Test Animals Research Articles

Acute toxicity test of tablets containing α-mangosteen, piperine, curcumin, methyl cinnamate and vitamin C in female wistar rats

Acute toxicity test of tablets containing α-mangosteen, piperine, curcumin, methyl cinnamate and vitamin C in female wistar rats

Сравнение радиопротекторных свойств рибоксина (инозина) и индралина при профилактическом введении в дозировках 100 мг/кг по критерию выживаемости облучённых мышей

Relevance: Due to the high chemical toxicity of all known effective radioprotectors, studies of the radioprotective properties of safer drugs are very relevant. A sufficient number of works are devoted to the radioprotective properties of ribonucleoside riboxin (inosine). However, studies comparing the direct radioprotective properties of riboxin and a recognized radioprotector, for example, indralin, using a survival test in irradiated animals have not yet been carried out. Purpose: Conduct a comparative assessment of the radioprotective properties of riboxin and indralin using a survival test in mice exposed to external X-ray radiation. Material and methods: The experiment was carried out on 200 male ICR (CD-1) mice of the SPF category in duplicate. In each experiment, the animals were divided into the following groups, separated by body weight, 10 animals each: vivar control, not exposed to drugs and radiation, radiation control, with preliminary intraperitoneal administration of sterile water and exposed to external X-ray radiation in doses of 6.0, 6.5 and 6.75 Gy, experimental groups exposed to irradiation in the indicated doses with preliminary intraperitoneal administration of riboxin at a dosage of 100 ml/kg body weight or indralin at a dosage of 100 ml/kg. Survival was assessed for 30 days after irradiation. The dose change factor was determined using probit analysis as the ratio of the radiation dose causing the death of half of the irradiated animals that received the drug to the radiation dose causing the death of half of the irradiated animals without administration of the drug. Results: The use of indralin before X-ray irradiation in doses of 6.0 Gy, 6.5 Gy and 6.75 Gy led to a statistically significant increase in the survival of animals compared to the group receiving Riboxin and control irradiation (р<0,05, log-rank test). Using equations derived from Phinney probit analysis, LD50 doses were calculated for indralin and riboxin, from which dose change factors were calculated to be 1.8 and 1.07, respectively. Conclusion: Since riboxin has not demonstrated radioprotective properties, its preventive use with intraperitoneal administration under the conditions described in this paper, for leveling the effects of radiation can be considered ineffective.

Course on Breeding, Handling, and Experimental Procedures of Zebrafish (Danio rerio): Training, Refinement, Paradigm Shift, and Dissemination of Knowledge from Brazil to the World.

Currently, in Brazil, all researchers involved in animal experimentation must undergo training in laboratory animal science to stay updated on biology, methodology, ethics, and legal considerations related to the use of animals. The training program presented in this study not only aims to fulfill a legal obligation but also intends to train students and professionals to effectively care for their biomodels. It seeks to help them understand the importance of this care, both for the welfare of the animals and for the results of their projects. In total, 58 participants were present at the event (pre-event and full-time course). These participants consisted students and professionals from 11 institutions and 5 different countries. These numbers demonstrate the successful attainment of the desired capillarity in the scientific community and the posterior dissemination of knowledge. Through this course, it was possible to train the participants and raise their awareness about the importance of applying scientific knowledge in their daily practices to maintain the animals, ensuring the welfare of the models and refining the research. Finally, the program presented in this study, as well as the strategies adopted, can serve as a model for other institutions aiming to achieve similar results.

The Effect of Pesticides on Public Health: A Review

Due to the widespread use of pesticides in agriculture and other contexts, human exposure to them persists. Despite premarket animal testing, epidemiologic studies show that current exposures are linked to dangers to the well-being of people. Within this analysis, we outline the current pathways for pesticide exposure and provide an overview and assessment of the epidemiologic research on adult neurotoxicity and carcinogenicity associated with pesticides. Assessing the hazards that pesticides bring to human health will be improved by a deeper comprehension of exposure patterns, the underlying diversity among people, and the connections between animals' toxicological data and their effects on human health. Public health policymakers will be able to evaluate the risks of exposure to pesticides to human health more precisely if epidemiological studies are improved and combined with toxicology data.

Energy metabolism disturbance, altered neuronal development and glutamatergic signalling in human derived neuronal cell models of ADHD

Abstract Despite major advances in research into the neurobiological basis of mental illness, there have been hardly any new developments in new drug therapies. As there are approximately 30% of affected individuals that do not respond sufficiently to available treatments, there is a significant unmet medical need for new therapeutic approaches. About 90% of novel substances that have shown promise in animal studies are not effective in clinical trials. Recent research on human induced pluripotent stem cells (hiPSC) could lead to the use of more human-tailored models in this field. IPSC-derived cell models and organoids may be very attractive for preclinical screening and bridge the gap between in vitro and in vivo studies, reducing animal testing. However, the next steps must first demonstrate the validity and reproducibility of the initial functional results from the hIPSC models of mental illness. In our own studies on neuronal cell models of patients with attention-deficit/hyperactivity disorder (ADHD) with rare PARK2 gene variants, we were able to show evidence of mitochondrial dysfunction and impaired energy metabolism. Additionally, we have first hints at a oxidative dysbalance which could be as well targeted by medication. In a model of cortical development of ADHD patients with common variants in the ADGRL3 gene, we found first evidence for altered neuronal maturation as well as abnormalities in calcium metabolism and glutamatergic functionality compared to cells from healthy controls. In summary, these first results are promising that hIPSC models can contribute new insights into cellular pathomechanisms of mental and neurodevelopmental disorders and the development of new, individualised therapeutic approaches.Disclosure of InterestS. Kittel-Schneider Grant / Research support from: The studies are funded by IZKF Wuerzburg and BBRF fund (to SKS and RMcN). SKS received speaker’s honoraria from Takeda, Medice and Janssen.

63 Allogeneic Recellularized Lung Orthotopic (ARLO) Transplant Research

OBJECTIVES/GOALS: As of 2021, the lung transplantation waiting list has a mortality rate of 7.6 deaths per 100 patient-years. Bioengineered human organs is an emerging field of tissue engineering with a goal of developing suitable organs for transplantation. The focus of the project is to evaluate the efficacy of bioengineered lungs using a human-to-swine model. METHODS/STUDY POPULATION: This project will involve designing and assessing the bioengineered lung by establishing a human-to-pig xenotransplantation survival model. The project aims to evaluate how well the bioengineered lung functions within a living model. The bioengineered lung is constructed using swine connective tissue scaffolding, which has been recellularized with human cells. Anatomically, the lung will resemble a swine lung but will possess the immunological and cellular markers of human tissue. The proposed model will initially assess the immunological response of swine to human lung tissue. Lung function will be assessed during surgery using pulmonary vein gas samples and tissue sampling. Following the end of the study, additional tissues samples will be taken to evaluate the immunological response to the tissue. RESULTS/ANTICIPATED RESULTS: Xenotransplantation and bioengineered organs are two new emerging fields of research that have just begun to enter the large animal testing phase. This model will provide a novel human-to-pig xenotransplant survival model that will be used to test the efficacy of bioengineered lungs function in a dynamic living organism. The design has taken the principles of immunology learned from the current clinical and xenotransplant research and has incorporated this knowledge into the known pig-to-pig transplant models. We anticipate that this model design will be easily reproducible and can be expanded to other bioengineered organs as an effective means to test functionality. DISCUSSION/SIGNIFICANCE: The COVID-19 pandemic’s aftermath may lead to an increased demand for lung transplants. Bioengineered lungs could provide an additional source of organs to supplement current availability. This novel approach has the potential to offer a means to test several different types of bioengineered organs in the future.

오가노이드를 이용한 동물대체시험법 표준동향과 전망

Animal testing for toxicity assessments has long been a point of contention due to concerns over animal welfare and ethics. As an integral component of product development and consumer safety, the evaluation of chemical toxicity is vital. Yet, the 21st century has witnessed a growing demand for alternative testing methods that can offer sustainable solutions without relying on animal experiments. This need has spurred major developed countries and international standardization organizations to pursue the development and standardization of novel testing approaches. Among these, organoid-based methods have emerged as a promising avenue for innovation.
 This study examines the concept of organoids, defining and categorizing alternative testing methods. It explores a broad spectrum of topics, including the standardization trends of alternative testing method regulations, organoid-based method standardization, and trends led by government entities, research institutions, academia, and corporations. It also examines the development of organ-specific manufacturing protocols for organoids. The research finds that international efforts to standardize alternative testing methods are in progress, with a particular focus on organoid-based strategies.
 The anticipated growth and evolution of such alternatives could revolutionize standard practices in safety assessments and drug efficacy evaluations. The standardization of organoid-based testing methods is poised to become a crucial milestone in advancing human health and environmental protection, driving the sector's growth and innovation. Moreover, advancements in organoid technology and the creation of models for various organs are likely to further refine alternative testing methods and introduce new evaluation techniques for drugs and chemicals. This study highlights to the biomedical community that standardizing new testing methods can address the ethical dilemmas of animal testing, underscoring a significant contribution towards ethical scientific practices.

Establishing porcine jejunum-derived intestinal organoids to study the function of intestinal epithelium as an alternative for animal testing

Establishing porcine jejunum-derived intestinal organoids to study the function of intestinal epithelium as an alternative for animal testing

Anti-obesity, phytochemical profiling and acute toxicity study of ethanolic extract of saffron (Crocus sativus L.)

IntroductionCrocus sativus (C. sativus), known as saffron, has been recognized for its medicinal benefits since ancient times. The plant was first introduced to the Chongming Island region of Shanghai, China, in 1980. Chinese Saffron, derived from this plant, is one of China's most valuable traditional herbs, yielding about 1 ton in 2013. Traditional Chinese Medicine (TCM) has used this plant as an antianginal for a long time. The plant has been extensively studied for its pharmacological benefits in treating obesity and gastrointestinal problems. This study aimed to conduct a comprehensive phytochemical analysis of C. sativus stigma (saffron) ethanolic extract using LCMS/MS, quantify its major compound, i.e., trans-4-GG-crocin, and study its acute toxicity. In addition, this study aimed to investigate the effect of the extract on anti-obesity in a high-fat diet-induced zebrafish model. MethodsBefore the anti-obesity study, we conducted a comprehensive profiling of saffron ethanolic extract using UHPLC-ESI-MS/MS in both positive and negative modes. Then, the principal constituent, i.e., trans-4-GG-crocin, was quantified using HPLC with a UV detector. The study was followed by an acute toxicity study for 72 h to evaluate the safety profile of this extract using five concentrations of extract, from 93.75–1500 mg/L. Six groups of wild-type zebrafish were then segregated according to food regimen, i.e., (1) control (5 mg artemia/fish/day), (2) overfeeding (60 mg of artemia/fish/day), (3–6) high-fat diet (5 mg artemia+30 mg egg yolk/fish/day), then starting week 9–16, groups 4–6 were received (4) 50 mg/L, (5) 100 mg/L and (6) 200 mg/L of saffron extract. ResultsNegative mode analysis revealed twenty phytoconstituents, while twenty-two were found in positive mode LCMS/MS analysis. The extract contained 536.91 µg/g of trans-4-GG-crocin. The highest concentration tested in the acute toxicity killed 100% of the testing animals within 24 h, and probit analysis showed that the LC50 of the extract was 1021 mg/L. Eight weeks of treatment with 100 and 200 mg/L of saffron significantly reduced the body mass index of the high-fat diet-induced zebrafish model (p<0.0001). ConclusionSaffron ethanolic extract contains mainly flavonoid, carotenoid, and terpenoid compounds. This extract is highly toxic in high concentrations (1500 mg/L), which kills 100% of the tested zebrafish. Eight weeks of treatment with 100 and 200 mg/L of saffron extract via immersion significantly reduced body mass index in the high-fat diet zebrafish model. The findings suggest that Saffron extract could be a promising natural anti-obesity agent that warrants further investigation. The study provides valuable insights into the potential of natural compounds as a source of anti-obesity agents and highlights the need for further research in this area. Even though this plant was studied for anti-obesity in rodent models, its mode of action is still unknown and warrants further investigation, which could be explored more in a cheaper and easier way by using another translational animal model, such as zebrafish.

Emotion Detection in Text: Advances in Sentiment Analysis Using Deep Learning

In the modern era of digital communication, the analysis of sentiment has emerged as a crucial tool for understanding and inferring public sentiment as communicated through written text. This is particularly relevant in the context of social media platforms such as Twitter, Facebook and Instagram. The present study focuses on the urgent matter of public opinion regarding the practice of animal testing, employing advanced deep-learning methodologies for sentiment analysis. A dataset of 15,360 tweets about animal testing was collected using the Twitter API. The data was prepared for analysis by undergoing careful preprocessing and word embedding it through the utilization of Word2vec. To classify tweets into positive and negative sentiment categories, a Long Short-Term Memory (LSTM) model was employed, given its suitability for processing sequential data. Remarkably, an accuracy rate of 88.7 percent was achieved by the model. It was determined that around 80% of tweets expressed criticism towards animal testing, indicating the presence of a substantial negative sentiment majority. These results show the topic's continuing significance by emphasizing its highly emotional and controversial nature. It is concluded that deep learning, and in particular LSTM models, can be used to effectively analyze large amounts of social media data and yield insightful understandings of public opinion. This study underlines the significance of sentiment analysis for gaining insight into public opinion and for its applications in policymaking and discourse analysis.

Pluripotent stem cells for target organ developmental toxicity testing.

Prenatal developmental toxicity research focuses on understanding the potential adverse effects of environmental agents, drugs, and chemicals on the development of embryos and fetuses. Traditional methods involve animal testing, but ethical concerns and the need for human-relevant models have prompted the exploration of alternatives. Pluripotent stem cells (PSCs) are versatile cells with the unique ability to differentiate into any cell type, serving as a foundational tool for studying human development. Two-dimensional (2D) PSC models are often chosen for their ease of use and reproducibility for high-throughput screening. However, they lack the complexity of an in vivo environment. Alternatively, three-dimensional (3D) PSC models, such as organoids, offer tissue architecture and intercellular communication more reminiscent of in vivo conditions. However, they are complicated to produce and analyze, usually requiring advanced and expensive techniques. This review discusses recent advances in the use of human PSCs differentiated into brain and heart lineages and emerging tools and methods that can be combined with PSCs to help address important scientific questions in the area of developmental toxicology. These advancements and new approach methods align with the push for more relevant and predictive developmental toxicity assessment, combining innovative techniques with organoid models to advance regulatory decision-making.

Exploring the therapeutic potential of DV-B-120 as an inhibitor of dengue virus infection.

Dengue fever, an infectious disease prevalent in subtropical and tropical regions, currently lacks effective small-molecule drugs as treatment. In this study, we used a fluorescence peptide cleavage assay to screen seven compounds to assess their inhibition of the dengue virus (DENV) NS2B-NS3 protease. DV-B-120 demonstrated superior inhibition of NS2B-NS3 protease activity and lower toxicity compared to ARDP0006. The selectivity index of DV-B-120 was higher than that of ARDP0006. In vivo assessments of the antiviral efficacy of DV-B-120 against DENV replication demonstrated delayed mortality of suckling mice treated with the compound, with 60-80% protection against life-threatening effects, compared to the outcomes of DENV-infected mice treated with saline. The lower clinical scores of DENV-infected mice treated with DV-B-120 indicated a reduction in acute-progressive illness symptoms, underscoring the potential therapeutic impact of DV-B-120. Investigations of DV-B-120's ability to restore the antiviral type I IFN response in the brain tissue of DENV-infected ICR suckling mice demonstrated its capacity to stimulate IFN and antiviral IFN-stimulated gene expression. DV-B-120 not only significantly delayed DENV-2-induced mortality and illness symptoms but also reduced viral numbers in the brain, ultimately restoring the innate antiviral response. These findings strongly suggest that DV-B-120 holds promise as a therapeutic agent against DENV infection and highlight its potential contribution in addressing the current lack of effective treatments for this infectious disease.IMPORTANCEThe prevalence of dengue virus (DENV) infection in tropical and subtropical regions is escalating due to factors like climate change and mosquito vector expansion. With over 300 million annual infections and potentially fatal outcomes, the urgent need for effective treatments is evident. While the approved Dengvaxia vaccine has variable efficacy, there are currently no antiviral drugs for DENV. This study explores seven compounds targeting the NS2B-NS3 protease, a crucial protein in DENV replication. These compounds exhibit inhibitory effects on DENV-2 NS2B-NS3, holding promise for disrupting viral replication and preventing severe manifestations. However, further research, including animal testing, is imperative to assess therapeutic efficacy and potential toxicity. Developing safe and potent treatments for DENV infection is critical in addressing the rising global health threat posed by this virus.

Creating Tiny Human “Organs” to Test medicines… and More!

Scientists have developed tiny cell models called microphysiological systems (MPSs) that mimic human organs, allowing medicines to be tested without using animals. MPS contain human cells carefully arranged to simulate a real organ’s structure and function. One type of MPS, called an organ-on-chip, also pumps fluids containing nutrients and oxygen through the model, similar to the function of blood flow in our bodies. These MPSs can test how medicines affect human cells and help scientists develop safer, more effective treatments for diseases. MPSs can also be personalized using a patient’s own cells, to find the best treatment for each person. While challenges remain, like cost and reliability, MPSs are steadily improving. Beyond testing medicines, they can be used to study dangerous environmental chemicals and to model diseases. We can even connect multiple “organs” to simulate the whole body. As these revolutionary technologies improve and become widely accepted, they could speed up drug development and reduce animal testing.

UJI AKTIVITAS ANALGESIK FRAKSI DAUN SENGGANI (Clidemia hirta [L] D.Don) PADA MENCIT PUTIH GALUR SWISS WEBSTER

Pain is a sensation that can interfere with activities and reduce the quality of life of the sufferer if left untreated and left prolonged. Senggani is one of the plants that contains flavonoid compounds and steroids that have analgesic activity. This study aimed to test the analgesic activity of senggani leaf fraction (Clidemia hirta [L] D.Don) in white mice swiss webster strain induced acetic acid and determine the fraction that most effectively produces analgesic activity. The type of research conducted is true experimental design based on Complete Randomized Design (RAL). The test animals used were 25 heads divided into 5 groups with 5 replications, namely negative control (CMC 1%), positive control (dexamethasone) and the test group of senggani leaf fraction, namely n-hexane, ethyl acetate, and ethanol fractions. The data obtained were analyzed using an alternative test from One Way ANOVA namely Kruskal Wallis with a follow-up test post hoc Mann Whitney test. The results showed that the fraction of senggani leaves in all types of solvents had analgesic activity compared to negative controls (CMC 1%) with a significant difference (p0.05). From these results it is concluded that the senggani leaf fraction can be used as an analgesic.

The Psychological Process Underlying Attitudes Toward Human-Animal Chimeric Brain Research: An Empirical Investigation

This study adopted an empirical method to investigate lay people’s attitudes toward the bioethical issues of human-animal chimeric brains. The results of online surveys showed that (1) people did not entirely reject chimeric brain research, but showed slightly more negative responses than ordinary animal testing; and that (2) their ethical concerns arose in connection with the perception that chimerism in the brain would humanize the animal. This means that people’s psychology was consistent with the ethical argument that crossing the human-animal boundary would bring moral confusion to our society. Meanwhile, it was not in line with another argument that moral status depended on having high capacities, and that chimerism would cause a problem if it enhanced animals’ capacities. Furthermore, this study analyzed additional psychological factors related to people’s moral judgment and the relationship among those factors. Several psychological factors, such as the perception that chimeric brain research is unnatural, were identified as mediating the relationship between perception of animal humanization and ethical concerns about creating and using chimeric brains. Introducing an empirical approach to the ethics of human-animal chimeric brains brought two findings: (1) this study informed us of socially shared intuition regarding this novel technology; and (2) it unveiled the psychological processes behind people’s ethical concerns in more detail than they spontaneously mentioned. These findings will help to build normative arguments and future policies that are understandable and acceptable to society.

3D Bioprinted Human Skin Model Recapitulating Native-Like Tissue Maturation and Immunocompetence as an Advanced Platform for Skin Sensitization Assessment.

Physiologically-relevant in vitro skin models hold the utmost importance for efficacy assessments of pharmaceutical and cosmeceutical formulations, offering valuable alternatives to animal testing. Here, an advanced immunocompetent 3D bioprinted human skin model is presented to assess skin sensitization. Initially, a photopolymerizable bioink is formulated using silk fibroin methacrylate, gelatin methacrylate, and photoactivated human platelet releasate. The developed bioink shows desirable physicochemical and rheological attributes for microextrusion bioprinting. The tunable physical and mechanical properties of bioink are modulated through variable photocuring time for optimization. Thereafter, the bioink is utilized to 3D bioprint "sandwich type" skin construct where an artificial basement membrane supports a biomimetic epidermal layer on one side and a printed pre-vascularized dermal layer on the other side within a transwell system. The printed construct is further cultured in the air-liquid interface for maturation. Immunofluorescence staining demonstrated a differentiated keratinocyte layer and dermal extracellular matrix (ECM)-remodeling by fibroblasts and endothelial cells. The biochemical estimations and gene-expression analysis validate the maturation of the printed model. The incorporation of macrophages further enhances the physiological relevance of the model. This model effectively classifies skin irritative and non-irritative substances, thus establishing itself as a suitable pre-clinical screening platform for sensitization tests.

Comparative Effects of Fermented and Unfermented Parkia biglobosa Bean Seeds in Diabetic Induced Experimental Rats

Aims: The present work investigates the hypoglycemic effect of fermented seeds of Parkia biglobosa (PB, African locust beans) in alloxan-induced diabetic rats. Its effect was compared with that of unfermented Parkia biglobosa.&#x0D; Methodology: In order to assess the hypoglycemic effects of the fermented seeds on experimental animals, fasting blood sugar (FBS), total cholesterol, triglycerides, high-density lipoprotein (HDL), and low-density lipoprotein (LDL) were determined. The microbiota analysis of the fecal sample was also determined.&#x0D; Result: Alloxan administered to the rats significantly increased the FBS of the test animals. However, dietary supplementation with fermented PB for three weeks ameliorated the FBS of the rats like that of the unfermented group. The animals fed with fermented Parkia biglobosa gained weight like those fed with unfermented Parkia biglobosa. High levels of HDL and low levels of LDL were observed in animals treated with fermented Parkia biglobosa compared with low levels of HDL and high levels of LDL observed in animals treated with unfermented Parkia biglobosa.&#x0D; Conclusion: The results of this present study demonstrate that both fermented and unfermented seeds of Parkia biglobosa exert a hypoglycemic effect. Hence, Parkia biglobosa has an anti-diabetic property with fermented form seemingly better.

Innovative Strategies for Photoallergy Assessment: Breaking Free from Animal Models in Cosmetic Ingredient Development

Photoallergy, a unique form of skin sensitization induced by specific compounds under ultraviolet irradiation, has traditionally been investigated using animals. However, the prohibition of animal testing for the assessment of cosmetic ingredients in Europe and other countries underscores the necessity for in vitro or in silico alternative methods. Currently, there are no validated methods for assessing photoallergy or photosensitization, presenting a significant challenge in the development of new cosmetic ingredients. This review examines the landscape of alternative methods for detecting photosensitization, emphasizing recent publications, and considering the underlying principles of the different proposed assays.

Abstract 6784: Cancer cell heterospheroids with stromal component used as drug testing and immune cell infiltration models

Abstract 3D cancer cell cultures have enabled new opportunities for replacing compound testing in experimental animals. However, driven by increased understanding of the need for more complex cancer cell models that consider drug resistance and penetration, hypoxia, and immuno-oncology applications, we have developed multicellular tumor heterospheroids composed of cancer cell lines, fibroblasts and immune cells. We have studied heterospheroids of cancer cell lines, HT29, MCF7, PANC1 and SW480 co-cultured with fibroblasts and 1) explored the architectural organization, 2) performed whole transcriptome analysis (WTA) of mono and co-cultures and 3) tested their capacity of immune cell infiltration. The architectural organization was studied in embedded heterospheroids. HT29 and MCF7 cells developed spheroids with the cancer cells surrounding the fibroblasts, whereas PANC1 cells mingled with the fibroblasts and SW480 cells were surrounded by fibroblasts. The fibroblasts expressed FAP-α mRNA as determined by ISH as well as collagen-1 as determined by immunohistochemistry. The hypoxia marker, Ca-IX, and the apoptosis marker, cleaved-Caspase-3, were dominating in the central core of the spheroids but the Ca-IX and Caspase-3 expression was not coordinated. In addition, WTA performed on day-7 heterospheroids, showed upregulated LOX, indicating that hypoxia-related markers known from genuine cancers are upregulated also in heterospheroids. WTA of day-7 mono-cell cultured and hetero-cultured spheroids, showed abundant ECM-, EMT-, and fibrosis-related expression in heterospheroids, thus reflecting a representative tumor-like microenvironment in heterospheroids compared to mono-culture spheroids. In particular, PANC1 spheroids showed a strong EMT profile with abundant FOXC2, TGF-β and TWIST expression. Genes related to Akt/mTOR and EGF pathways were consistently expressed in the tested heterospheroids. Evaluation of the cytotoxicity of anti-cancer compounds was performed using both cell viability and apoptosis assays. Exposure of the NCI drug library composed of 166 cytotoxic compounds to the heterospheroids, interestingly, revealed a complex output with limited correlation between cytotoxic and apoptotic effects. A subset of drug candidates is currently being further examined. PBMC-derived immune cells infiltrate heterospheroids and their infiltration is strongly enhanced after activation. The T cells' ability to infiltrate heterospheroids of different cancer types may serve as models of cold and hot tumors. We conclude that the cancer cell lines determine the architectural structure of heterospheroids, that ECM development is a prevalent part of the heterospheroids, and that the different heterospheroids possess varying ability of immune cell infiltration. Citation Format: Boye Schnack Nielsen, Natasha H. Madsen, Jesper Larsen, Monika Gad, Kim Holmstrøm. Cancer cell heterospheroids with stromal component used as drug testing and immune cell infiltration models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6784.

Abstract 4183: Utility of the highly immunodeficient SRG rat for combined drug efficacy, pharmacokinetics, and toxicology studies in tumor-bearing animals

Abstract Immunodeficient rodents are vital pre-clinical disease models. Historically, immunodeficient mice have been standard for hosting human cell and tissue xenografts, enabling cancer drug efficacy and tolerability testing. However, an immunodeficient rat that supports a variety of human cancer types provides a larger model for easier surgical manipulation and serial blood and tumor tissue sampling, allowing for efficacy, pharmacokinetics, and toxicology testing in the same animal. We created a Sprague Dawley Rag2 -/-, Il2rg -/- rat (SRG Rat®; SRG) that is highly supportive of human cell and tissue engraftment. The SRG rat lacks B, T, and NK cells and is more immunodeficient than the Nude rat. Here we show comparative pathology between the SRG rat and its parental strain, the CRL Sprague Dawley (CD). We also present data from an efficacy study in tumor bearing SRG rats, demonstrating that serial blood draws and tumor biopsies can be performed in tumor bearing animals to assess drug pharmacokinetics and pharmacodynamics within a single study. First, we assessed 10 SRG (5/sex) and 10 CD (5/sex) at 8-10 weeks of age. Body weight, hematology, and clinical chemistry parameters were measured. Microscopic examinations were performed on routine H&amp;E slide preparations. Male and female SRG displayed lower mean body weight when compared to sex- and age-matched CD. Compared to CD, SRG had reduced lymphoid tissue and circulating monocytes, and highly reduced circulating lymphocytes, neutrophils, eosinophils, and basophils; expected phenotypes for this severely immunodeficient rat. Thymus was not present in SRG rats, spleens were grossly smaller, and bone marrow cellularity was decreased compared to CD. Microscopically, mandibular lymph nodes and lymphoid tissue in spleen, lung, and intestine were not observed in SRG rats, consistent with the 92% decrease in circulating lymphocytes. To demonstrate the utility of the SRG for tumor inhibition, pharmacokinetics, and pharmacodynamics, we inoculated rats with human VCaP prostate cancer cell-derived tumors, then treated these animals and collected serial blood samples and tumor biopsies to assess tumor biomarkers. Treatment led to decreased PSA and AR, two key proteins expressed in prostate cancer, correlating to decreased tumor growth. These data demonstrate that the SRG rat is suitable for toxicology studies and has comparable pathology to the CD rat, except for reduced lymphoid tissue and WBCs, which are expected phenotypes. In addition, the SRG rat supports the growth of human cancer tissue for treatment studies. The large size of the SRG rat and comparative pathology to the CD rat support its value as an immunodeficient model for assessing toxicity during efficacy testing for de-risking safety concerns in the presence of the human target tissue. Citation Format: Diane Begemann, Cynthia Dunn, Darrell Robertson, Lauren Keach, Emily Baldwin, R. Grace Walton, Alex Van Engelenburg, Jessica Durrant, Laura Healy, Koh Meng Aw Yong, Michael J. Schlosser, Fallon K. Noto. Utility of the highly immunodeficient SRG rat for combined drug efficacy, pharmacokinetics, and toxicology studies in tumor-bearing animals [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4183.