic exposures, highlighting the impact of organ-selectivity in reducing systemic safety findings.