Primate Studies Research Articles

ProMENDA: an updated resource for proteomic and metabolomic characterization in depression

Depression is a prevalent mental disorder with a complex biological mechanism. Following the rapid development of systems biology technology, a growing number of studies have applied proteomics and metabolomics to explore the molecular profiles of depression. However, a standardized resource facilitating the identification and annotation of the available knowledge from these scattered studies associated with depression is currently lacking. This study presents ProMENDA, an upgraded resource that provides a platform for manual annotation of candidate proteins and metabolites linked to depression. Following the establishment of the protein dataset and the update of the metabolite dataset, the ProMENDA database was developed as a major extension of its initial release. A multi-faceted annotation scheme was employed to provide comprehensive knowledge of the molecules and studies. A new web interface was also developed to improve the user experience. The ProMENDA database now contains 43,366 molecular entries, comprising 20,847 protein entries and 22,519 metabolite entries, which were manually curated from 1370 human, rat, mouse, and non-human primate studies. This represents a significant increase (more than 7-fold) in molecular entries compared to the initial release. To demonstrate the usage of ProMENDA, a case study identifying consistently reported proteins and metabolites in the brains of animal models of depression was presented. Overall, ProMENDA is a comprehensive resource that offers a panoramic view of proteomic and metabolomic knowledge in depression. ProMENDA is freely available at https://menda.cqmu.edu.cn.

From lab bench to hope: a review of gene therapies in clinical trials for Parkinson's disease and challenges.

Parkinson's disease (PD) is a chronic neurological disorder that is identified by a characteristic combination of symptoms such as bradykinesia, resting tremor, rigidity, and postural instability. It is the second most common neurodegenerative disease after Alzheimer's disease and is characterized by the progressive loss of dopamine-producing neurons in the brain. Currently, available treatments for PD are symptomatic and do not prevent the disease pathology. There is growing interest in developing disease-modifying therapy that can reduce disease progression and improve patients' quality of life. One of the promising therapeutic approaches under evaluation is gene therapy utilizing a viral vector, adeno-associated virus (AAV), to deliver transgene of interest into the central nervous system (CNS). Preclinical studies in small animals and nonhuman primates model of PD have shown promising results utilizing the gene therapy that express glial cell line-derived neurotrophic factor (GDNF), cerebral dopamine neurotrophic factor (CDNF), aromatic L-amino acid decarboxylase (AADC), and glutamic acid decarboxylase (GAD). This study provides a comprehensive review of the current state of the above-mentioned gene therapies in various phases of clinical trials for PD treatment. We have highlighted the rationale for the gene-therapy approach and the findings from the preclinical and nonhuman primates studies, evaluating the therapeutic effect, dose safety, and tolerability. The challenges associated with gene therapy for heterogeneous neurodegenerative diseases, such as PD, have also been described. In conclusion, the review identifies the ongoing promising gene therapy approaches in clinical trials and provides hope for patients with PD.

Is retrograde menstruation a universal, recurrent, physiological phenomenon? A systematic review of the evidence in humans and non-human primates.

What are the quantitative, qualitative, and temporal patterns of retrograde mentruation? The extreme quantitative and qualitative heterogeneity of the available studies prevents the definitive conclusion that retrograde menstruation is a universal and consistent phenomenon during the reproductive period. Retrograde menstruation has been defined as a universal, physiological phenomenon that occurs similarly in about 90% of menstruators during the reproductive period. However, uncertainties still exist in terms of the event frequency, total amount, and cellular composition of retrograde menstruation and the differences between individuals with versus those without endometriosis. Two systematic reviews were performed, one for human studies, and one for non-human primate studies. We retrieved studies from the PubMed and Embase databases published between 1 January 1980 and 1 November 2023. Studies published in the English language were included and identified using a combination of MeSH terms. References from relevant publications were systematically screened and further articles were identified using PubMed's 'similar articles' and 'cited by' functions. Results were reported in accordance with the PRISMA guidelines. Studies that did not report original data or provided a review of the field were excluded. Bias analysis was completed for each included human study by using the Newcastle-Ottawa scoring system. Fifteen studies were finally included in the human systematic review, mostly with limited sample sizes. The macroscopic visualization of blood in PF during menses was reported with a frequency ranging from 9% to 100%. A prevalence of endometrial cells detected in peritoneal fluid ranging from 8% to 75% was reported in the various studies. Controversial findings were reported in relation to patients with endometriosis. Retrograde menstruation has been evaluated cross-sectionally on single occasions, and no information is available on the course of the phenomenon within an entire cycle and between subsequent cycles. Two studies were included in the non-human primate systematic review; one of them showed that retrograde menstruation was observed more frequently in baboons with naturally occurring endometriosis (83%) than in those with a normal pelvis (51%). In humans, peritoneal fluid has often been collected at different cycle phases and not systematically during menstruation. The indication for laparoscopy was not always clear for all participants. A wide variety of methods were used to detect endometrial cells, including cytological staining, cell block analysis, immunocytochemistry, and various methods of cell culture. The idea that almost all women experience retrograde menstruation regularly and similarly during their reproductive life is currently unsubstantiated. It is an academic notion accepted uncritically. Development of endometriosis may derive from differences in the frequency or severity of the event. The review was partially funded by Italian Ministry of Health-Current Research IRCCS. P.Vi. serves as co-editor in Chief of Journal of Endometriosis and Uterine Disorders. E.S. serves as Editor in Chief of Human Reproduction Open and discloses research grants from Ferring, Ibsa, Gedeon Richter, and Theramex, and honoraria from Ibsa and Gedeon Richter. P.Ve. serves as Associate Editor for Human Reproduction Open; is a member of the Editorial Board of the Journal of Obstetrics and Gynaecology Canada, of the Italian Journal of Obstetrics and Gynaecology, and of the International Editorial Board of Acta Obstetricia et Gynecologica Scandinavica; has received royalties from Wolters Kluwer for chapters on endometriosis management in the clinical decision support resource UpToDate; and maintains both a public and private gynecological practice. All other authors declare they have no conflict of interest. N/A.

First noncontact millimeter-wave radar measurement of heart rate in great apes: Validation in chimpanzees (Pan troglodytes).

Heart rate is a crucial vital sign and a valuable indicator for assessing the physical and psychological condition of a target animal. Heart rate contributes to (1) fundamental information for cognitive research, (2) an indicator of psychological and physical stress, and (3) improving the animal welfare of captive animals, especially in nonhuman primate studies. Heart rate has been measured using a contact-type device; however, the device burdens the target animals and that there are risks associated with anesthesia during installation. This study explores the application of heartbeat measurement techniques using millimeter-wave radar, primarily developed for humans, as a remote and noninvasive method for measuring the heart rate of nonhuman primates. Through a measurement test conducted on two chimpanzees, we observed a remarkable correspondence between the peak frequency spectrum of heart rate estimated using millimeter-wave radar and the mean value obtained from electrocardiograph data, thereby validating the accuracy of the method. To the best of our knowledge, this is the first demonstration of the precise measurement of great apes' heart rate using millimeter-wave radar technology. Compared to heart rate measurement using video analysis, the method using millimeter-wave radar has the advantage that it is less susceptible to weather and lighting conditions and that measurement techniques for multiple individuals have been developed for human subjects, while its disadvantage is that validation of measurement from long distances has not been completed. Another disadvantage common to both methods is that measurement becomes difficult when the movement of the target individual is large. The possibility of noncontact measurement of heart rate in wild and captive primates will undoubtedly open up a new research area while taking animal welfare into consideration.

Stabilized trimeric peptide immunogens of the complete HIV-1 gp41 N-heptad repeat and their use as HIV-1 vaccine candidates

Efforts to develop an HIV-1 vaccine include those focusing on conserved structural elements as the target of broadly neutralizing monoclonal antibodies. MAb D5 binds to a highly conserved hydrophobic pocket on the gp41 N-heptad repeat (NHR) coiled coil and neutralizes through prevention of viral fusion and entry. Assessment of 17-mer and 36-mer NHR peptides presenting the D5 epitope in rodent immunogenicity studies showed that the longer peptide elicited higher titers of neutralizing antibodies, suggesting that neutralizing epitopes outside of the D5 pocket may exist. Although the magnitude and breadth of neutralization elicited by NHR-targeting antigens are lower than that observed for antibodies directed to other epitopes on the envelope glycoprotein complex, it has been shown that NHR-directed antibodies are potentiated in TZM-bl cells containing the FcγRI receptor. Herein, we report the design and evaluation of covalently stabilized trimeric 51-mer peptides encompassing the complete gp41 NHR. We demonstrate that these peptide trimers function as effective antiviral entry inhibitors and retain the ability to present the D5 epitope. We further demonstrate in rodent and nonhuman primate immunization studies that our 51-mer constructs elicit a broader repertoire of neutralizing antibody and improved cross-clade neutralization of primary HIV-1 isolates relative to 17-mer and 36-mer NHR peptides in A3R5 and FcγR1-enhanced TZM-bl assays. These results demonstrate that sensitive neutralization assays can be used for structural enhancement of moderately potent neutralizing epitopes. Finally, we present expanded trimeric peptide designs which include unique low-molecular-weight scaffolds that provide versatility in our immunogen presentation strategy.

Pharmacokinetics and safety of LEAD-452, an EGFR-specific 4-1BB-agonistic trimerbody in non-human primates

LEAD-452 is a humanized bispecific EGFR-targeted 4-1BB-agonistic trimerbody with a unique trimeric configuration compared to other 4-1BB-specific antibodies that are currently in development. Indeed, enhanced tumor-specific costimulation and very remarkable safety and efficacy profiles have been observed in mouse models. Here, we conducted for the first time a preclinical pharmacokinetic and toxicity study in non-human primates (NHP) (Macaca fascicularis). LEAD-452 exhibits comparable binding affinity for human and macaque targets, indicating its pharmacological significance for safety testing across species. The NHP were administered LEAD-452 in a series of ascending doses, ranging from 0.1 mg/kg to 10 mg/kg, and repeated doses up to 20 mg/kg. The administration of LEAD-452 was found to be clinically well tolerated, with no major related adverse effects observed. Furthermore, there have been no reported cases of liver toxicity, thrombocytopenia, and neutropenia, which are commonly associated with treatments using conventional anti-4-1BB IgG-based antibodies. In addition, neither IgM nor IgG-based anti-drug antibodies were detected in serum samples from NHP during the study, regardless of the dose of LEAD-452 administered. These results support the clinical development of LEAD-452 for the treatment of solid tumors.

Preliminary investigation into long-term stress by isolated captivity-related changes of reproduction hormones in Cynomolgus monkey.

Stress profoundly affects physical and emotional well-being, extending its physiological influence to the female menstrual cycle, impeding the hypothalamus-pituitary-gonadal (HPG) axis, and affecting fertility by suppressing sex-stimulating hormones. In this study, we meticulously analyzed menstrual cycles and corresponding hormonal fluctuations in three female Cynomolgus monkeys. The preliminary findings indicated lower-than-normal levels of cortisol, follicle-stimulating hormone (FSH), and estradiol. Anovulatory bleeding occurred in one monkey, which could be linked to stress. In contrast to cortisol, alkaline phosphatase (ALP), which is correlated to cortisol levels, was consistently elevated in menstruating monkeys, suggesting its potential as a stress indicator. The non-menstruating group exhibited stress-related weight loss, emphasizing the observed ALP trends. Non-menstruating monkeys may experience more stress than menstruating monkeys. The implications of this study extend beyond the confines of primate studies and offer a valuable method for enhancing the welfare of female Cynomolgus monkeys.

Integrative multi-omics profiling in human decedents receiving pig heart xenografts.

In a previous study, heart xenografts from 10-gene-edited pigs transplanted into two human decedents did not show evidence of acute-onset cellular- or antibody-mediated rejection. Here, to better understand the detailed molecular landscape following xenotransplantation, we carried out bulk and single-cell transcriptomics, lipidomics, proteomics and metabolomics on blood samples obtained from the transplanted decedents every 6 h, as well as histological and transcriptomic tissue profiling. We observed substantial early immune responses in peripheral blood mononuclear cells and xenograft tissue obtained from decedent 1 (male), associated with downstream T cell and natural killer cell activity. Longitudinal analyses indicated the presence of ischemia reperfusion injury, exacerbated by inadequate immunosuppression of T cells, consistent with previous findings of perioperative cardiac xenograft dysfunction in pig-to-nonhuman primate studies. Moreover, at 42 h after transplantation, substantial alterations in cellular metabolism and liver-damage pathways occurred, correlating with profound organ-wide physiological dysfunction. By contrast, relatively minor changes in RNA, protein, lipid and metabolism profiles were observed in decedent 2 (female) as compared to decedent 1. Overall, these multi-omics analyses delineate distinct responses to cardiac xenotransplantation in the two human decedents and reveal new insights into early molecular and immune responses after xenotransplantation. These findings may aid in the development of targeted therapeutic approaches to limit ischemia reperfusion injury-related phenotypes and improve outcomes.

Hyperphosphorylated tau in Alzheimer's disease disseminates along pathways predicted by the Structural Model for Cortico-cortical Connections.

In Alzheimer´s disease (AD), hyperphosphorylated tau spreads along the cerebral cortex in a stereotypical pattern that parallels cognitive deterioration. Tau seems to spread transsynaptically along cortico-cotical pathways that, according to synaptic tract-tracing studies in nonhuman primates, have specific laminar patterns related to the cortical type of the connected areas. This relation is described in the Structural Model. In the present article, we study the laminar distribution of hyperphosphorylated tau, labeled with the antibody AT8, along temporal cortical types in postmortem human brains with different AD stages to test the predictions of the Structural Model. Brains from donors without dementia had scant AT8-immunorreactive (AT8-ir) neurons in allo-, meso-, and isocortical types. In early AD stages, the mesocortical dysgranular type, including part of the transentorhinal cortex, had the highest AT8 immunostaining and AT8-ir neurons density. In advanced AD stages, AT8 immunostaining increased along the isocortical types until reaching the auditory koniocortex. Regarding laminar patterns, in early AD stages there were more AT8-ir neurons in supragranular layers in each de novo involved neocortical type; in advanced AD stages, AT8-ir neurons were equally distributed in supra- and infragranular layers. These AT8-ir laminar patterns are compatible with the predictions of the Structural Model. In summary, we show that hyperphosphorylated tau initially accumulates in allo-, meso-, and isocortical types, offer a proof of concept for the validity of the Structural Model to predict synaptic pathway organization in the human cerebral cortex, and highlight the relevance of nonhuman primate tract-tracing studies to understand human neuropathology.

Adaptations to maternal WSD-feeding and metformin use on fetal islets from non-human primate offspring

Given the current increased prevalence of diabetes in pregnancy, the potential relative risks and benefits of oral glycemic agents on the developing offspring should be evaluated and considered. Metformin (Met) is a common oral glycemic agent prescribed in pregnancy that is not metabolized, but is renally excreted and actively transported across the placenta. Thus, the developing fetus is exposed to near equivalent maternal adult doses with subsequent risk of adverse consequences. Using a non-human primate model of maternal overnutrition, our group previously demonstrated that developmental exposure to a Western-style diet (WSD) results in an inappropriate increase in glucose-stimulated insulin secretion and changes in expression of several ion channels involved in insulin secretion in islets from juvenile offspring. Given that maternal overnutrition results in an excess energy supply to the developing fetus while Met inhibits mitochondrial respiration, we hypothesize that maternal WSD-feeding and Met will result in maladaptive beta-cell function in the fetus. Static insulin secretion assays on islets from chow (CD) and WSD +/- Met-exposed offspring show no differences in glucose-stimulated insulin secretion. Nevertheless, islets from a mWSD+Met-exposed fetus showed lower oxygen consumption and extracellular acidification rate relative to mWSD+placebo fetal islets, suggesting that maternal diet influences fetal programming and adaptation to Met in fetal islets. Therefore, we predict that the metabolic response to intrauterine Met exposure differs depending on the maternal diet. Elucidation of the mechanisms mediating adaptations to maternal diet and Met exposure during development may provide insight on interventions to avoid beta-cell failure in the offspring as they age. All nonhuman primate studies were supported in part by the Oregon National Primate Research Center grant P51 OD011092 from the National Institutes of Health/Offce of the Director. D.T.C. was supported by the Vanderbilt University Training Program in Molecular Endocrinology (5T32 DK563-30) and a predoctoral fellowship from the NIH/NIDDK (1F31 DK135164).T.A.D., C.E.M., K.M.A., J.E.F., P.K., and M.G. were supported by NIH/NIDDK Grants R24 DK090964 and R01 DK128187. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Common anesthetic used in preclinical PET imaging inhibits metabolism of the PET tracer [18F]3F4AP.

Positron emission tomography (PET) imaging studies in laboratory animals are almost always performed under isoflurane anesthesia to ensure that the subject stays still during the image acquisition. Isoflurane is effective, safe, and easy to use, and it is generally assumed to not have an impact on the imaging results. Motivated by marked differences observed in the brain uptake and metabolism of the PET tracer 3-[18F]fluoro-4-aminopyridine [(18F]3F4AP) between human and nonhuman primate studies, this study investigates the possible effect of isoflurane on this process. Mice received [18F]3F4AP injection while awake or under anesthesia and the tracer brain uptake and metabolism was compared between groups. A separate group of mice received the known cytochrome P450 2E1 inhibitor disulfiram prior to tracer administration. Isoflurane was found to largely abolish tracer metabolism in mice (74.8 ± 1.6 vs. 17.7 ± 1.7% plasma parent fraction, % PF) resulting in a 4.0-fold higher brain uptake in anesthetized mice at 35 min post-radiotracer administration. Similar to anesthetized mice, animals that received disulfiram showed reduced metabolism (50.0 ± 6.9% PF) and a 2.2-fold higher brain signal than control mice. The higher brain uptake and lower metabolism of [18F]3F4AP observed in anesthetized mice compared to awake mice are attributed to isoflurane's interference in the CYP2E1-mediated breakdown of the tracer, which was confirmed by reproducing the effect upon treatment with the known CYP2E1 inhibitor disulfiram. These findings underscore the critical need to examine the effect of isoflurane in PET imaging studies before translating tracers to humans that will be scanned without anesthesia.

Echocardiographic characterization of age- and sex-associated differences in cardiac function and morphometry in nonhuman primates.

Aging per se is a major risk factor for cardiovascular diseases and is associated with progressive changes in cardiac structure and function. Rodent models are commonly used to study cardiac aging, but do not closely mirror differences as they occur in humans. Therefore, we performed a 2D echocardiographic study in non-human primates (NHP) to establish age- and sex-associated differences in cardiac function and morphometry in this animal model. M mode and 2D echocardiography and Doppler analyses were performed cross-sectionally in 38 healthy rhesus monkeys (20 females and 18 males), both young (age 7-12years; n = 20) and old (age 19-30years; n = 18). The diameters of the cardiac chambers did not differ significantly by age group, but males had larger left ventricular diameters (2.43 vs 2.06cm in diastole and 1.91 vs 1.49cm in systole, p = 0.0004 and p = 0.0001, respectively) and left atrial diameter (1.981 vs 1.732cm; p = 0.0101). Left ventricular mass/body surface area did not vary significantly with age and sex. Ejection fraction did not differ by age and females presented a higher ejection fraction than males (54.0 vs 50.8%, p = 0.0237). Diastolic function, defined by early to late mitral peak flow velocity ratio (E/A), was significantly lower in old rhesus monkeys (2.31 vs 1.43, p = 0.0020) and was lower in females compared to males (1.595 vs 2.230, p = 0.0406). Right ventricular function, evaluated by measuring the Tricuspid Annular Plane Systolic Excursion, did not differ by age or sex, and Right Ventricular Free Wall Longitudinal Strain, did not differ with age but was lower in males than in females (-22.21 vs -17.95%, p = 0.0059). This is the first echocardiographic study to evaluate age- and sex-associated changes of cardiac morphometry and function in young and old NHP. The findings of this work will provide a reference to examine the effect of age and sex on cardiac diseases in NHP.

Chst9 marks a spatially and transcriptionally unique population of Oprm1-expressing neurons in the nucleus accumbens

Opioids produce addictive, analgesic, and euphoric effects via actions at mu opioid receptors (µORs). The µOR is encoded by the Oprm1 gene and is expressed in multiple brain regions that regulate reward and motivation, such as the nucleus accumbens (NAc). Oprm1 expression in NAc medium spiny neurons (MSNs) mediates opioid place preference, seeking, and consumption. However, recent single nucleus RNA sequencing (snRNA-seq) studies have revealed that multiple subpopulations of NAc neurons express Oprm1 mRNA, making it unclear which populations mediate diverse behaviors resulting from µOR activation. Using published snRNA-seq datasets from the rat NAc, we identified a novel population of MSNs that express the highest levels of Oprm1 of any NAc cell type. Here, we show that this population is selectively marked by expression of Chst9, a gene encoding a carbohydrate sulfotransferase. Notably, Chst9+ neurons exhibited more abundant expression of Oprm1 as compared to other cell types, and formed discrete cellular clusters along the medial and ventral borders of the NAc shell subregion. Moreover, CHST9 mRNA was also found to mark specific MSN populations in published human and primate snRNA-seq studies, indicating that this unique population may be conserved across species. Together, these results identify a spatially and transcriptionally distinct NAc neuron population characterized by the expression of Chst9. The abundant expression of Oprm1 in this population and the conservation of these cells across species suggests that they may play a key functional role in opioid response and identify this subpopulation as a target for further investigation.

A pig kidney supporting human physiology

Because of the global shortage of donor kidneys, xenotransplantation emerges as a potential solution for individuals with kidney failure who face challenges in securing a suitable donor kidney. A study featured in this month's issue of Kidney International assesses the kidney physiology of a porcine kidney transplanted into a brain-dead human with kidney failure, demonstrating life-sustaining physiological function for 7 days. Together with preclinical nonhuman primate studies, decedent models provide complementary data for development of clinical kidney xenotransplantation.

Two-photon autofluorescence lifetime assay of rabbit photoreceptors and retinal pigment epithelium during light-dark visual cycles in rabbit retina

Two-photon excited fluorescence (TPEF) is a powerful technique that enables the examination of intrinsic retinal fluorophores involved in cellular metabolism and the visual cycle. Although previous intensity-based TPEF studies in non-human primates have successfully imaged several classes of retinal cells and elucidated aspects of both rod and cone photoreceptor function, fluorescence lifetime imaging (FLIM) of the retinal cells under light-dark visual cycle has yet to be fully exploited. Here we demonstrate a FLIM assay of photoreceptors and retinal pigment epithelium (RPE) that reveals key insights into retinal physiology and adaptation. We found that photoreceptor fluorescence lifetimes increase and decrease in sync with light and dark exposure, respectively. This is likely due to changes in all-trans-retinol and all-trans-retinal levels in the outer segments, mediated by phototransduction and visual cycle activity. During light exposure, RPE fluorescence lifetime was observed to increase steadily over time, as a result of all-trans-retinol accumulation during the visual cycle and decreasing metabolism caused by the lack of normal perfusion of the sample. Our system can measure the fluorescence lifetime of intrinsic retinal fluorophores on a cellular scale, revealing differences in lifetime between retinal cell classes under different conditions of light and dark exposure.

Visual perception of highly memorable images is mediated by a distributed network of ventral visual regions that enable a late memorability response.

Behavioral and neuroscience studies in humans and primates have shown that memorability is an intrinsic property of an image that predicts its strength of encoding into and retrieval from memory. While previous work has independently probed when or where this memorability effect may occur in the human brain, a description of its spatiotemporal dynamics is missing. Here, we used representational similarity analysis (RSA) to combine functional magnetic resonance imaging (fMRI) with source-estimated magnetoencephalography (MEG) to simultaneously measure when and where the human cortex is sensitive to differences in image memorability. Results reveal that visual perception of High Memorable images, compared to Low Memorable images, recruits a set of regions of interest (ROIs) distributed throughout the ventral visual cortex: a late memorability response (from around 300 ms) in early visual cortex (EVC), inferior temporal cortex, lateral occipital cortex, fusiform gyrus, and banks of the superior temporal sulcus. Image memorability magnitude results are represented after high-level feature processing in visual regions and reflected in classical memory regions in the medial temporal lobe (MTL). Our results present, to our knowledge, the first unified spatiotemporal account of visual memorability effect across the human cortex, further supporting the levels-of-processing theory of perception and memory.

Differential induction of donor-reactive Foxp3+ regulatory T cell via blockade of CD154 vs CD40

Recently published studies in both murine models and a meta-analysis of non-human primate renal transplant studies showed that anti-CD154 reagents conferred a significant survival advantage over CD40 blockers in both animal models and across multiple organs. Here we sought to compare the induction of donor-reactive forkhead box P3+-induced regulatory T cells (Foxp3+ iTreg) in mice treated with anti-CD154 versus anti-CD40 monoclonal antibodies (mAbs). Results indicated that while treatment with anti-CD154 mAb resulted in a significant increase in the frequency of donor-reactive CD4+ Foxp3+ iTreg following transplantation, treatment with anti-CD40 or Cd40 deficiency failed to recapitulate this result. Because we recently identified CD11b as an alternate receptor for CD154 during alloimmunity, we interrogated the role of CD154:CD11b interactions in the generation of Foxp3+ iTreg and found that blockade of CD11b in Cd40−/− recipients resulted in increased donor-reactive Foxp3+ iTreg as compared with CD40 deficiency alone. Mechanistically, CD154:CD11b inhibition decreased interleukin (IL)-1β from CD11b+ and CD11c+ dendritic cells, and blockade of IL-1β synergized with CD40 deficiency to promote Foxp3+ iTreg induction and prolong allograft survival. Taken together, these data provide a mechanistic basis for the observed inferiority of anti-CD40 blockers as compared with anti-CD154 mAb and illuminate an IL-1β-dependent mechanism by which CD154:CD11b interactions prevent the generation of donor-reactive Foxp3+ iTreg during transplantation.

Sitagliptin elevates plasma and CSF incretin levels following oral administration to nonhuman primates: relevance for neurodegenerative disorders

The endogenous incretins glucagon-like peptide-1 (GLP-1) and glucose-dependent insulinotropic polypeptide (GIP) possess neurotrophic, neuroprotective, and anti-neuroinflammatory actions. The dipeptidyl peptidase 4 (DPP-4) inhibitor sitagliptin reduces degradation of endogenous GLP-1 and GIP, and, thereby, extends the circulation of these protective peptides. The current nonhuman primate (NHP) study evaluates whether human translational sitagliptin doses can elevate systemic and central nervous system (CNS) levels of GLP-1/GIP in naive, non-lesioned NHPs, in line with our prior rodent studies that demonstrated sitagliptin efficacy in preclinical models of Parkinson’s disease (PD). PD is an age-associated neurodegenerative disorder whose current treatment is inadequate. Repositioning of the well-tolerated and efficacious diabetes drug sitagliptin provides a rapid approach to add to the therapeutic armamentarium for PD. The pharmacokinetics and pharmacodynamics of 3 oral sitagliptin doses (5, 20, and 100 mg/kg), equivalent to the routine clinical dose, a tolerated higher clinical dose and a maximal dose in monkey, were evaluated. Peak plasma sitagliptin levels were aligned both with prior reports in humans administered equivalent doses and with those in rodents demonstrating reduction of PD associated neurodegeneration. Although CNS uptake of sitagliptin was low (cerebrospinal fluid (CSF)/plasma ratio 0.01), both plasma and CSF concentrations of GLP-1/GIP were elevated in line with efficacy in prior rodent PD studies. Additional cellular studies evaluating human SH-SY5Y and primary rat ventral mesencephalic cultures challenged with 6-hydroxydopamine, established cellular models of PD, demonstrated that joint treatment with GLP-1 + GIP mitigated cell death, particularly when combined with DPP-4 inhibition to maintain incretin levels. In conclusion, this study provides a supportive translational step towards the clinical evaluation of sitagliptin in PD and other neurodegenerative disorders for which aging, similarly, is the greatest risk factor.

Disruption of awake sharp-wave ripples does not affect memorization of locations in repeated-acquisition spatial memory tasks.

Storing and accessing memories is required to successfully perform day-to-day tasks, for example for engaging in a meaningful conversation. Previous studies in both rodents and primates have correlated hippocampal cellular activity with behavioral expression of memory. A key role has been attributed to awake hippocampal replay - a sequential reactivation of neurons representing a trajectory through space. However, it is unclear if awake replay impacts immediate future behavior, gradually creates and stabilizes long-term memories over a long period of time (hours and longer), or enables the temporary memorization of relevant events at an intermediate time scale (seconds to minutes). In this study, we aimed to address the uncertainty around the timeframe of impact of awake replay by collecting causal evidence from behaving rats. We detected and disrupted sharp wave ripples (SWRs) - signatures of putative replay events - using electrical stimulation of the ventral hippocampal commissure in rats that were trained on three different spatial memory tasks. In each task, rats were required to memorize a new set of locations in each trial or each daily session. Interestingly, the rats performed equally well with or without SWR disruptions. These data suggest that awake SWRs - and potentially replay - does not affect the immediate behavior nor the temporary memorization of relevant events at a short timescale that are required to successfully perform the spatial tasks. Based on these results, we hypothesize that the impact of awake replay on memory and behavior is long-term and cumulative over time.

Abstract 1897: Preclinical development of MGC028, an ADAM9-targeted, glycan-linked, exatecan-based antibody-drug conjugate for the treatment of solid cancers

Abstract Introduction: Antibody-drug conjugates (ADCs) seek to increase the therapeutic window of potent cytotoxic agents by linking them with monoclonal antibodies (mAbs) to selectively deliver cytotoxic payloads to tumor cells. The effectiveness and safety of ADCs rely on both the mAb specificity, and the linker-payload employed. Several approved ADCs using microtubule inhibitor payloads are impacted by ocular adverse events that have been observed both preclinically and in patients. A class of linker-payloads incorporating topoisomerase 1 inhibitors (TOP1i) has emerged recently as an effective alternative to tubulin inhibitor-based ADCs. To date, TOP1i ADCs have not been associated with dose-limiting ocular toxicity seen with microtubule inhibitor payloads. Here we report the preclinical development of an ADAM9 (a disintegrin and metalloprotease domain 9)-targeted ADC that incorporates a novel glycan-linked TOP1i. ADAM9, a member of the ADAM family of multifunctional type 1 transmembrane proteins, plays a role in tumorigenesis and cancer progression and is overexpressed in multiple cancers, making it an attractive target for cancer treatment. Methods: MGC028 incorporates the cleavable linker-payload, bicyclononyne carbamoyl sulfamide Val-Ala-PABC exatecan (SYNtecan E™), site-specifically conjugated at asparagine 297 of the heavy chain through enzymatic glycan remodeling and metal-free click chemistry using Synaffix’s GlycoConnect™ technology. In vivo efficacy studies were performed in immunodeficient mice with ADAM9-expressing human tumor cell-line (CDX) or patient-derived (PDX) xenografts. A non-human primate toxicology study was conducted in which MGC028 was administered by 15-minute IV infusion every two weeks at dose levels of 22.5 and 55 mg/kg for a total of two doses. Results: MGC028 exhibited specific, dose-dependent in vivo antitumor activity toward ADAM9-positive CDX models representing gastric, lung, pancreatic and colorectal cancer, and head and neck squamous cell carcinoma. Furthermore, MGC028 demonstrated antitumor activity toward ADAM9-positive PDX models of lung and pancreatic cancer, and cholangiocarcinoma. MGC028 was well tolerated in a repeat-dose non-human primate toxicology study, up to 55 mg/kg, the highest dose level tested. Observations were limited to mild, reversible increases in liver enzymes, without microscopic correlates, and decreased lymphoid cellularity in the thymus. In particular, ocular toxicities were not observed. Conclusions: MGC028 exhibited potent antitumor activity in in vivo models representing various solid cancer indications and was well tolerated in non-human primates at exposure levels exceeding those required for antitumor activity. Our findings support continued investigation of MGC028 as an ADC therapeutic for the treatment of ADAM9-expressing solid cancers. Citation Format: Juniper A. Scribner, Jennifer G. Brown, Thomas Son, Linda Jin, Carroll McKenzie, Viktoriya Nam, Curtis Bush, Dienis Quinonez, Delta Ford, Verlene Gonzalez, James Tamura, Sergey Gorlatov, Hua Li, Shelley Butler, Ezio Bonvini, Deryk Loo. Preclinical development of MGC028, an ADAM9-targeted, glycan-linked, exatecan-based antibody-drug conjugate for the treatment of solid cancers [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 1897.