Preclinical Settings Research Articles

The Roles of Imaging Biomarkers in the Management of Chronic Neuropathic Pain.

Chronic neuropathic pain (CNP) affects around 10% of the general population and has a significant social, emotional, and economic impact. Current diagnosis techniques rely mainly on patient-reported outcomes and symptoms, which leads to significant diagnostic heterogeneity and subsequent challenges in management and assessment of outcomes. As such, it is necessary to review the approach to a pathology that occurs so frequently, with such burdensome and complex implications. Recent research has shown that imaging methods can detect subtle neuroplastic changes in the central and peripheral nervous system, which can be correlated with neuropathic symptoms and may serve as potential markers. The aim of this paper is to review available imaging methods used for diagnosing and assessing therapeutic efficacy in CNP for both the preclinical and clinical setting. Of course, further research is required to standardize and improve detection accuracy, but available data indicate that imaging is a valuable tool that can impact the management of CNP.

The Role of Interleukin-7 in the Formation of Tertiary Lymphoid Structures and Their Prognostic Value in Gastrointestinal Cancers.

Immunotherapies for the treatment of solid tumors continue to develop in preclinical and clinical research settings. Unfortunately, for many patients the tumor fails to respond or becomes resistant to therapies such as checkpoint inhibitors (CPIs) targeting programmed cell death protein-1 (PD-1), programmed death-ligand 1 (PD-L1), and cytotoxic T lymphocyte antigen-4 (CTLA-4). In many cancers, failed response to CPIs can be attributed to poor T cell infiltration, dominant immunosuppression, and exhausted immune responses. In gastrointestinal (GI) cancers T cell infiltration can be dismal, with several reports finding that CD8+ T cells compose less than 2% of all cells within the tumor. Organized aggregates of lymphocytes, antigen-presenting cells, and vessels, together termed tertiary lymphoid structures (TLSs), are hypothesized to be a major source of T cells within solid tumors. The intratumoral formation of these organized immune centers appears to rely on intricate cytokine and chemokine signaling to heterogeneous cell populations such as B and T cells, innate lymphoid cells, fibroblasts, and dendritic cells. In GI cancers, the presence and density of TLSs provide prognostic value for predicting outcome and survival. Further, TLS presence and density associates with favorable responses to CPIs in many cancers. This review highlights the prognostic value of TLSs in GI cancers, the role of the homeostatic cytokine interleukin-7 (IL-7) in TLS formation, and the induction of TLSs in solid tumors by novel therapeutics.

Enrichment of hepatic glycogen and plasma glucose from H₂18 O informs gluconeogenic and indirect pathway fluxes in naturally feeding mice.

Deuterated water (2 H2 O) is a widely used tracer of carbohydrate biosynthesis in both preclinical and clinical settings, but the significant kinetic isotope effects (KIE) of 2 H can distort metabolic information and mediate toxicity. 18 O-water (H2 18 O) has no significant KIE and is incorporated into specific carbohydrate oxygens via well-defined mechanisms, but to date it has not been evaluated in any animal model. Mice were given H2 18 O during overnight feeding and 18 O-enrichments of liver glycogen, triglyceride glycerol (TG), and blood glucose were quantified by 13 C NMR and mass spectrometry (MS). Enrichment of oxygens 5 and 6 relative to body water informed indirect pathway contributions from the Krebs cycle and triose phosphate sources. Compared with mice fed normal chow (NC), mice whose NC was supplemented with a fructose/glucose mix (i.e., a high sugar [HS] diet) had significantly higher indirect pathway contributions from triose phosphate sources, consistent with fructose glycogenesis. Blood glucose and liver TG 18 O-enrichments were quantified by MS. Blood glucose 18 O-enrichment was significantly higher for HS versus NC mice and was consistent with gluconeogenic fructose metabolism. TG 18 O-enrichment was extensive for both NC and HS mice, indicating a high turnover of liver triglyceride, independent of diet. Thus H2 18 O informs hepatic carbohydrate biosynthesis in similar detail to 2 H2 O but without KIE-associated risks.

The Feasibility of Biologically Guided Radiation Therapy Implemented in a Small-Animal Radiation Therapy Platform Integrated with Quad-Modal On-Board Imaging

<h3>Purpose/Objective(s)</h3> Unparallel evolution between clinical radiation therapy (RT) paradigms and preclinical RT techniques, especially in biologically guided RT, has posed a challenge to completely validate the benefits and pitfalls of novel RT techniques prior to their translation to bedside practices. In this study, we firstly investigated the feasibility of biologically guided RT implemented within a small animal radiation therapy (SART) platform, with quad-modal on-board positron emission tomography (PET), single-photon emission computed tomography (SPECT), spectral CT, and cone-beam computed tomography (CBCT) imaging integrated, in a Monte Carlo model as a proof-of-concept. <h3>Materials/Methods</h3> The SART platform with quad-modal imaging guidance was modeled by using a planning tool. A preclinical biologically guided RT workflow with quad-modal imaging guidance was designed, in which image-guided RT and emission-guided radiation therapy (EGRT) can be implemented. To demonstrate the potential of contrast-enhanced spectral CT used in biologically guided RT for simultaneously guiding dose prescription based on tracer uptakes and dose calculation based on underlying tissue electron densities in the platform, iodine uptake fractions and virtual non-contrast electron densities in the Kidney1 inserts within a simulated phantom, mixed with an iodine contrast agent at electron fractions of 0.02 and 0.03, were decomposed quantitatively by using the Bayesian eigen tissue decomposition method and the maximum a posterior (MAP) estimation. To demonstrate EGRT implemented in the platform, the preliminary algorithm of EGRT was illustrated by introducing a 2-mm setup error to a target in the simulated phantom. The dose calculation was implemented after the set of binary micro-multileaf collimator (bmMLC) configurations was determined. The dose distributions delivered by EGRT were compared to that delivered by the conventional RT method with three treatment beams with and without setup error. <h3>Results</h3> For the Kidney1 insert with an iodine contrast agent fraction of 0.02, the average relative errors in the iodine contrast agent fraction and the underlying electron density estimated were 0.64% and 0.95%, respectively. In EGRT, the simulations resulted in 4339 and 5197 beamlet responses in an exemplary dose slice delivery with and without the 2-mm setup error, respectively. Compared to the conventional RT method, EGRT scheme introduced into the SART platform was validated to be more robust to setup errors while providing improved tumor targeting and a lesser dose burden on normal tissues during the treatment beam delivery in preclinical RT setting. <h3>Conclusion</h3> The results demonstrated the successful implementation of biologically guided RT within the SART platform with quad-model imaging guidance. With the comprehensive image- and emission-guided RT schemes integrated, this novel SART platform affords one-stop preclinical RT studies.

Alternative routes for 64Cu production using an 18 MeV medical cyclotron in view of theranostic applications

Radiometals play a fundamental role in the development of personalized nuclear medicine. In particular, copper radioisotopes are attracting increasing interest since they offer a varying range of decay modes and half-lives and can be used for imaging (60Cu, 61Cu, 62Cu and 64Cu) and targeted radionuclide therapy (64Cu and 67Cu), providing two of the most promising true theranostic pairs, namely 61Cu/67Cu and 64Cu/67Cu. Currently, the most widely used in clinical applications is 64Cu, which has a unique decay scheme featuring β+-, β−-decay and electron capture. These characteristics allow its exploitation in both diagnostic and therapeutic fields. However, although 64Cu has extensively been investigated in academic research and preclinical settings, it is still scarcely used in routine clinical practice due to its insufficient availability at an affordable price. In fact, the most commonly used production method involves proton irradiation of enriched 64Ni, which has a very low isotopic abundance and is therefore extremely expensive. In this paper, we report on the study of two alternative production routes, namely the 65Cu(p,pn)64Cu and 67Zn(p, α)64Cu reactions, which enable low and high 64Cu specific activities, respectively. To optimize the 64Cu production, while minimizing the mass of copper used as a target in the first case, or the co-production of other copper radioisotopes in the second case, an accurate knowledge of the production cross sections is of paramount importance. For this reason, the involved nuclear reaction cross sections were measured at the Bern medical cyclotron laboratory by irradiating enriched 65CuO and enriched 67ZnO targets. On the basis of the obtained results, the production yield and purity were calculated to assess the optimal irradiation conditions. Several production tests were performed to confirm these findings.

Deferiprone attenuates neuropathology and improves outcome following traumatic brain injury.

Traumatic brain injury (TBI) remains a leading cause of mortality and morbidity in young adults. The role of iron in potentiating neurodegeneration following TBI has gained recent interest as iron deposition has been detected in the injured brain in the weeks to months post-TBI, in both the preclinical and clinical setting. A failure in iron homeostasis can lead to oxidative stress, inflammation and excitotoxicity; and whether this is a cause or consequence of the long-term effects of TBI remains unknown. We investigated the role of iron and the effect of therapeutic intervention using a brain-permeable iron chelator, deferiprone, in a controlled cortical impact mouse model of TBI. An extensive assessment of cognitive, motor and anxiety/depressive outcome measures were examined, and neuropathological and biochemical changes, over a 3-month period post-TBI. Lesion volume was significantly reduced at 3 months, which was preceded by a reduction in astrogliosis, microglia/macrophages and preservation of neurons in the injured brain at 2 weeks and/or 1 month post-TBI in mice receiving oral deferiprone. Deferiprone treatment showed significant improvements in neurological severity scores, locomotor/gait performance and cognitive function, and attenuated anxiety-like symptoms post-TBI. Deferiprone reduced iron levels, lipid peroxidation/oxidative stress and altered expression of neurotrophins in the injured brain over this period. Our findings support a detrimental role of iron in the injured brain and suggest that deferiprone (or similar iron chelators) may be promising therapeutic approaches to improve survival, functional outcomes and quality of life following TBI.

Chlorin e6-loaded goat milk-derived extracellular vesicles for Cerenkov luminescence-induced photodynamic therapy.

Photodynamic therapy (PDT) is a promising cancer treatment strategy with rapid progress in preclinical and clinical settings. However, the limitations in penetration of external light and precise delivery of photosensitizers hamper its clinical translation. As such, the internal light source such as Cerenkov luminescence (CL) from decaying radioisotopes offers new opportunities. Herein, we show that goat milk-derived extracellular vesicles (GEV) can act as a carrier to deliver photosensitizer Chlorin e6 (Ce6) and tumor-avid 18F-FDG can activate CL-induced PDT for precision cancer theranostics. GEV was isolated via differential ultracentrifugation of commercial goat milk and photosensitizer Ce6 was loaded by co-incubation to obtain Ce6@GEV. Tumor uptake of Ce6@GEV was examined using confocal microscopy and flow cytometry. To demonstrate the ability of 18F-FDG to activate photodynamic effects against cancer cells, apoptosis rates were measured using flow cytometry, and the production of 1O2 was measured by reactive oxygen species (ROS) monitoring kit. Moreover, we used the IVIS device to detect Cherenkov radiation and Cerenkov radiation energy transfer (CRET). For animal experiments, a small-animal IVIS imaging system was used to visualize the accumulation of the GEV drug delivery system in tumors. PET/CT and CL images of the tumor site were performed at 0.5, 1, and 2 h. For in vivo antitumor therapy, changes of tumor volume, survival time, and body weight in six groups of tumor-bearing mice were monitored. Furthermore, the blood sample and organs of interest (heart, liver, spleen, lungs, kidneys, and tumor) were collected for hematological analysis, immunohistochemistry, and H&E staining. Confocal microscopy of 4T1 cells incubated with Ce6@GEV for 4 h revealed strong red fluorescence signals in the cytoplasm, which demonstrated that Ce6 loaded in GEV could be efficiently delivered into tumor cells. When Ce6@GEV and 18F-FDG co-existed incubated with 4T1 cells, the cell viability plummeted from more than 88.02 ± 1.30% to 23.79 ± 1.59%, indicating excellent CL-induced PDT effects. In vivo fluorescence images showed a peak tumor/liver ratio of 1.36 ± 0.09 at 24 h after Ce6@GEV injection. For in vivo antitumor therapy, Ce6@GEV + 18F-FDG group had the best tumor inhibition rate (58.02%) compared with the other groups, with the longest survival rate (35 days, 40%). During the whole treatment process, neither blood biochemical analysis nor histological observation revealed vital organ damage, suggesting the biosafety of this treatment strategy. The simultaneous accumulation of 18F-FDG and Ce6 in tumor tissues is expected to overcome the deficiency of traditional PDT. This strategy has the potential to extend PDT to a variety of tumors, including metastases, using targeted radiotracers to provide internal excitation of light-responsive therapeutics. We expect that our method will play a critical role in precision treatment of deep solid tumors.

A novel assessment of fine-motor function reveals early hindlimb and detectable forelimb deficits in an experimental model of ALS

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disorder associated with the loss of cortical and spinal motor neurons (MNs) and muscle degeneration (Kiernan et al. in Lancet 377:942–955, 2011). In the preclinical setting, functional tests that can detect early changes in motor function in rodent models of ALS are critical to understanding the etiology of the disease and treatment development. Here, we established a string-pulling paradigm that can detect forelimb and hindlimb motor deficits in the SOD1 mouse model of ALS earlier than traditional motor performance tasks. Additionally, our findings indicate that early loss of forelimb and hindlimb function is correlated with cortical and spinal MN loss, respectively. This task is not only ecological, low-cost, efficient, and non-onerous, it also requires little animal handling and reduces the stress placed on the animal. It has long been a concern in the field that the SOD1 mouse does not display forelimb motor deficits and does not give researchers a complete picture of the disease. Here, we provide evidence that the SOD1 model does in fact develop early forelimb motor deficits due to the task’s ability to assess fine-motor function, reconciling this model with the various clinical presentation of ALS. Taken together, the string-pulling paradigm may provide novel insights into the pathogenesis of ALS, offer nuanced evaluation of prospective treatments, and has high translational potential to the clinic.

Combinatory anti-tumor activities of 1,4-bis[2-(dimethylamino)ethylamino]-5,8-dihydroxyanthracene-9,10-dione (AQ4) and temsirolimus against colorectal cancer cells.

Banoxantrone is a topoisomerase II inhibitor that is selectively activated in hypoxia. Although it has exhibited anti-tumor activity against several types of cancers in preclinical models, its efficacy against colorectal cancer (CRC) remains unclear. We examined the antitumor effects of 1,4-bis[2-(dimethylamino)ethylamino]-5,8-dihydroxyanthracene-9,10-dione (AQ4), an activated metabolite of banoxantrone, in CRC cell lines (HT-29, CaR-1) using in vitro experiments under normoxic and hypoxic conditions. The inhibition of cell growth was assessed using a proliferation assay. The induction of apoptosis and changes in the cell cycle were measured using flow cytometry. Signaling pathways involved in apoptosis and hypoxia were analyzed. The anti-tumor activity of temsirolimus, an inhibitor of mammalian target of rapamycin, and the combined effects of temsirolimus and AQ4 were also evaluated. Regardless of the oxygen condition, a single drug treatment with AQ4 or temsirolimus inhibited proliferation and induced apoptosis in both cell lines, accompanied by a reduction in the phosphorylation of S6. AQ4 induced G2/M cell cycle arrest, whereas temsirolimus induced G0/G1 arrest. Moreover, the combined treatment markedly reduced the proportion of cells in the S phase and enhanced apoptosis, as evidenced by an increased Bax/Bcl-2 ratio. The hypoxia-induced activation of the HIF-1α pathway was suppressed by AQ4 and temsirolimus. Based on the cooperative anti-tumor activity of AQ4 and temsirolimus in vitro, the combination of banoxantrone plus temsirolimus has potential as a treatment option for CRC in preclinical and clinical settings.

Therapeutic Use of an Inhaled Drug Delivery in Pulmonary Hypertension: A Review.

Pulmonary arterial hypertension (PAH) is a seriouscondition in which there is increased blood pressure in arteries of the lungs (pulmonary arteries). The therapies or drugs for PAH have expanded with the revelation of three key pathological processes - encompassing prostacyclin, nitric oxide (NO), and endothelin pathways. An outlook for patients suffering from PAH is still mediocre amidst recent advancements. The evolution of pre-clinical and clinical research on PAH has facilitated the identification of several new targeted therapies for the disease. In this article, we examine recent data on new pulmonary hypertension physiological pathways, primarily concentrating on administering drugs through the inhalation route and their effects. Although they have been given clinical use approval, medications based on these routes are presently being studied in clinical or pre-clinical settings. To confirm these innovative medicines' therapeutic efficacy and safety, extensive clinical trials are needed.

Senescence plays a role in myotonic dystrophy type 1.

Myotonic dystrophy type 1 (DM1; MIM #160900) is an autosomal dominant disorder, clinically characterized by progressive muscular weakness and multisystem degeneration. The broad phenotypes observed in patients with DM1 resemble the appearance of an accelerated aging process. However, the molecular mechanisms underlying these phenotypes remain largely unknown. Transcriptomic analysis of fibroblasts derived from patients with DM1 and healthy individuals revealed a decrease in cell cycle activity, cell division, and DNA damage response in DM1, all of which related to the accumulation of cellular senescence. The data from transcriptome analyses were corroborated in human myoblasts and blood samples, as well as in mouse and Drosophila models of the disease. Serial passage studies in vitro confirmed the accelerated increase in senescence and the acquisition of a senescence-associated secretory phenotype in DM1 fibroblasts, whereas the DM1 Drosophila model showed reduced longevity and impaired locomotor activity. Moreover, functional studies highlighted the impact of BMI1 and downstream p16INK4A/RB and ARF/p53/p21CIP pathways in DM1-associated cellular phenotypes. Importantly, treatment with the senolytic compounds Quercetin, Dasatinib, or Navitoclax reversed the accelerated aging phenotypes in both DM1 fibroblasts in vitro and in Drosophila in vivo. Our results identify the accumulation of senescence as part of DM1 pathophysiology and, therefore, demonstrate the efficacy of senolytic compounds in the preclinical setting.

Update on the recent development of allosteric modulators for adenosine receptors and their therapeutic applications.

Adenosine receptors (ARs) have been identified as promising therapeutic targets for countless pathological conditions, spanning from inflammatory diseases to central nervous system disorders, from cancer to metabolic diseases, from cardiovascular pathologies to respiratory diseases, and beyond. This extraordinary therapeutic potential is mainly due to the plurality of pathophysiological actions of adenosine and the ubiquitous expression of its receptors. This is, however, a double-edged sword that makes the clinical development of effective ligands with tolerable side effects difficult. Evidence of this is the low number of AR agonists or antagonists that have reached the market. An alternative approach is to target allosteric sites via allosteric modulators, compounds endowed with several advantages over orthosteric ligands. In addition to the typical advantages of allosteric modulators, those acting on ARs could benefit from the fact that adenosine levels are elevated in pathological tissues, thus potentially having negligible effects on normal tissues where adenosine levels are maintained low. Several A1 and various A3AR allosteric modulators have been identified so far, and some of them have been validated in different preclinical settings, achieving promising results. Less fruitful, instead, has been the discovery of A2A and A2BAR allosteric modulators, although the results obtained up to now are encouraging. Collectively, data in the literature suggests that allosteric modulators of ARs could represent valuable pharmacological tools, potentially able to overcome the limitations of orthosteric ligands.

Confocal laser imaging in neurosurgery: A comprehensive review of sodium fluorescein-based CONVIVO preclinical and clinical applications.

Given the established direct correlation that exists among extent of resection and postoperative survival in brain tumors, obtaining complete resections is of primary importance. Apart from the various technological advancements that have been introduced in current clinical practice, histopathological study still remains the gold-standard for definitive diagnosis. Frozen section analysis still represents the most rapid and used intraoperative histopathological method that allows for an intraoperative differential diagnosis. Nevertheless, such technique owes some intrinsic limitations that limit its overall potential in obtaining real-time diagnosis during surgery. In this context, confocal laser technology has been suggested as a promising method to have near real-time intraoperative histological images in neurosurgery, thanks to the results of various studies performed in other non-neurosurgical fields. Still far to be routinely implemented in current neurosurgical practice, pertinent literature is growing quickly, and various reports have recently demonstrated the utility of this technology in both preclinical and clinical settings in identifying brain tumors, microvasculature, and tumor margins, when coupled to the intravenous administration of sodium fluorescein. Specifically in neurosurgery, among different available devices, the ZEISS CONVIVO system probably boasts the most recent and largest number of experimental studies assessing its usefulness, which has been confirmed for identifying brain tumors, offering a diagnosis and distinguishing between healthy and pathologic tissue, and studying brain vessels. The main objective of this systematic review is to present a state-of-the-art summary on sodium fluorescein-based preclinical and clinical applications of the ZEISS CONVIVO in neurosurgery.

Comparison of PSMA-TO-1 and PSMA-617 labeled with gallium-68, lutetium-177 and actinium-225

BackgroundPSMA-TO-1 (“Tumor-Optimized-1”) is a novel PSMA ligand with longer circulation time than PSMA-617. We compared the biodistribution in subcutaneous tumor-bearing mice of PSMA-TO-1, PSMA-617 and PSMA-11 when labeled with 68Ga and 177Lu, and the survival after treatment with 225Ac-PSMA-TO-1/-617 in a murine model of disseminated prostate cancer. We also report dosimetry data of 177Lu-PSMA-TO1/-617 in prostate cancer patients.MethodsFirst, PET images of 68Ga-PSMA-TO-1/-617/-11 were acquired on consecutive days in three mice bearing subcutaneous C4-2 xenografts. Second, 50 subcutaneous tumor-bearing mice received either 30 MBq of 177Lu-PSMA-617 or 177Lu-PSMA-TO-1 and were sacrificed at 1, 4, 24, 48 and 168 h for ex vivo gamma counting and biodistribution. Third, mice bearing disseminated lesions via intracardiac inoculation were treated with either 40 kBq of 225Ac-PSMA-617, 225Ac-PSMA-TO-1, or remained untreated and followed for survival. Additionally, 3 metastatic castration-resistant prostate cancer patients received 500 MBq of 177Lu-PSMA-TO-1 under compassionate use for dosimetry purposes. Planar images with an additional SPECT/CT acquisition were acquired for dosimetry calculations.ResultsTumor uptake measured by PET imaging of 68Ga-labeled agents in mice was highest using PSMA-617, followed by PSMA-TO-1 and PSMA-11. 177Lu-PSMA tumor uptake measured by ex vivo gamma counting at subsequent time points tended to be greater for PSMA-TO-1 up to 1 week following treatment (p > 0.13 at all time points). This was, however, accompanied by increased kidney uptake and a 26-fold higher kidney dose of PSMA-TO-1 compared with PSMA-617 in mice. Mice treated with a single-cycle 225Ac-PSMA-TO-1 survived longer than those treated with 225Ac-PSMA-617 and untreated mice, respectively (17.8, 14.5 and 7.7 weeks, respectively; p < 0.0001). Kidney, salivary gland, bone marrow and mean ± SD tumor dose coefficients (Gy/GBq) for 177Lu-PSMA-TO-1 in patients #01/#02/#03 were 2.5/2.4/3.0, 1.0/2.5/2.3, 0.14/0.11/0.10 and 0.42 ± 0.03/4.45 ± 0.07/1.8 ± 0.57, respectively.ConclusionsPSMA-TO-1 tumor uptake tended to be greater than that of PSMA-617 in both preclinical and clinical settings. Mice treated with 225Ac-PSMA-TO-1 conferred a significant survival benefit compared to 225Ac-PSMA-617 despite the accompanying increased kidney uptake. In humans, PSMA-TO-1 dosimetry estimates suggest increased tumor absorbed doses; however, the kidneys, salivary glands and bone marrow are also exposed to higher radiation doses. Thus, additional preclinical studies are needed before further clinical use.

Spironolactone alleviates schizophrenia-related reversal learning in Tcf4 transgenic mice subjected to social defeat

Cognitive deficits are a hallmark of schizophrenia, for which no convincing pharmacological treatment option is currently available. Here, we tested spironolactone as a repurposed compound in Tcf4 transgenic mice subjected to psychosocial stress. In this ‘2-hit’ gene by environment mouse (GxE) model, the animals showed schizophrenia-related cognitive deficits. We had previously shown that spironolactone ameliorates working memory deficits and hyperactivity in a mouse model of cortical excitatory/inhibitory (E/I) dysbalance caused by an overactive NRG1-ERBB4 signaling pathway. In an add-on clinical study design, we used spironolactone as adjuvant medication to the standard antipsychotic drug aripiprazole. We characterized the compound effects using our previously established Platform for Systematic Semi-Automated Behavioral and Cognitive Profiling (PsyCoP). PsyCoP is a widely applicable analysis pipeline based on the Research Domain Criteria (RDoC) framework aiming at facilitating translation into the clinic. In addition, we use dimensional reduction to analyze and visualize overall treatment effect profiles. We found that spironolactone and aripiprazole improve deficits of several cognitive domains in Tcf4tg x SD mice but partially interfere with each other’s effect in the combination therapy. A similar interaction was detected for the modulation of novelty-induced activity. In addition to its strong activity-dampening effects, we found an increase in negative valence measures as a side effect of aripiprazole treatment in mice. We suggest that repurposed drug candidates should first be tested in an adequate preclinical setting before initiating clinical trials. In addition, a more specific and effective NRG1-ERBB4 pathway inhibitor or more potent E/I balancing drug might enhance the ameliorating effect on cognition even further.

Melatonin for Neuropathic Pain: Protocol for a Double-blind, Randomized Controlled Trial.

BackgroundNeuropathic pain (NP), a complication of several conditions (eg, diabetic neuropathy and varicella zoster), is a common challenging problem, and there is a growing need to develop safe and effective nonopioid treatments. Sleep disturbance is commonly associated with NP because pain intensity in NP conditions is often worse at night. The pineal hormone melatonin has been shown to reduce pain in both preclinical and clinical settings, in addition to multiple trials demonstrating efficacy for primary insomnia and delayed sleep phase syndrome.ObjectiveWe propose to conduct a clinical trial to evaluate the efficacy and safety of melatonin for NP.MethodsUsing a double-blind, placebo-controlled, crossover design, 30 adults with NP will be randomly allocated to one of two sequences of treatment with melatonin and placebo. During each of the two treatment periods, participants will take capsules containing melatonin or placebo for 4 weeks, followed by a 7-day washout period. The primary outcome will be mean daily pain intensity (scored 0-10) at maximally tolerated doses (MTDs) during each period. Secondary outcomes, assessed at MTDs, will include global improvement, adverse events, mood, and quality of life.ResultsThis trial was registered in the International Standard Randomized Controlled Trial registry May 4, 2022 (ISRCTN #16215617), attained conditional ethics approval May 9, 2022 (Queen’s University Health Sciences & Affiliated Teaching Hospitals Research Ethics Board protocol number ANAE-387-22), and recruitment is set to start August 2022.ConclusionsThis trial will provide rigorous evidence comparing the efficacy of melatonin to that of placebo in the treatment of NP.Trial RegistrationISRCTN Registry 16215617; https://www.isrctn.com/ISRCTN16215617International Registered Report Identifier (IRRID)PRR1-10.2196/40025

Ischemic Preconditioning and Postconditioning Protect the Heart by Preserving the Mitochondrial Network.

Background Mitochondria fuse to form elongated networks which are more tolerable to stress and injury. Ischemic pre- and postconditioning (IPC and IPost, respectively) are established cardioprotective strategies in the preclinical setting. Whether IPC and IPost modulates mitochondrial morphology is unknown. We hypothesize that the protective effects of IPC and IPost may be conferred via preservation of mitochondrial network. Methods IPC and IPost were applied to the H9c2 rat myoblast cells, isolated adult primary murine cardiomyocytes, and the Langendorff-isolated perfused rat hearts. The effects of IPC and IPost on cardiac cell death following ischemia-reperfusion injury (IRI), mitochondrial morphology, and gene expression of mitochondrial-shaping proteins were investigated. Results IPC and IPost successfully reduced cardiac cell death and myocardial infarct size. IPC and IPost maintained the mitochondrial network in both H9c2 and isolated adult primary murine cardiomyocytes. 2D-length measurement of the 3 mitochondrial subpopulations showed that IPC and IPost significantly increased the length of interfibrillar mitochondria (IFM). Gene expression of the pro-fusion protein, Mfn1, was significantly increased by IPC, while the pro-fission protein, Drp1, was significantly reduced by IPost in the H9c2 cells. In the primary cardiomyocytes, gene expression of both Mfn1 and Mfn2 were significantly upregulated by IPC and IPost, while Drp1 was significantly downregulated by IPost. In the Langendorff-isolated perfused heart, gene expression of Drp1 was significantly downregulated by both IPC and IPost. Conclusion IPC and IPost-mediated upregulation of pro-fusion proteins (Mfn1 and Mfn2) and downregulation of pro-fission (Drp1) promote maintenance of the interconnected mitochondrial network, ultimately conferring cardioprotection against IRI.

Mucosal COVID-19 vaccines: Risks, benefits and control of the pandemic.

Based on mucosal immunization to promote both mucosal and systemic immune responses, next-generation coronavirus disease 2019 (COVID-19) vaccines would be administered intranasally or orally. The goal of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccines is to provide adequate immune protection and avoid severe disease and death. Mucosal vaccine candidates for COVID-19 including vector vaccines, recombinant subunit vaccines and live attenuated vaccines are under development. Furthermore, subunit protein vac-cines and virus-vectored vaccines have made substantial progress in preclinical and clinical settings, resulting in SARS-CoV-2 intranasal vaccines based on the previously successfully used nasal vaccines. Additional to their ability to trigger stable, protective immune responses at the sites of pathogenic infection, the development of ‘specific’ mucosal vaccines targeting coronavirus antigens could be an excellent option for preventing future pandemics. However, their efficacy and safety should be confirmed.

Nanoparticles and Radioisotopes: A Long Story in a Nutshell.

The purpose of this narrative review was to assess the use of nanoparticles (NPs) to deliver radionuclides to targets, focusing on systems that have been tested in pre-clinical and, when available, clinical settings. A literature search was conducted in PubMed and Web of Science databases using the following terms: “radionuclides” AND “liposomes” or “PLGA nanoparticles” or “gold nanoparticles” or “iron oxide nanoparticles” or “silica nanoparticles” or “micelles” or “dendrimers”. No filters were applied, apart from a minimum limit of 10 patients enrolled for clinical studies. Data from some significant studies from pre-clinical and clinical settings were retrieved, and we briefly describe the information available. All the selected seven classes of nanoparticles were highly tested in clinical trials, but they all present many drawbacks. Liposomes are the only ones that have been tested for clinical applications, though they have never been commercialized. In conclusion, the application of NPs for imaging has been the object of much interest over the years, albeit mainly in pre-clinical settings. Thus, we think that, based on the current state, radiolabeled NPs must be investigated longer before finding their place in nuclear medicine.

Improving the predictive power of xenograft and syngeneic anti-tumour studies using mice humanised for pathways of drug metabolism.

Drug development is an expensive and time-consuming process, with only a small fraction of drugs gaining regulatory approval from the often many thousands of candidates identified during target validation. Once a lead compound has been identified and optimised, they are subject to intensive pre-clinical research to determine their pharmacodynamic, pharmacokinetic and toxicological properties, procedures which inevitably involve significant numbers of animals - mainly mice and rats, but also dogs and monkeys in much smaller numbers and for specific types of drug candidates. Many compounds that emerge from this process, having been shown to be safe and efficacious in pre-clinical studies, subsequently fail to replicate this outcome in clinical trials, therefore wasting time, money and, most importantly, animals. The poor predictive power of animal models in pre-clinical studies is predominantly due to lack of efficacy or safety reasons, which in turn can be attributed mainly to the significant species differences in drug metabolism between humans and animals. To circumvent this, we have developed a complex transgenic mouse model - 8HUM - which faithfully replicates human Phase I drug metabolism (and its regulation), and which will generate more human-relevant data [ REFINEMENT] from fewer animals [ REDUCTION] in a pre-clinical setting and reduce attrition in the clinic. One key area for the pre-clinical application of animals in an oncology setting - almost exclusively mice - is their use in anti-tumour studies. We now further demonstrate the utility of the 8HUM mouse using a murine melanoma cell line as a syngeneic tumour and also present an immunodeficient version 8HUM_Rag2 -/- - for use in xenograft studies. These models will be of significant benefit not only to Pharma for pre-clinical drug development work, but also throughout the drug efficacy, toxicology, pharmacology, and drug metabolism communities, where fewer animals will be needed to generate more human-relevant data.