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Certain immune-checkpoint inhibitors have a narrow therapeutic window (TW) as cancer therapeutics, and engineered dual-/multi-targeting agents could potentially widen the TW to bring true clinical benefits. We report a new rationally-designed bispecific-antibody (BsAb), HX009, simultaneously targeting PD1 and CD47 to improve both the efficacy and safety over the respective single-targeting agents by grafting the extracellular domain of SIRPα onto the parental anti-PD1-monoclonal antibody, HX008. This resulted in an IgG4-based "2 × 2" symmetric structure but with an intentionally-reduced CD47-binding affinity, suggesting a novel candidate cancer immunotherapy. Specifically, HX009 has binding affinity constant of 8.951 × 10–9 M for human PD1 and 2.557 × 10–8 M for human CD47, respectively, where the CD47 binding is significantly weaker as compared to the binding affinity of HX008 to PD1 as well as the binding affinity of SIRPα-Fc to CD47, leading to little binding to RBCs and platelets and is contrasting to many CD47-agents in development. However, HX009 effectively and simultaneously binds to the PD1 and CD47 on PD1+CD47+ T-cells via cis-binding and elicits enhanced T cell activation compared to the parental HX008. HX009 caused little cytokine-release in human peripheral blood mononuclear cells. HX009 cross-species binds to cynomolgus monkey PD1/CD47 but not to rodents, making cynomolgus monkeys the choice of species to investigate the pharmacokinetics (PK) and toxicology of HX009. HX009's anti-tumor activities were confirmed in several humanized preclinical mouse models by determining either its anti-PD1 (humanized hu-CD47-MC38 models) or anti-CD47 (HuT-102 lymphoma CDX and three PDX-AML models) functions, although limited available humanized models have hindered broadly demonstration of enhanced anti-tumor activities contributed from the dual targeting of the BsAb. The expanded DLBCL-PDX trial data suggested that both EBV-status and OX40 expression could potentially be two positive predictors for response to HX009. An intravenous (IV) infusion PK study in cynomolgus monkey revealed its largely vasculature distribution, terminal half-life (T1/2) of ~ 50 h, and dose-proportional exposure without accumulation. The anti-drug antibody (ADA) was observed in all monkeys as expected, affecting the PK parameters of repeated administration. The IV single-dose toxicology study with a 14-day observation revealed a maximum tolerated dose of 150 mg/kg, while the repeated-dose (once weekly for 4 weeks, 5 doses in total) study showed a highest non-severely toxic dose (HNSTD) of 15 mg/kg. The desired preclinical PK and safety profiles, along with its antitumor activity, support HX009's candidacy for its clinical development. [ABSTRACT FROM AUTHOR]

Background: Repeated inhalation of granulocyte-macrophage colony-stimulating factor (GM-CSF) was recently approved in Japan as a treatment for autoimmune pulmonary alveolar proteinosis. However, the detailed physiological and pathological effects of repeated inhalation in the long term, especially at increasing doses, remain unclear. Methods: In this chronic safety study, we administered 24 cynomolgus monkeys (Macaca fascicularis) aged 2–3 years with aerosolized sargramostim (a yeast-derived recombinant human GM-CSF [rhGM-CSF]) biweekly for 26 weeks across four dosing groups (0, 5, 100, and 500 µg/kg/day). We measured the serum GM-CSF antibody (GM-Ab) concentration by an ELISA and assessed the neutralizing capacity of GM-Ab using the GM-CSF-dependent cell line TF-1. We subjected lung tissue samples taken from all monkeys at 27 weeks to histopathological assessment using a sargramostim-specific monoclonal antibody to detect localization of residual sargramostim. Results: All the animals maintained good body condition and showed steady weight gain throughout the study. The pathological analyses of the lung revealed the formation of induced bronchus-associated lymphoid tissue (iBALT) in the lower respiratory tract, even at the clinical dose of 5 µg/kg/day. There was a relationship between the number or size of BALT and sargramostim dose or the serum GM-Ab levels. Immunohistochemical analyses revealed GM-Ab–producing cells in the follicular region of iBALT, with residual sargramostim in the follicles. Leucocyte counts were inversely correlated with GM-Ab levels in the high-dose groups. Additionally, serum GM-Ab from the treated animals significantly suppressed the alveolar macrophage proliferation activity of both Cynomolgus recombinant and rhGM-CSF in vitro. Conclusion: Long-term repeated inhalation of sargramostim led to iBALT formation in the lower respiratory tract, even at the clinical dose of 5 µg/kg/day, with the extent of iBALT formation increasing in a dose-dependent manner. Inhaled sargramostim was localized to the follicular region of iBALT nodules, which may induce the production of GM-Ab. [ABSTRACT FROM AUTHOR]

The article discusses the assessment of IgE and basophil activity in blood samples from nonhuman primates, specifically cynomolgus monkeys, in the context of developing new drugs to combat allergies. The study compares IgE biology between humans and cynomolgus monkeys, finding similarities in key residues and binding affinities for FcεRI. The research suggests that cynomolgus monkeys could serve as a relevant model for testing novel anti-IgE therapies, with potential applications in drug development and clinical trials. [Extracted from the article]

Abstract Background Repeated inhalation of granulocyte-macrophage colony-stimulating factor (GM-CSF) was recently approved in Japan as a treatment for autoimmune pulmonary alveolar proteinosis. However, the detailed physiological and pathological effects of repeated inhalation in the long term, especially at increasing doses, remain unclear. Methods In this chronic safety study, we administered 24 cynomolgus monkeys (Macaca fascicularis) aged 2–3 years with aerosolized sargramostim (a yeast-derived recombinant human GM-CSF [rhGM-CSF]) biweekly for 26 weeks across four dosing groups (0, 5, 100, and 500 µg/kg/day). We measured the serum GM-CSF antibody (GM-Ab) concentration by an ELISA and assessed the neutralizing capacity of GM-Ab using the GM-CSF-dependent cell line TF-1. We subjected lung tissue samples taken from all monkeys at 27 weeks to histopathological assessment using a sargramostim-specific monoclonal antibody to detect localization of residual sargramostim. Results All the animals maintained good body condition and showed steady weight gain throughout the study. The pathological analyses of the lung revealed the formation of induced bronchus-associated lymphoid tissue (iBALT) in the lower respiratory tract, even at the clinical dose of 5 µg/kg/day. There was a relationship between the number or size of BALT and sargramostim dose or the serum GM-Ab levels. Immunohistochemical analyses revealed GM-Ab–producing cells in the follicular region of iBALT, with residual sargramostim in the follicles. Leucocyte counts were inversely correlated with GM-Ab levels in the high-dose groups. Additionally, serum GM-Ab from the treated animals significantly suppressed the alveolar macrophage proliferation activity of both Cynomolgus recombinant and rhGM-CSF in vitro. Conclusion Long-term repeated inhalation of sargramostim led to iBALT formation in the lower respiratory tract, even at the clinical dose of 5 µg/kg/day, with the extent of iBALT formation increasing in a dose-dependent manner. Inhaled sargramostim was localized to the follicular region of iBALT nodules, which may induce the production of GM-Ab.

Introduction: Assisted reproductive technologies (ARTs), such as intracytoplasmic sperm injection and embryo transfer, are essential for generating genetically edited monkeys. Despite their importance, ARTs face challenges in recipient selection in terms of time and the number of animals required. The potential of superovulated monkeys, commonly used as oocyte donors, to serve as surrogate mothers, remains underexplored. The study aimed to compare the efficacy of superovulated and uterine-embryo synchronized recipients of embryo transfer in cynomolgus monkeys (Macaca fascicularis). Methods: This study involved 23 cynomolgus monkeys divided into two groups-12 superovulated recipients and 11 synchronized recipients. The evaluation criteria included measuring endometrial thickness on the day of embryo transfer and calculating pregnancy and implantation rates to compare outcomes between groups. Results: The study found no statistically significant differences in endometrial thickness (superovulated: 4.48 ± 1.36 mm, synchronized: 5.15 ± 1.58 mm), pregnancy rates (superovulated: 30.8%, synchronized: 41.7%), and implantation rates (superovulated: 14.3%, synchronized: 21.9%) between the groups (p > 0.05). Conclusion: The observations indicate that superovulated recipients are as effective as synchronized recipients for embryo transfer in cynomolgus monkeys. This suggests that superovulated recipients can serve as viable options, offering an efficient and practical approach to facilitate the generation of gene-edited models in this species. [ABSTRACT FROM AUTHOR]

Increasing complexity of mAbs in development creates challenges in predicting human pharmacokinetic (PK) parameters from preclinical data. The aim of this analysis was to identify optimal allometric scaling exponents. Data were extracted from literature to create a central database (currently the largest available published database) of two-compartment model parameters for mAbs (n = 59) in cynomolgus monkey (CM) and human. Global allometric exponents were calculated and drug-dependent factors were investigated as potential variables in determining the optimal scaling factor. The global exponents for scaling CM mAb PK data were 0.74 (CL), 0.80 (CL with Fc-modified mAbs excluded), 0.44 (CL with Fc-modified mAbs only), 0.71 (Q), 1.12 (V1), and 0.99 (V2). These values are in line with previously published literature values. [ABSTRACT FROM AUTHOR]

Background/Aim: Ischemic stroke is a major health concern globally and developing reliable animal models is crucial for understanding its pathophysiology. This study evaluated the relationship between cerebral angiographic findings and neurologic dysfunction in an acute non-human primate thromboembolic stroke model and determined the minimum clot length for suitable middle cerebral artery (MCA) occlusion. Materials and Methods: A thromboembolic stroke model was developed by injecting autologous blood clots (length: 1, 2, 3, 4, 5, and 10 cm, n=1 to 3, 14 monkeys in total) into the internal carotid artery of male cynomolgus monkeys. Digital subtraction angiography (DSA) and neurologic deficit observation were performed pre-; immediately after (DSA only); and 1, 3, 6, and 24 h after embolization, and the relationship between clot length, neurologic deficits, and cerebral infarction was assessed. Results: DSA confirmed MCA occlusion in all animals after the clot injection. Recanalization of the MCA was observed within 6 h post-embolization in animals with shorter clots (≤3 cm). Neurologic deficits were evident in animals with MCA occlusion and correlated with the clot length. Larger clots (≥5 cm) led to permanent MCA occlusion, significant neurologic deficits, and extensive cerebral infarction. Histopathological examination revealed ischemic damage in brain regions corresponding to the infarcted areas. Conclusion: Clot length is critical in determining the extent of neurologic dysfunction and cerebral infarction, with larger clots producing more severe outcomes. Furthermore, the minimum clot length required for model creation is 5 cm. [ABSTRACT FROM AUTHOR]

BACKGROUND: The clinical application of human induced pluripotent stem cell--derived cardiomyocytes (CMs) for cardiac repair commenced with the epicardial delivery of engineered cardiac tissue; however, the feasibility of the direct delivery of human induced pluripotent stem cell--derived CMs into the cardiac muscle layer, which has reportedly induced electrical integration, is unclear because of concerns about poor engraftment of CMs and posttransplant arrhythmias. Thus, in this study, we prepared purified human induced pluripotent stem cell--derived cardiac spheroids (hiPSC-CSs) and investigated whether their direct injection could regenerate infarcted nonhuman primate hearts. METHODS: We performed 2 separate experiments to explore the appropriate number of human induced pluripotent stem cell--derived CMs. In the first experiment, 10 cynomolgus monkeys were subjected to myocardial infarction 2 weeks before transplantation and were designated as recipients of hiPSC-CSs containing 2107 CMs or the vehicle. The animals were euthanized 12 weeks after transplantation for histological analysis, and cardiac function and arrhythmia were monitored during the observational period. In the second study, we repeated the equivalent transplantation study using more CMs (6107 CMs). RESULTS: Recipients of hiPSC-CSs containing 2107 CMs showed limited CM grafts and transient increases in fractional shortening compared with those of the vehicle (fractional shortening at 4 weeks after transplantation [mean ± SD]: 26.2±2.1%; 19.3±1.8%; P<0.05), with a low incidence of posttransplant arrhythmia. Transplantation of increased dose of CMs resulted in significantly greater engraftment and long-term contractile benefits (fractional shortening at 12 weeks after transplantation: 22.5±1.0%; 16.6±1.1%; P<0.01, left ventricular ejection fraction at 12 weeks after transplantation: 49.0±1.4%; 36.3±2.9%; P<0.01). The incidence of posttransplant arrhythmia slightly increased in recipients of hiPSC-CSs containing 6107 CMs. [ABSTRACT FROM AUTHOR]

AbstractThe pharmacokinetics and tissue distribution of renadirsen sodium, a dystrophin exon-skipping phosphorothioate-modified antisense oligonucleotide with 2’-O,4’-C-ethylene-bridged nucleic acid (ENA), after subcutaneous or intravenous administration to cynomolgus monkeys were investigated. The plasma concentration of renadirsen after subcutaneous administration at 1, 3, and 10 mg/kg increased with the dose. The absolute bioavailability at 3 mg/kg after subcutaneous administration was calculated as 88.6%, and the time to reach maximum plasma concentration of renadirsen was within 4 h, indicating the efficient and rapid absorption following subcutaneous administration. The exposure of muscle tissues to renadirsen was found to increase with repeated dosing at 6 mg/kg, and higher exposure was observed in the diaphragm and heart than in the quadriceps femoris and anterior tibialis muscles. Renadirsen achieved more exon 45-skipped dystrophin mRNA in the diaphragm and heart than in the quadriceps femoris and anterior tibialis muscles. Renadirsen also showed a cumulative skipping effect in a repeated-dose study. The findings on exon 45-skipped dystrophin mRNA in these muscle tissues were consistent with the concentration of renadirsen in these tissues. Because it is not feasible to directly evaluate drug concentration and exon skipping in the heart and diaphragm in humans, the pharmacokinetics and pharmacodynamics of renadirsen in these tissues in monkeys are crucial for the design and interpretation of clinical settings. [ABSTRACT FROM AUTHOR]

INTRODUCTION: More robust non‐human primate models of Alzheimer's disease (AD) will provide new opportunities to better understand the pathogenesis and progression of AD. METHODS: We designed a CRISPR/Cas9 system to achieve precise genomic deletion of exon 9 in cynomolgus monkeys using two guide RNAs targeting the 3′ and 5′ intron sequences of PSEN1 exon 9. We performed biochemical, transcriptome, proteome, and biomarker analyses to characterize the cellular and molecular dysregulations of this non‐human primate model. RESULTS: We observed early changes of AD‐related pathological proteins (cerebrospinal fluid Aβ42 and phosphorylated tau) in PSEN1 mutant (ie, PSEN1‐ΔE9) monkeys. Blood transcriptome and proteome profiling revealed early changes in inflammatory and immune molecules in juvenile PSEN1‐ΔE9 cynomolgus monkeys. DISCUSSION: PSEN1 mutant cynomolgus monkeys recapitulate AD‐related pathological protein changes, and reveal early alterations in blood immune signaling. Thus, this model might mimic AD‐associated pathogenesis and has potential utility for developing early diagnostic and therapeutic interventions. Highlights: A dual‐guide CRISPR/Cas9 system successfully mimics AD PSEN1‐ΔE9 mutation by genomic excision of exon 9.PSEN1 mutant cynomolgus monkey‐derived fibroblasts exhibit disrupted PSEN1 endoproteolysis and increased Aβ secretion.Blood transcriptome and proteome profiling implicate early inflammatory and immune molecular dysregulation in juvenile PSEN1 mutant cynomolgus monkeys.Cerebrospinal fluid from juvenile PSEN1 mutant monkeys recapitulates early changes of AD‐related pathological proteins (increased Aβ42 and phosphorylated tau). [ABSTRACT FROM AUTHOR]

In a 3-month toxicity study in cynomolgus monkeys at a European contract laboratory, animals were infected with HAV, initially resulting in hepatic injury being incorrectly attributed to the test compound. Elevated serum ALT/AST/GLDH (5- to 10-fold) were noted in individual animals from all groups including controls, with no apparent dose, exposure, or time-related relationship. Liver histopathology revealed minimal to slight inflammatory cell accumulation in periportal zones of most animals, and minimal to slight hepatocyte degeneration/necrosis in 10/42 animals from all groups. As these findings were more pronounced in 6 drug-treated animals, including 2/6 in the low dose group, the draft report concluded: " treatment-related hepatotoxicity at all dose levels precluded determination of a NOAEL." However, the unusual pattern of hepatotoxicity suggested a factor other than drug exposure might have caused the hepatic effects. Therefore, snap-frozen liver samples were tested for hepatitis viruses using a PCR method. Tests for hepatitis B, C, and E virus were negative; however, 20/42 samples were positive for hepatitis A virus (HAV). Infection was strongly associated with increased serum ALT/GLDH, and/or hepatocyte degeneration/necrosis. Re-evaluation of the study in light of these data concluded that the hepatic injury was not drug-related. A subsequent 6-month toxicology study in HAV-vaccinated cynomolgus monkeys confirmed the absence of hepatotoxicity. Identification of HAV infection supported progression of the drug candidate into later clinical trials. Although rarely investigated, subclinical HAV infection has occasionally been reported in laboratory primates, including those used for toxicology studies and it may be more prevalent than the literature indicates. [ABSTRACT FROM AUTHOR]

Background: Cynomolgus monkeys (Macaca fascicularis) are essential in biomedical research, including reproductive studies. However, the application of human estimated foetal weight (EFW) formulas using ultrasonography (USG) in these non‐human primates is not well established. Objectives: This study aims to evaluate the applicability of human EFW formulas for estimating foetal weight in cynomolgus monkeys at approximately 130 days of gestation. Methods: Our study involved nine pregnant cynomolgus monkeys. We measured foetal parameters, including biparietal diameter, head circumference, abdominal circumference and femur length using USG. The EFW was calculated using 11 human EFW formulas. The actual birthweight (ABW) was recorded following Cesarean section, the day after the EFW calculation. For comparing EFW and ABW, we employed statistical methods such as mean absolute percentage error (APE) and Bland–Altman analysis. Results: The ABW ranged between 200.36 and 291.33 g. Among the 11 formulas, the Combs formula showed the lowest APE (4.3%) and highest correlation with ABW (p < 0.001). Notably, EFW and ABW differences for the Combs formula were ≤5% in 66.7% and ≤10% in 100% of cases. The Bland–Altman analysis supported these results, showing that all cases fell within the limits of agreement. Conclusions: The Combs formula is applicable for estimating the weight of cynomolgus monkey fetuses with USG at approximately 130 days of gestation. Our observations suggest that the Combs formula can be applied in the prenatal care and biomedical research of this species. [ABSTRACT FROM AUTHOR]

A captive 17‐year‐old male cynomolgus monkey (Macaca fascicularis) developed diffuse large B‐cell lymphoma (DLBCL). This was the first report of DLBCL presenting with a mandible mass and violation of the paranasal sinus in a cynomolgus monkey. The neoplasm showed marked microscopical malignant aspects. Immunohistochemical staining showed strong positive expression of CD20. These features may contribute to the diagnosis and therapeutics of DLBCL in NHPs. [ABSTRACT FROM AUTHOR]

Abstract Mesenchymal stem cells (MSCs) have demonstrated promising advantages in the therapies of many diseases, while its multi-directional differentiation potential and immunotoxicity are the major concerns hindered their clinical translation. In this study, human umbilical Mesenchymal stem cell (hUC-MSCs) were labeled with a near-infrared fluorescent dye DiR before infused into cynomolgus monkeys, and the amount of hUC-MSCs in the peripheral blood were dynamically estimated from 5 min to 28 days post a single administration at 3 × 106 cells/kg and 2 × 107 cells/kg intravenously. As results, some hUC-MSCs distributed to the whole body within 5 min, while most of the cells accumulate in the lungs along with the systemic blood circulation, and subsequently released into the blood. The toxicity potentials of hUC-MSCs were investigated in another 30 cynomolgus monkeys, and the cells were repeatedly administrated at doses of 3 × 106 cells/kg and 2 × 107 cells/kg for 5 times on a weekly basis, with a recovery period of 1 months. hUC-MSCs showed no obvious toxic effects in cynomolgus monkeys, except xenogeneic immune rejection to human stem cells. Low levels of the hUC-MSC gene were detected in the peripheral blood of a few animals administered 2 × 107 cells/kg at 30 min subsequent to the first and last administration, and there was no significant difference in the copy number of the hUC-MSC gene in the blood samples compared with the first and last administration, indicating that the hUC-MSC was not significantly amplified in vivo, and it its safe in non-human primates. Our study for the first time verified the safety of long-term use of hUC-MSCs in primates. We have pioneered a technology for the real-time detection of hUC-MSCs in peripheral blood and provide dynamicand rapid monitoring of the distribution characteristics of hUC-MSCs in vivo. Here, we provide data supporting the application of such products for clinical treatment and the application of stem cells in major refractory diseases and regenerative medicine.

Mesenchymal stem cells (MSCs) have demonstrated promising advantages in the therapies of many diseases, while its multi-directional differentiation potential and immunotoxicity are the major concerns hindered their clinical translation. In this study, human umbilical Mesenchymal stem cell (hUC-MSCs) were labeled with a near-infrared fluorescent dye DiR before infused into cynomolgus monkeys, and the amount of hUC-MSCs in the peripheral blood were dynamically estimated from 5 min to 28 days post a single administration at 3 × 106 cells/kg and 2 × 107 cells/kg intravenously. As results, some hUC-MSCs distributed to the whole body within 5 min, while most of the cells accumulate in the lungs along with the systemic blood circulation, and subsequently released into the blood. The toxicity potentials of hUC-MSCs were investigated in another 30 cynomolgus monkeys, and the cells were repeatedly administrated at doses of 3 × 106 cells/kg and 2 × 107 cells/kg for 5 times on a weekly basis, with a recovery period of 1 months. hUC-MSCs showed no obvious toxic effects in cynomolgus monkeys, except xenogeneic immune rejection to human stem cells. Low levels of the hUC-MSC gene were detected in the peripheral blood of a few animals administered 2 × 107 cells/kg at 30 min subsequent to the first and last administration, and there was no significant difference in the copy number of the hUC-MSC gene in the blood samples compared with the first and last administration, indicating that the hUC-MSC was not significantly amplified in vivo, and it its safe in non-human primates. Our study for the first time verified the safety of long-term use of hUC-MSCs in primates. We have pioneered a technology for the real-time detection of hUC-MSCs in peripheral blood and provide dynamicand rapid monitoring of the distribution characteristics of hUC-MSCs in vivo. Here, we provide data supporting the application of such products for clinical treatment and the application of stem cells in major refractory diseases and regenerative medicine. [ABSTRACT FROM AUTHOR]

Neurotensin (NTS) is a 13‐amino acid peptide which is highly expressed in the mammalian ovary in response to the luteinizing hormone surge. Antibody neutralization of NTS in the ovulatory follicle of the cynomolgus macaque impairs ovulation and induces follicular vascular dysregulation, with excessive pooling of red blood cells in the follicle antrum. We hypothesize that NTS is an essential intrafollicular regulator of vascular permeability. In the present study, follicle injection of the NTS receptor antagonist SR142948 also resulted in vascular dysregulation. To measure vascular permeability changes in vitro, primary macaque ovarian microvascular endothelial cells (mOMECs) were enriched from follicle aspirates and studied in vitro. When treated with NTS, permeability of mOMECs decreased. RNA sequencing (RNA‐Seq) of mOMECs revealed high mRNA expression of the permeability‐regulating adherens junction proteins N‐cadherin (CDH2) and K‐cadherin (CDH6). Immunofluorescent detection of CDH2 and CDH6 confirmed expression and localized these cadherins to the cell–cell boundaries, consistent with function as components of adherens junctions. mOMECs did not express detectable levels of the typical vascular endothelial cadherin, VE‐cadherin (CDH5) as determined by RNA‐Seq, qPCR, western blot, and immunofluorescence. Knockdown of CDH2 or CDH6 via siRNA abrogated the NTS effect on mOMEC permeability. Collectively, these data suggest that NTS plays an ovulation‐critical role in vascular permeability maintenance, and that CDH2 and CDH6 are involved in the permeability modulating effect of NTS on the ovarian microvasculature. NTS can be added to a growing number of angiogenic regulators which are critical for successful ovulation. [ABSTRACT FROM AUTHOR]

Abstract Background Cynomolgus monkeys (Macaca fascicularis) are essential in biomedical research, including reproductive studies. However, the application of human estimated foetal weight (EFW) formulas using ultrasonography (USG) in these non‐human primates is not well established. Objectives This study aims to evaluate the applicability of human EFW formulas for estimating foetal weight in cynomolgus monkeys at approximately 130 days of gestation. Methods Our study involved nine pregnant cynomolgus monkeys. We measured foetal parameters, including biparietal diameter, head circumference, abdominal circumference and femur length using USG. The EFW was calculated using 11 human EFW formulas. The actual birthweight (ABW) was recorded following Cesarean section, the day after the EFW calculation. For comparing EFW and ABW, we employed statistical methods such as mean absolute percentage error (APE) and Bland–Altman analysis. Results The ABW ranged between 200.36 and 291.33 g. Among the 11 formulas, the Combs formula showed the lowest APE (4.3%) and highest correlation with ABW (p

Neural tube defects (NTDs) are severe congenital neurodevelopmental disorders arising from incomplete neural tube closure. Although folate supplementation has been shown to mitigate the incidence of NTDs, some cases, often attributable to genetic factors, remain unpreventable. The SHROOM3 gene has been implicated in NTD cases that are unresponsive to folate supplementation; at present, however, the underlying mechanism remains unclear. Neural tube morphogenesis is a complex process involving the folding of the planar epithelium of the neural plate. To determine the role of SHROOM3 in early developmental morphogenesis, we established a neuroepithelial organoid culture system derived from cynomolgus monkeys to closely mimic the in vivo neural plate phase. Loss of SHROOM3 resulted in shorter neuroepithelial cells and smaller nuclei. These morphological changes were attributed to the insufficient recruitment of cytoskeletal proteins, namely fibrous actin (F-actin), myosin II, and phospho-myosin light chain (PMLC), to the apical side of the neuroepithelial cells. Notably, these defects were not rescued by folate supplementation. RNA sequencing revealed that differentially expressed genes were enriched in biological processes associated with cellular and organ morphogenesis. In summary, we established an authentic in vitro system to study NTDs and identified a novel mechanism for NTDs that are unresponsive to folate supplementation. [ABSTRACT FROM AUTHOR]

Abstract Sepsis is a life‐threatening condition that requires urgent medical attention. In this study, we investigated the potential of soybean‐derived peptides (soy peptides) as an anti‐inflammatory supplement for patients with sepsis. Cynomolgus monkeys were induced with sepsis by intravenous administration of three different concentrations of Escherichia coli (6.3 × 103, 6.3 × 105, and 6.3 × 107 CFU/kg). Changes in body weight and inflammatory markers were analyzed, and the high dose of 6.3 × 107 CFU/kg was found to be the most appropriate for sepsis modeling. Controlled enzymatic hydrolysis was then used to release bioactive soy peptides from soy protein isolate, which were administered to monkeys with sepsis. The results showed that soy peptide supplementation significantly suppressed inflammatory responses and improved coagulation abnormalities in the monkeys. Our findings suggest that soy peptides could be an effective clinical nutritional supplement for patients with sepsis, with potential benefits for reducing inflammation and improving coagulation. These results provide a foundation for further research into the development of novel nutritional supplements for patients with sepsis.

Animal models are an integral part of the drug development and evaluation process. However, they are unsurprisingly imperfect reflections of humans, and the extent and nature of many immunological differences are unknown. With the rise of targeted and biological therapeutics, it is increasingly important that we understand the molecular differences in the immunological behavior of humans and model organisms. However, very few antibodies are raised against non-human primate antigens, and databases of cross-reactivity between species are incomplete. Thus, we screened 332 antibodies in five immune cell populations in blood from humans and four non-human primate species generating a comprehensive cross-reactivity catalog that includes cell type-specificity. We used this catalog to create large mass cytometry universal cross-species phenotyping and signaling panels for humans, along with three of the model organisms most similar to humans: rhesus and cynomolgus macaques and African green monkeys; and one of the mammalian models most widely used in drug development: C57BL/6 mice. As a proof-of-principle, we measured immune cell signaling responses across all five species to an array of 15 stimuli using mass cytometry. We found numerous instances of different cellular phenotypes and immune signaling events occurring within and between species, and detailed three examples (double-positive T cell frequency and signaling; granulocyte response to Bacillus anthracis antigen; and B cell subsets). We also explore the correlation of herpes simian B virus serostatus on the immune profile. Antibody panels and the full dataset generated are available online as a resource to enable future studies comparing immune responses across species during the evaluation of therapeutics.

This review on cynomolgus monkey (Macaca fascicularis) blastoids discusses a breakthrough in modeling early non‐human primate embryogenesis, offering insights into embryonic development and implantation processes. It acknowledges ethical challenges and animal welfare considerations in developmental biology, suggests potential applications in human reproductive medicine, and highlights the need for ongoing ethical and technical refinement. [ABSTRACT FROM AUTHOR]

Abstract Background As a supplement for promoting hair health, Shi-Bi-Man (SBM) is a prescription comprising various traditional Chinese medicines. Though SBM has been reported to promote hair regeneration, its molecular mechanism remains unclear. Cynomolgus monkeys (Macaca fascicularis) are non-human primates with a gene expression profile similar to that of humans. The purpose of this research is to evaluate the effect of SBM on promoting hair regeneration in cynomolgus monkeys and to reveal the underlying mechanism. Methods The effect of SBM on hair regeneration was observed by skin administration on 6 cynomolgus monkeys with artificial back shaving. The molecular mechanism of SBM was studied using single-cell RNA sequencing (scRNA-seq) in combination with quantitative polymerase chain reaction (qPCR) detection for gene transcription level, and immunofluorescence staining verification for protein level. Results SBM significantly induced hair regeneration in cynomolgus monkeys, increased hair follicle number and facilitated hair follicle development. ScRNA-seq revealed an increase in the number of hair follicle stem cells (HFSCs) with a higher activation state, as evidenced by the higher expression of activation marker LDHA related to metabolism and the proliferation marker MKI67. Immunofluorescence analysis at the protein level and qPCR at the mRNA level confirmed the sequencing data. Cellchat analysis revealed an enrichment of ligand-receptor pairs involved in intercellular communication in Laminin-related pathways. Conclusion SBM significantly promotes hair regeneration in cynomolgus monkeys. Mechanically, SBM can up-regulate LDHA-mediated lactic acid metabolism and drive HFSC activation, which in turn promotes the proliferation and differentiation of HFSCs.

Background: A novel buprenorphine (BUP) extended‐release formulation (BUP‐XR) produced as a lipid‐encapsulated, low viscosity BUP suspension for subcutaneous (SC) injection to control pain was evaluated for pharmacokinetics and safety in four adult male cynomolgus monkeys. Methods: Each animal was given 0.2 mg/kg reformulated BUP‐XR SC. Clinical observations were made during the course of the study. Blood samples were obtained from each animal before BUP‐XR administration, 6, 24, 48, 72, and 96 h post‐BUP‐XR injection. Plasma levels of buprenorphine were analyzed using HPLC‐MS/MS. The PK values calculated included peak plasma concentration of the BUP analyte, time to peak plasma concentration, plasma half‐life, area under the plasma concentration–time curve, clearance, apparent volume of distribution, and elimination rate constant (Cmax, Tmax, T½, AUC0‐t, CL, Vd, and Ke, respectively). Results: Observable adverse clinical signs were not detected. BUP concentration peaked from 6 to 48 h, then declined in a linear fashion. Quantifiable plasma BUP was measured in all monkeys at all time points. Results indicate that a single BUP‐XR dose at 0.2 mg/kg can reliably provide plasma levels of BUP reported in the literature to be therapeutically relevant for up to 96 h. Conclusions: Because of the lack of any clinical observations or adverse effects at the injection site or absence of observable abnormal behaviors, it may be concluded that the use of BUP‐XR is safe and efficacious in this species of non‐human primate at the dose regimen described in this study for up to 96 h post‐administration. [ABSTRACT FROM AUTHOR]

ASP8062 is an orally available GABAB receptor positive allosteric modulator (PAM). This study assessed the potential of ASP8062 for treating opioid use disorder (OUD). Three rhesus monkeys were pretreated with ASP8062 (0.3, 1 or 3 mg/kg) by oral administration 1 h prior to a 2-h morphine self-administration session (0.03 mg/kg, iv, per injection) under a fixed-ratio 5 schedule. We further examined the potential worsening of morphine-induced respiratory suppression by ASP8062 after coadministration of morphine (10 mg/kg, sc) and ASP8062 (10 mg/kg, po) in cynomolgus monkeys using a custom-made whole-body plethysmograph. Plasma concentrations of ASP8062 (3 or 10 mg/kg, po) were assessed in cynomolgus monkeys using liquid chromatography-tandem mass spectroscopy (LC-MS/MS). ASP8062 at 3 mg/kg, po decreased the morphine self-administrations with significant differences from the vehicle-treated group (IC50 = 0.97 ± 0.36 mg/kg). Exposure levels at 3 mg/kg observed in monkeys were comparable to the clinical exposure levels which positive pharmacodynamic effects were previously shown. Further, ASP8062 did not potentiate morphine-induced respiratory suppression up to exposure levels higher than the clinically relevant dose. ASP8062 may reduce opioid use in OUD patients without affecting respiratory system, providing justification for further ASP8062 development as a potential treatment option for OUD.

Acute gastric dilatation (AGD) is one of the most prevalent and life-threatening diseases in nonhuman primates worldwide. However, the etiology of this syndrome has not been determined. Recently, sudden death occurred in a 7-year-old female cynomolgus monkey with a history of fecal microbiota transplantation using diarrheic stools. The monkey had undergone surgery previously. On necropsy, gastric dilatation and rupture demonstrated a tetrad arrangement on histopathologic examination. On 16S rRNA sequencing, a high population of Clostridium ventriculi was identified in the duodenum adjacent to stomach but not in the colon. This paper is the first report of Clostridium ventriculi infection in a cynomolgus macaque with acute gastric dilatation and rupture. [ABSTRACT FROM AUTHOR]

Screws coated with fibroblast growth factor 2 (FGF-2)-calcium phosphate (CP) composite layers exhibit enhanced soft tissue and bone formation and angiogenesis because of the biological activity of FGF-2. Furthermore, the mitogenic activity of the FGF-2 within the composite layers remains unchanged after gamma-ray sterilization, which may improve the storage stability prior to clinical use. However, the in vivo safeties of these screws as spinal implants remain unknown. Here, a randomized controlled trial, involving non-human primates, investigated the safety of using FGF-2-CP composite layer-coated screws after either gamma-ray sterilization or aseptic processing. Titanium alloy screws coated with FGF-2-CP composite layers and subjected to either gamma-ray sterilization at 25 kGy (GS group) or aseptic storage (AS group) were implanted into the vertebral bodies of two cynomolgus monkeys exceeding 12 weeks (day 99). Physiological, histological, and radiographic investigations were performed to evaluate the safeties of the screws. There were no serious adverse events, such as surgical site infection, significant loss of body weight, or abnormal blood test results. No radiolucent areas were observed around the screws from the GS or AS group throughout the study. In the intraosseous region, no significant differences were observed in bone and fibrous tissue apposition rates and rate of bone formation between the two groups (p = 0.49, 0.77, and 0.11, respectively). Neither tumor lesions nor accumulation of lymphocytes and neutrophils were observed in either group. Our data suggest that FGF-2-CP composite layer-coated screws subjected to terminal gamma-ray sterilization are as safe as those fabricated in aseptic processing. [ABSTRACT FROM AUTHOR]

Background: As a supplement for promoting hair health, Shi-Bi-Man (SBM) is a prescription comprising various traditional Chinese medicines. Though SBM has been reported to promote hair regeneration, its molecular mechanism remains unclear. Cynomolgus monkeys (Macaca fascicularis) are non-human primates with a gene expression profile similar to that of humans. The purpose of this research is to evaluate the effect of SBM on promoting hair regeneration in cynomolgus monkeys and to reveal the underlying mechanism. Methods: The effect of SBM on hair regeneration was observed by skin administration on 6 cynomolgus monkeys with artificial back shaving. The molecular mechanism of SBM was studied using single-cell RNA sequencing (scRNA-seq) in combination with quantitative polymerase chain reaction (qPCR) detection for gene transcription level, and immunofluorescence staining verification for protein level. Results: SBM significantly induced hair regeneration in cynomolgus monkeys, increased hair follicle number and facilitated hair follicle development. ScRNA-seq revealed an increase in the number of hair follicle stem cells (HFSCs) with a higher activation state, as evidenced by the higher expression of activation marker LDHA related to metabolism and the proliferation marker MKI67. Immunofluorescence analysis at the protein level and qPCR at the mRNA level confirmed the sequencing data. Cellchat analysis revealed an enrichment of ligand-receptor pairs involved in intercellular communication in Laminin-related pathways. Conclusion: SBM significantly promotes hair regeneration in cynomolgus monkeys. Mechanically, SBM can up-regulate LDHA-mediated lactic acid metabolism and drive HFSC activation, which in turn promotes the proliferation and differentiation of HFSCs. [ABSTRACT FROM AUTHOR]

Objective To investigate the establishment of a six-gene-edited pig-to-non-human primate kidney xenotransplantation model. Methods The kidney of humanized genetically-edited pig (GTKO/β4GalNT2KO/CMAHKO/hCD55/hCD46/hTBM) was transplanted into a cynomolgus monkey. The survival of the recipient and kidney condition after blood perfusion were observed. The parenchymal echo, blood flow changes, and size of the kidney were monitored on a regular basis. Routine blood test, kidney function test and electrolyte assessment were carried out. Dynamic changes of urine, feces and body mass were monitored. At the end of life, the transplant kidney, heart, liver, spleen, lung, and cecum were collected for pathological examination. Results The recipient died at postoperative 7 d. After blood flow was restored, the kidney was properly perfused, the organ was soft and the color was normal. At the end of the recipient’s life, a slight amount of purulent secretion was attached to the ventral side of the kidney, with evident congestion and swelling, showing the appearance of "red kidney". Postoperatively, the echo of renal parenchyma was increased, blood flow was decreased, the cortex was gradually thickened, and a slight amount of effusion surrounded the kidney and abdominal cavity over time. In the recipient, the amount of peripheral red blood cells, hemoglobin, albumin, and platelets was progressively decreased, and serum creatinine level was increased to 308 μmol/L at postoperative 7 d, whereas the K+ concentration did not significantly change. Light yellow urine was discharged immediately after surgery, diet and drinking water were resumed within postoperative 3 h, and light yellow and normal-shape stool was discharged. The reddish urine was gradually restored to normal color within postoperative 1 d, which were consistent with the results of the routine urine test. A large amount of brown bloody stool was discharged twice in the morning of 2 d after surgery. Omeprazole was given for acid suppression, and the stool returned to normal at postoperative 4 d. The β2-microglobulin level was increased to 0.75 mg/L at postoperative 7 d. The body mass was increased by 1.7 kg. Autopsy pathological examination showed interstitial edema and bleeding of the transplant kidney, a large amount of infiltration of lymphocytes and macrophages, infiltration of lymphocytes in the arteriole wall and arterial cavity, accompanied by arteritis changes, lymphocyte infiltration in the cecal stroma and congestion in the spleen tissues. No significant abnormal changes were observed in other organs. Conclusions The humanized genetically-edited pig-to-non-human primate kidney xenotransplantation model is successfully established, and postoperative survival of the recipient is 1 week. [ABSTRACT FROM AUTHOR]

Fc-engineering is commonly used to improve the therapeutic potency of antibody (Ab) treatments. Because FcƳRIIb is the only inhibitory FcƳR that contains an immunoreceptor tyrosinebased inhibition motif (ITIM), Fc-engineered Abs with enhanced binding affinity to FcƳRIIb might provide immune suppression in clinical contexts. GYM329 is an anti-latent myostatin Fc-engineered Ab with increased affinity to FcƳRIIb which is expected to improve muscle strength in patients with muscular disorders. Cross-linking of FcƳRIIb by immune complex (IC) results in phosphorylation of ITIM to inhibit immune activation and apoptosis in B cells. We examined whether the IC of Fc-engineered Abs with enhanced binding affinity to FcƳRIIb causes phosphorylation of ITIM or B cell apoptosis using GYM329 and its Fc variant Abs in human and cynomolgus-monkey (cyno) immune cells in vitro. IC of GYM329 with enhanced binding affinity to human FcƳRIIb (×5) induced neither ITIM phosphorylation nor B cell apoptosis. As for GYM329, FcƳRIIb should work as an endocytic receptor of small IC to sweep latent myostatin, so it is preferable that GYM329 induces neither ITIM phosphorylation nor B cell apoptosis to prevent immune suppression. In contrast, IC of myo-HuCy2b, the Ab with enhanced binding affinity to human FcƳRIIb (×4), induced ITIM phosphorylation and B cell apoptosis. The result of the present study demonstrated that Fc-engineered Abs with similar binding affinity to FcƳRIIb had different effects. Thus, it is important to also investigate FcƳR-mediated immune functions other than binding to fully understand the biological effects of Fc-engineered Abs. [ABSTRACT FROM AUTHOR]

ObjectiveBy observing the impact of Bevacizumab injection on the speed of wound healing and the expression of CD34 in cynomolgus monkeys, to verify its delayed wound healing mechanism, and to provide clinical reference for the dosage and frequency of Bevacizumab treatment for oncology surgery patients.MethodsSix male cynomolgus monkeys underwent full-thickness skin resection on the back to establish a wound healing evaluation model. Three cynomolgus monkeys were randomly selected as the saline group, and the other three cynomolgus monkeys were allocated to the Bevacizumab treatment group. The monkeys of Bevacizumab group were administrated with Bevacizumab injection (30 mg/kg, i.v.) on day 0, day 4, day 8, and day 11, respectively. The monkeys of saline group were injected with the same volume of saline as the group receiving Bevacizumab. The blood routine test was conducted, and the wound healing of each group of cynomolgus monkeys was observed 28 days after operation. The wound healing rate was analyzed by software Image J, and the severity of the wound was assessed by scoring method. Wound healing skin samples of 6 cynomolgus monkeys were obtained after 4 weeks, and the expression level of CD34 in the wound skin tissues was detected using immunohistochemistry.ResultsA skin excision wound model has already been established in cynomolgus monkeys. The number of white blood cells and neutrophils in the blood of cynomolgus monkeys increased significantly on the 3rd day after operation (P

ObjectiveTo study the skeletal characteristics of cynomolgus monkeys at different ages, and to provide reference values for skeletal research using cynomolgus monkeys as model animals.MethodsA total of 283 cynomolgus monkeys aged 1-19 years were selected and divided into 8 groups according to sex and age :1 year≤age

Background: Melatonin is a multifunctional hormone synthesized in the pineal gland and peripheral reproductive tissues that regulates many biological processes. There is increasing evidence for a role of melatonin in oocyte maturation and embryonic development in various mammals. However, no study has reported evidence for the existence of melatonergic system, such as melatonin synthesis enzymes, melatonin membrane receptors, or melatonin binding sites in non‐human primate cumulus–oocyte complexes (COCs). Methods: Reverse transcription polymerase chain reaction (RT‐PCR) and immunocytochemistry were performed to detect transcripts and proteins of the rate‐limiting enzyme in melatonin synthesis (arylalkylamine N‐acetyltransferase, AANAT), melatonin membrane receptors (MT1 and MT2), and a melatonin binding site (NRH: quinone oxidoreductase 2, NQO2) in cynomolgus monkey COCs. Results: RT‐PCR analyses revealed the presence of AANAT, MT1, MT2, and NQO2 transcripts in granulosa cells, germinal vesicle (GV)‐ and metaphase II (MII)‐stage cumulus cells, and oocytes. Immunocytochemistry revealed the presence of AANAT, MT1, MT2, and NQO2 proteins in GV‐ and MII‐stage COCs. Conclusions: Our results provide the first evidence for the existence of the rate‐limiting enzyme required for melatonin synthesis, melatonin membrane receptors, and a melatonin binding site in non‐human primate COCs. [ABSTRACT FROM AUTHOR]

The spermatogonial compartment maintains spermatogenesis throughout the reproductive lifespan. Single-cell RNA sequencing (scRNA-seq) has revealed the presence of several spermatogonial clusters characterized by specific molecular signatures. However, it is unknown whether the presence of such clusters can be confirmed in terms of protein expression and whether protein expression in the subsets overlaps. To investigate this, we analyzed the expression profile of spermatogonial markers during the seminiferous epithelial cycle in cynomolgus monkeys and compared the results with human data. We found that in cynomolgus monkeys, as in humans, undifferentiated spermatogonia are largely quiescent, and the few engaged in the cell cycle were immunoreactive to GFRA1 antibodies. Moreover, we showed that PIWIL4+ spermatogonia, considered the most primitive undifferentiated spermatogonia in scRNA-seq studies, are quiescent in primates. We also described a novel subset of early differentiating spermatogonia, detectable from stage III to stage VII of the seminiferous epithelial cycle, that were transitioning from undifferentiated to differentiating spermatogonia, suggesting that the first generation of differentiating spermatogonia arises early during the epithelial cycle. Our study makes key advances in the current understanding of male germline premeiotic expansion in primates. [ABSTRACT FROM AUTHOR]

Species selection plays a pivotal part during non-clinical safety assessment in drug development. If possible, use of non-human primates (NHPs) should be avoided due to ethical considerations. However, limiting factors as lack of pharmacologic activity in other species could necessitate use of NHPs. LAI-PCSK9i is a bi-functional molecule combining a long-acting insulin analogue with a PCSK9 inhibitor peptide aiming to provide glycaemic control and to reduce plasma LDL concentrations. The NHP was chosen for the safety assessment of LAI-PCSK9i being the most relevant species with basal levels and plasma lipid composition closest to humans, while the dog and initially also the minipig were deemed irrelevant due to lack of pharmacologic activity on LDL-lowering and biological differences in lipid profiles. An in vivo tolerability and toxicokinetic study of LAI-PCSK9i in NHPs showed recurrent and severe hypoglycaemia at very low doses. Therefore, the minipig was re-evaluated and a follow-up study thoroughly assessing blood glucose and cholesterol levels and clinical signs illustrated that minipigs dosed with LAI-PCSK9i, tolerated the compound and LAI-PCSK9i decreased glucose and LDL over time. This work underlines that careful consideration is required when selecting species during safety assessment in drug development. The tolerability issue in NHPs led to the subsequent selection of the minipig for safety evaluation of LAI-PCSK9i although as a suboptimal alternative, which unexpectedly had a measurable pharmacologic response on LDL lowering. In conclusion, the NHPs may be unsuitable as test species for safety assessment of long-acting insulin analogues due to high sensitivity to recurring hypoglycaemic episodes. [ABSTRACT FROM AUTHOR]

Aim: To develop an obese, insulin‐resistant cynomolgus monkey model of non‐alcoholic steatohepatitis (NASH) with fibrosis with a high fat/high cholesterol (HFHC) diet (with or without high fructose) and test its responsiveness to caloric restriction or pioglitazone. Methods: First, two groups of monkeys (n = 24/group) with histologically proven NASH and fibrosis were fed the HFHC diet for 17 weeks. The treatment group was subjected to a 40% caloric restriction (CR) and had their diet switched from the HFHC diet to a chow diet (DSCR). Paired liver biopsies were taken before and 17 weeks after DSCR. Subsets of monkeys (nine/group) had whole liver fat content assessed by MRI. Next, two groups of monkeys with histologically proven NASH and fibrosis were treated with vehicle (n = 9) or pioglitazone (n = 20) over 24 weeks. Results: The HFHC and DSCR groups lost 0.9% and 11.4% of body weight, respectively. After 17 weeks, non‐alcoholic fatty liver disease activity score (NAS) improvement was observed in 66.7% of the DSCR group versus 12.5% of the HFHC group (P <.001). Hepatic fat was reduced to 5.2% in the DSCR group versus 23.0% in the HFHC group (P =.0001). After 24 weeks, NAS improvement was seen in 30% of the pioglitazone group versus 0% of the vehicle group (P =.08). Conclusions: Both weight loss induced by DSCR and treatment with pioglitazone improve the histological features of NASH in a diet‐induced cynomolgus monkey model. This model provides a translational preclinical model for testing novel NASH therapies. [ABSTRACT FROM AUTHOR]

Primate-specific genes (PSGs) tend to be expressed in the brain and testis. This phenomenon is consistent with brain evolution in primates but is seemingly contradictory to the similarity of spermatogenesis among mammals. Here, using whole-exome sequencing, we identified deleterious variants of X-linked SSX1 in six unrelated men with asthenoteratozoospermia. SSX1 is a PSG expressed predominantly in the testis, and the SSX family evolutionarily expanded independently in rodents and primates. As the mouse model could not be used for studying SSX1 , we used a non-human primate model and tree shrews, which are phylogenetically similar to primates, to knock down (KD) Ssx1 expression in the testes. Consistent with the phenotype observed in humans, both Ssx1- KD models exhibited a reduced sperm motility and abnormal sperm morphology. Further, RNA sequencing indicated that Ssx1 deficiency influenced multiple biological processes during spermatogenesis. Collectively, our experimental observations in humans and cynomolgus monkey and tree shrew models highlight the crucial role of SSX1 in spermatogenesis. Notably, three of the five couples who underwent intra-cytoplasmic sperm injection treatment achieved a successful pregnancy. This study provides important guidance for genetic counseling and clinical diagnosis and, significantly, describes the approaches for elucidating the functions of testis-enriched PSGs in spermatogenesis. The importance of primate-specific genes (PSGs) in spermatogenesis is largely unknown. This study identified variants in an X-linked PSG, SSX1 , associated with male infertility. By using cynomolgus monkey and tree shrew models, this study provides a powerful experimental system for elucidating the functions of testis-enriched PSGs in spermatogenesis. [ABSTRACT FROM AUTHOR]

Obesity is a metabolic disorder resulting from behavioral, environmental and heritable causes, and can have a negative impact on male reproduction. There have been few experiments in mice, rats, and rabbits on the effects of obesity on reproduction, which has inhibited the development of better treatments for male subfertility caused by obesity. Nonhuman primates are most similar to human beings in anatomy, physiology, metabolism, and biochemistry and are appropriate subjects for obesity studies. In this investigation, we conducted a transcriptome analysis of the testes of cynomolgus monkeys on high-fat, high-fructose, and cholesterol-rich diets to determine the effect of obesity on gene expression in testes. The results showed that the testes of obese monkeys had abnormal morphology, and their testes transcriptome was significantly different from that of non-obese animals. We identified 507 differentially abundant genes (adjusted p value < 0.01, log2 [FC] > 2) including 163 up-regulated and 344 down-regulated genes. Among the differentially abundant genes were ten regulatory genes, including IRF1, IRF6, HERC5, HERC6, IFIH1, IFIT2, IFIT5, IFI35, RSAD2, and UBQLNL. Gene ontology (GO) and KEGG pathway analysis was conducted, and we found that processes and pathways associated with the blood testes barrier (BTB), immunity, inflammation, and DNA methylation in gametes were preferentially enriched. We also found abnormal expression of genes related to infertility (TDRD5, CLCN2, MORC1, RFX8, SOHLH1, IL2RB, MCIDAS, ZPBP, NFIA, PTPN11, TSC22D3, MAPK6, PLCB1, DCUN1D1, LPIN1, and GATM) and down-regulation of testosterone in monkeys with dietetic obesity. This work not only provides an important reference for research and treatment on male infertility caused by obesity, but also valuable insights into the effects of diet on gene expression in testes. [ABSTRACT FROM AUTHOR]

Type II transmembrane serine proteases represent pharmacological targets for blocking entry and spread of influenza or coronaviruses. In this study, the depletion rates of the 3-amidinophenylalanine (3-APhA)-derived matriptase/TMPRSS2 inhibitors MI-463, MI-472, MI-485 or MI-1900 were determined by LC-MS/MS measurements over a period of 300 min using suspensions of rat, dog and cynomolgus monkey primary hepatocytes. From these in vitro pharmacokinetic (PK) experiments, intrinsic clearance values (Clint) were evaluated, and in vivo pharmacokinetic parameters (hepatic clearance, hepatic extraction ratio and bioavailability) were predicted. It was found that rat hepatocytes were the most active in the metabolism of 3-APhA derivatives (Clint 31.9–37.8 mL/min/kg), whereas dog and monkey cells displayed somewhat lower clearance of these compounds (Clint 6.6–26.7 mL/min/kg). These data support elucidation of important PK properties of anti-TMPRSS2/anti-matriptase 3-APhAs using mammalian hepatocyte models and thus contribute to the optimization of lead compounds. [ABSTRACT FROM AUTHOR]

We studied time course of pathological remodeling occurring in the cynomolgus monkey hearts against persistent atrioventricular block condition (n = 10). The atrioventricular block induced the ventricular and atrial dilation followed by the ventricular hypertrophy. Interstitial fibrosis in the ventricle was also observed along with gradual increases in the plasma angiotensin II and aldosterone concentrations. These adaptations were associated with the changes in gene expression profiling reflecting fibrosis and hypertrophy. Atrioventricular block reduced the ventricular rate and cardiac output, but the ejection fraction and stroke volume increased, whereas the cardiac output was gradually restored to its basal level. Systolic/diastolic blood pressure after the atrioventricular block was kept equal to or lower than that before the block, according with lack of increase in the plasma catecholamine levels. Chronic atrioventricular block gradually prolonged the QRS width and JT interval, leading to the QT interval prolongation in conscious state. 10 mg/kg of dl-sotalol hydrochloride induced torsade de pointes (TdP) in 6 out of 10 animals by 15 months. Animals showing longer QTcF under anesthesia after the atrioventricular block developed dl-sotalol-induced TdP earlier. No marked difference was observed in pharmacokinetics of dl-sotalol between 1 and 7 months after the atrioventricular block. Each TdP spontaneously terminated, reflecting a monkey’s relatively small “effective size of the heart (=∛(left ventricular weight)/wavelength of reentry)”. These fundamental knowledge will help better utilize the chronic atrioventricular block monkeys as an in vivo proarrhythmia model for detecting drug-induced TdP. [ABSTRACT FROM AUTHOR]

Advances in stem cell biology have facilitated cardiac regeneration, and many animal studies and several initial clinical trials have been conducted using human pluripotent stem cell-derived cardiomyocytes (PSC-CMs). Most preclinical and clinical studies have typically transplanted PSC-CMs via the following two distinct approaches: direct intramyocardial injection or epicardial delivery of engineered heart tissue. Both approaches present common disadvantages, including a mandatory thoracotomy and poor engraftment. Furthermore, a standard transplantation approach has yet to be established. In this study, we tested the feasibility of performing intracoronary administration of PSC-CMs based on a commonly used method of transplanting somatic stem cells. Six male cynomolgus monkeys underwent intracoronary administration of dispersed human PSC-CMs or PSC-CM aggregates, which are called cardiac spheroids, with multiple cell dosages. The recipient animals were sacrificed at 4 weeks post-transplantation for histological analysis. Intracoronary administration of dispersed human PSC-CMs in the cynomolgus monkeys did not lead to coronary embolism or graft survival. Although the transplanted cardiac spheroids became partially engrafted, they also induced scar formation due to cardiac ischemic injury. Cardiac engraftment and scar formation were reasonably consistent with the spheroid size or cell dosage. These findings indicate that intracoronary transplantation of PSC-CMs is an inefficient therapeutic approach. [Display omitted] • We tested the feasibility of intracoronary administration of PSC-CMs in primates. • This study is the first to show PSC-CM spheroids engrafted via coronary injection. • PSC-CM spheroids induced substantial scar formation due to coronary embolism. • Dispersed human iPSC-CMs resulted in neither coronary embolism nor graft survival. • Intracoronary injection of PSC-CMs was an inefficient approach for cardiac repair. [ABSTRACT FROM AUTHOR]

Objective By observing the impact of Bevacizumab injection on the speed of wound healing and the expression of CD34 in cynomolgus monkeys, to verify its delayed wound healing mechanism, and to provide clinical reference for the dosage and frequency of Bevacizumab treatment for oncology surgery patients. Methods Six male cynomolgus monkeys underwent full-thickness skin resection on the back to establish a wound healing evaluation model. Three cynomolgus monkeys were randomly selected as the saline group, and the other three cynomolgus monkeys were allocated to the Bevacizumab treatment group. The monkeys of Bevacizumab group were administrated with Bevacizumab injection (30 mg/kg, i.v.) on day 0, day 4, day 8, and day 11, respectively. The monkeys of saline group were injected with the same volume of saline as the group receiving Bevacizumab. The blood routine test was conducted, and the wound healing of each group of cynomolgus monkeys was observed 28 days after operation. The wound healing rate was analyzed by software Image J, and the severity of the wound was assessed by scoring method. Wound healing skin samples of 6 cynomolgus monkeys were obtained after 4 weeks, and the expression level of CD34 in the wound skin tissues was detected using immunohistochemistry. Results A skin excision wound model has already been established in cynomolgus monkeys. The number of white blood cells and neutrophils in the blood of cynomolgus monkeys increased significantly on the 3rd day after operation (P<0.05), and the number of white blood cells gradually returned to normal on the 7th day, while the number of red blood cells, hemoglobin and hematocrit did not change significantly, suggesting that the animals were in good nutritional status. The Bevacizumab injection group wound healing rate was significantly slower than that of the saline group on day 7, and day 28 (P<0.01, P<0.001). Microvessel density in the wound skin of Bevacizumab group appeared to be significantly lower than that of the saline group (P<0.01). Conclusion The injection of Bevacizumab may delay the wound healing by inhibiting the angiogenesis in the new skin tissue of the cynomolgus monkey. In clinical practice, the timing of giving a tumor patient a bevacizumab injection after surgery should be based on the pros and cons, and the duration, dosage, and frequency of the intervention should be chosen in a rational way. [ABSTRACT FROM AUTHOR]

Background Early post-implantation development, especially gastrulation in primates, is accompanied by extensive drastic chromatin reorganization, which remains largely elusive. Results To delineate the global chromatin landscape and understand the molecular dynamics during this period, a single-cell assay for transposase accessible chromatin sequencing (scATAC-seq) was applied to in vitro cultured cynomolgus monkey (Macaca fascicularis , hereafter referred to as monkey) embryos to investigate the chromatin status. First, we delineated the cis -regulatory interactions and identified the regulatory networks and critical transcription factors involved in the epiblast (EPI), hypoblast, and trophectoderm/trophoblast (TE) lineage specification. Second, we observed that the chromatin opening of some genome regions preceded the gene expression during EPI and trophoblast specification. Third, we identified the opposing roles of FGF and BMP signaling in pluripotency regulation during EPI specification. Finally, we revealed the similarity between EPI and TE in gene expression profiles and demonstrated that PATZ1 and NR2F2 were involved in EPI and trophoblast specification during monkey post-implantation development. Conclusions Our findings provide a useful resource and insights into dissecting the transcriptional regulatory machinery during primate post-implantation development. [ABSTRACT FROM AUTHOR]

Gnetum montanum Markgr. (Gnetaceae) is a commonly used traditional herbal medicine among the Yao ethnic group, with potential effects in preventing and treating tumors. However, the substance basis of its anti-tumor properties remains unclear. This study utilized ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC–Q–TOF–MS) to identify the chemical components of G. montanum extract (GME) and its absorbed prototypes in cynomolgus monkey plasma after oral administration. A total of 57 compounds were detected in the GME, with 14 compounds in positive ion mode and 43 compounds in negative ion mode. In the cynomolgus monkey plasma, 17 compounds were identified, with 3 compounds in positive ion mode and 14 compounds in negative ion mode. Subsequently, we utilized high content screening technology to investigate the anti-tumor effects of GME on colon cancer, lung cancer, breast cancer, gastric cancer, liver cancer, and esophageal cancer. We found that the GME exhibited significant proliferation inhibition on colon cancer cells SW480, with an IC 50 value of 50.77 μg/mL. Further research using component separation and pharmacological tracking revealed that the F2 component of the GME demonstrated notable anti-tumor effects. Through UPLC-MS identification, the chemical components in the F2 fraction were identified as pinoresinol diglucoside, (+)-pinoresinol-4-O-beta-D-glucopyranoside, ursolic acid, and gnetol. In conclusion, this study contributes to elucidating the anti-tumor pharmacological basis of GME and provides robust support for future drug design and development. • Identifying the absorption prototype substance of cynomolgus monkey plasma in G. montanum extract. • The G. montanum extract exhibits a significant anti-tumor effect. • Screening active substances through component separation and pharmacological tracking. • Revealing the anti-tumor mechanism of G. montanum extract is of great significance. [ABSTRACT FROM AUTHOR]

Controlling the immune response with only clinically approved immunosuppressant drugs is difficult in renal heterotransplantation from pigs to nonhuman primates. Moreover, to the best of our knowledge, no reports exist on the use of fetal pigs as kidney donors. This study aimed to compare the degree of transplant rejection between neonatal and fetal kidneys, with genetically unmodified pigs as donors and cynomolgus monkeys as recipients. The left kidneys of the recipient monkeys were removed, followed by transplantation of neonatal as well as fetal pig kidneys, which had undergone vascular anastomosis at the same site, into the retroperitoneum. Immunosuppression was performed with only US Food and Drug Administration-approved drugs. The fetal kidneys were transplanted into the omentum and para-aortic regions of cynomolgus monkeys. Consequently, the engraftment and development of the transplanted tissues were pathologically examined by sampling over time (twice in each experiment). An acute rejection was observed after a few weeks in neonatal renal grafts with vascular anastomosis. However, fetal pig kidneys were spared from rejection despite the administration of the same immunosuppressive protocol to the monkeys and the recipient blood vessels flowing into the fetal kidneys. The immunogenicity of fetal kidneys in pig–monkey renal heterotransplantation was lower than that of neonatal kidneys.

IntroductionPolycystic kidney disease (PKD) is a common autosomal dominant or recessive genetic disease, often accompanied by polycystic liver disease (PLD). Many cases of PKD in animals have been reported. However, little is known about the genes that cause PKD in animals.MethodsIn this study, we evaluated the clinical phenotypes of PKD in two spontaneously aged cynomolgus monkeys and explored the genetic etiology using whole-genome sequencing (WGS). Ultrasonic and histological consequences were further investigated in PKD- and PLD-affected monkeys.ResultsThe results indicated that the kidneys of the two monkeys had varying degrees of cystic changes, and the renal cortex was thinned and accompanied by fluid accumulation. As for hepatopathy, inflammatory cell infiltration, cystic effusion, steatosis of hepatocytes, and pseudo-lobular were found. Based on WGS results, the variants of PKD1:(XM_015442355: c.1144G>C p. E382Q) and GANAB: (NM_001285075.1: c.2708T>C/p. V903A) are predicted to be likely pathogenic heterozygous mutations in PKD- and PLD-affected monkeys.DiscussionOur study suggests that the cynomolgus monkey PKD and PLD phenotypes are very similar to those in humans, and are probably caused by pathogenic genes homologous to humans. The results indicate that cynomolgus monkeys can be used as the most appropriate animal model for human PKD pathogenesis research and therapeutic drug screening.

Background: Using untargeted metabolomics techniques, the goal of the study is to differentially screen serum and feces metabolite profiles of spontaneously diabetic and healthy cynomolgus monkeys, to explore potential serum and fecal biomarkers and analyze affected metabolic pathways. Methods: We adopted the diagnostic criteria for T2DM recommended by ADA for humans: FSG ≥7.0 mmol/L (126 mg/dl) and HbA1c ≥ 6.5%. The serum and feces samples from three diagnosed spontaneously T2DM cynomolgus monkeys and 11 age‐matched healthy controls were enrolled in the study. We employed LC–MS/MS‐based untargeted metabolomic methods to reveal the differential metabolite profiles of serum and feces samples between the two groups and to analyze the affected metabolic pathways in MetaboAnalyst 5.0 based on KEGG library. Results: Six and 44 differential metabolites were identified in serum and feces samples, respectively, and the corresponding affected commonly metabolic pathways involved several metabolic ways, such as arginine biosynthesis, pantothenate and CoA biosynthesis, alanine, aspartate and glutamate metabolism, valine, leucine and isoleucine biosynthesis, and histidine metabolism. Conclusion: The differential potential serum and feces biomarkers obtained from the LC–MS/MS based untargeted metabolomic may help to explain the potential pathophysiological mechanisms of T2DM and offer pivotal information for the early diagnosis and treatment of DM. [ABSTRACT FROM AUTHOR]

Immunotoxicity assessment is an important part of non-clinical safety evaluation of biotechnology-derived pharmaceuticals. The reference ranges of evaluated parameters, which depend on the sex, age and geographical origin of animals, play a significant role in interpreting the study results. The aim of this study was to determine the reference ranges of parameters commonly used for non-clinical immunotoxicity studies in cynomolgus monkeys (Macaca fascicularis) of different ages. The percentages of CD3+, CD4+, CD8+, CD20+, CD16/56+ lymphocytes, and the serum levels of immunoglobulins A, M, G and E in clinically healthy cynomolgus monkeys of both sexes and different ages (37–104 months) were analysed. The lymphocyte subsets were assessed by flow cytometry and serum immunoglobulin levels were determined by enzyme-linked immunosorbent assay. Statistical analysis showed a weak rank correlation of CD3+, CD20+ lymphocyte levels with the ages of female and male cynomolgus monkeys, and no significant correlation with age for the combined data of both sexes. The serum levels of immunoglobulins A, M, G and E did not have a significant rank correlation with age for the combined data of both sexes, whereas IgA levels in females and IgG levels in males were weakly correlated with age. Overall, these results justify the use of animals aged three to eight years for non-clinical immunotoxicity evaluation of biotechnology-derived pharmaceuticals. It is important to avoid the formation of animal groups with extreme ages (three years or eight years) in one group during randomization. [ABSTRACT FROM AUTHOR]

Although the new coronavirus disease 2019 (COVID-19) outbreak occurred in late 2019, it is still endemic worldwide, and has become a global public health problem. Vaccination against SARS-CoV-2 is considered to be the most effective intervention to prevent the spread of COVID-19. ZF2001 is a recombinant protein vaccine based on SARS-CoV-2 receptor-binding domain (RBD) subunit which contains aluminum adjuvant. In order to advance our research on ZF2001 into clinical trial, we investigated the general toxicity and immunogenicity of ZF2001 in cynomolgus monkeys and assessed the possible target organs for vaccine-induced toxicity. In the present research, we observed no significant systemic toxicities and abnormal cardiovascular and respiratory events following four times injections of intramuscular ZF2001 in cynomolgus monkeys. Histological examination revealed recoverable inflammatory changes in quadricep muscle and adjacent lymph node at the vaccine injection site. As expected, the vaccine can produce a strongly specific binding antibody and neutralizing antibodies in cynomolgus monkeys after inoculation. Taken together, our regulatory toxicology research proves the safety and immunogenicity of the ZF2001 vaccine, supporting its entry into large scale clinical trials. [ABSTRACT FROM AUTHOR]

Validation of risk-stratification method for the chronic atrioventricular block cynomolgus monkey model and its mechanistic interpretation was performed using 6 pharmacologically distinct drugs. The following drugs were orally administered in conscious state, astemizole: 1, 5, and 10 mg/kg (n  = 6); haloperidol: 1, 10, and 30 mg/kg (n  = 5); amiodarone: 30 mg/kg (n  = 4); famotidine: 10 mg/kg (n  = 4); levofloxacin: 100 mg/kg (n  = 4); and tolterodine: 0.2, 1, and 4.5 mg/kg (n  = 4). Astemizole of 5 and 10 mg/kg significantly prolonged ΔΔQTcF, whereas no significant change was observed by the others. Torsade de pointes (TdP) was induced by astemizole of 5 and 10 mg/kg in 3/6 and 6/6, and by haloperidol of 10 and 30 mg/kg in 1/5 and 1/5, respectively, which was not observed in the others. Torsadogenic risk of the drugs was quantified using the criteria for the monkey model specified in our previous study. Namely, high-risk drugs induced TdP at ≤ 3 times of their maximum clinical daily dose. Intermediate-risk drugs did not induce TdP at this dose range, but induced it at higher doses. Low/no-risk drugs never induced TdP at any dose tested. The magnitude of risk was intermediate for astemizole and haloperidol, and low/no risk for the others. The prespecified, risk-stratification method for the monkey model may solve the issue existing between nonclinical models and patients with labile repolarization, which can reinforce the regulatory decision-making and labeling at time of marketing application of nondouble-negative drug candidate (hERG assay positive and/or in vivo QT study positive). [ABSTRACT FROM AUTHOR]

The use of stereotaxic brain templates in automatic neuroimage analysis has become the predominant technology. However, in the context of experiment studies involving cynomolgus monkeys, there is a lack of adequate brain templates. This gap poses a challenge to effectively utilize cynomolgus monkeys in neuroscience research. Considering the dynamic changes in structural morphology and functional maturity during development and aging, particularly post-birth, improving the accuracy of image data analysis necessitates the construction of a series of stereotaxic brain templates spanning all age groups. Therefore, this study developed a comprehensive set of stereotaxic templates for development cynomolgus brain with fine substructure delineations, including T1WI MRI structural template, tissue probability map (TPM), [ 18 F]FDG PET functional template aligned in a standardized space. This template set enables precise localization and quantification of cynomolgus monkey brain tissue across different developmental stages. Additionally, it could also be used for automatic individualized substructure segmentation. By automatically generating multiple precision subregion segmentations within individual spaces, it preserves the integrity of individual data with high precision and repeatability. Our constructed cynomolgus template sets can further facilitate the accurate study of brain structure and glucose metabolism function in non-human being studies., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier Inc.)