Aprepitant Restores Corneal Sensitivity and Reduces Pain in DED.

Purpose: This study aims to assess the efficacy of two aprepitant formulations (X1 and X2), in a preclinical model of dry eye disease (DED) induced by benzalkonium chloride (BAK). Methods: Two aprepitant formulations were tested on 7 to 8-week-old male mice for their efficacy. In vivo corneal fluorescein staining assessed epithelial damage as the primary end point on days 0, 3, 5, 7, 9, 12, and 14 using slit-lamp microscopy. The DED model was induced with 0.2% BAK twice daily for the first week and once daily for the next week. Mice were randomly assigned to 5 treatment groups: Aprepitant X1 (n = 10) and X2 (n = 10) formulation, 2 mg/mL dexamethasone (n = 10), control vehicle X (n = 10), 0.2% hyaluronic acid (n = 10), or no treatment (n = 10). Eye wiping, phenol red, and Cochet Bonnet tests assessed ocular pain, tear fluid secretion, and nerve function. After 7 days, the mice were euthanized to quantify leukocyte infiltration and corneal nerve density. Results: Topical aprepitant X1 reduced BAK-induced corneal damage and pain compared to gel vehicle X (P = 0.007) and dexamethasone (P = 0.021). Aprepitant X1 and X2 improved corneal sensitivity versus gel vehicle X and dexamethasone (P < 0.001). Aprepitant X1 reduced leukocyte infiltration (P < 0.05) and enhanced corneal nerve density (P < 0.001). Tear fluid secretion remained statistically unchanged in both the X1 and X2 groups. Conclusions: Aprepitant formulation X1 reduced pain, improved corneal sensitivity and nerve density, ameliorated epitheliopathy, and reduced leukocyte infiltration in male mouse corneas. Translational Relevance: Aprepitant emerges as a safe, promising therapeutic prospect for the amelioration of DED's associated symptoms.

The combined anti-tumor effects of 5-fluorouracil and neurokinin receptor inhibitor, aprepitant, against colorectal cancer: In vitro and in vivo study.

BACKGROUND: Colorectal cancer (CRC) is one of the world's largest health concerns with growing global incidence and mortality. The potential value of the neurokinin-1 receptor as a therapeutic target has been reported in several tumor types, including CRC. Here we examined the potential anti-tumor effects of a clinically approved neurokinin-1 receptor antagonist, aprepitant, alone and its combination with 5-Fluorouracil (5-FU) as a first choice CRC chemotherapeutic drug, in both in vitro and in vivo models of CRC. METHODS: MTT assay was employed for assessing cell proliferation. mRNA expression levels were determined by quantitative real-time PCR (qRT-PCR). Flow cytometric analysis of apoptosis was performed using an Annexin-V/propidium iodide assay kit. We finally conducted an in vivo experiment in a mouse model of CRC to confirm the in vitro antiproliferative activity of aprepitant and 5-FU. RESULTS: We found that aprepitant and 5-FU significantly reduced CRC cell viability. The combination of drugs exhibited potent synergistic growth inhibitory effects on CRC cells. Moreover, aprepitant and 5-FU induced apoptosis and altered the levels of apoptotic genes (up-regulation of Bax, and p53 along with downregulation of Bcl-2). Importantly, the aprepitant and 5-FU combination showed a more pronounced impact on apoptosis and associated genes than either of the agents alone. Furthermore, aprepitant reduced tumor growth in vivo and led to significantly longer survival time, and this effect was more prominent when using the aprepitant and 5-FU combination. CONCLUSIONS: Collectively, combinatory treatment with aprepitant and 5-FU potentially exerts synergistic growth inhibition and apoptosis induction in CRC, deserving further consideration as a novel strategy for CRC patients.

Biological evaluation of carbohydrate-based aprepitant analogs for neuroblastoma treatment.

Different studies using Aprepitant, a NK1R antagonist currently used as a clinical drug for treating chemotherapy-related nausea and vomiting, have demonstrated that pharmacological inhibition of NK1R effectively reduces the growth of several tumor types such as neuroblastoma (NB). In a previous work, we demonstrated that a series of carbohydrate-based Aprepitant analogs, derived from either d-galactose or l-arabinose, have shown high affinity and NK1R antagonistic activity with a broad-spectrum anticancer activity and an important selectivity. In this new study, we explore the selective cytotoxic effects of these derivatives for the treatment of NB. Furthermore, we describe the design and stereoselective synthesis of a new generation of d-glucose derivatives as Aprepitant analogs, supported by docking studies. This approach showed that most of our carbohydrate-based analogs are significantly more selective than Aprepitant. The galactosyl derivative 2α, has demonstrated a marked in vitro selective cytotoxic activity against NB, with IC50 values in the same range as those of Aprepitant and its prodrug Fosaprepitant. Interestingly, the derivative 2α has shown similar apoptotic effect to that of Aprepitant. Moreover, we can select the glucosyl amino derivative 10α as an interesting hit exhibiting higher in vitro cytotoxic activity against NB than Aprepitant, being 1.2 times more selective.

The Repurposing of Non-Peptide Neurokinin-1 Receptor Antagonists as Antitumor Drugs: An Urgent Challenge for Aprepitant.

The substance P (SP)/neurokinin-1 receptor (NK-1R) system is involved in cancer progression. NK-1R, activated by SP, promotes tumor cell proliferation and migration, angiogenesis, the Warburg effect, and the prevention of apoptosis. Tumor cells overexpress NK-1R, which influences their viability. A typical specific anticancer strategy using NK-1R antagonists, irrespective of the tumor type, is possible because these antagonists block all the effects mentioned above mediated by SP on cancer cells. This review will update the information regarding using NK-1R antagonists, particularly Aprepitant, as an anticancer drug. Aprepitant shows a broad-spectrum anticancer effect against many tumor types. Aprepitant alone or in combination therapy with radiotherapy or chemotherapy could reduce the sequelae and increase the cure rate and quality of life of patients with cancer. Current data open the door to new cancer research aimed at antitumor therapeutic strategies using Aprepitant. To achieve this goal, reprofiling the antiemetic Aprepitant as an anticancer drug is urgently needed.

The SP/NK1R system promotes the proliferation of breast cancer cells through NF-κB-mediated inflammatory responses.

BACKGROUND: Numerous molecules have been introduced to participate in the formation of breast cancer, the most common malignancy in women. Among them, neuropeptide substance P (SP) and its related receptor neurokinin-1 receptor (NK1R) have attracted unprecedented attention in tumorigenesis processes. In this study, we investigated the effect of the SP/NK1R pathway on the induction of oxidative stress in breast cancer and examine the therapeutic potential of NK1R inhibition in this malignancy. METHODS: MCF-7 cells were treated with varying concentrations of SP and aprepitant, an FDA-approved NK1R antagonist, either as a single drug or in a combined modality. Resazurin assay was used to evaluate the anti-cancer ability of aprepitant. The alteration in the intracellular levels of reactive oxygen species (ROS) and gene expression were determined using ROS assay and the qRT-PCR analysis, respectively. RESULTS: The stimulation of the SP/NK1R axis in the MCF-7 cells was coupled with the accumulation of ROS as well as upregulation of NF-κB and its related pro-inflammatory cytokines, including tumor necrosis factor (TNF)-α and IL-6. In contrast, the suppression of NK1R by aprepitant halted the viability of MCF-7 cells, at least partly due to p53-mediated upregulation of p21. Moreover, aprepitant attenuated the oncogenic properties of SP by preventing the oxidative property of this neuropeptide. CONCLUSION: Overall, our results suggest that the SP/NK1R pathway might play a critical role in breast cancer pathogenesis, probably through inducing ROS/NF-κB-mediated inflammatory responses. Moreover, it seems that blockage of the axis has promising therapeutic value against breast cancer cells. Schematic representation proposed for the plausible mechanism by which the stimulation of the SP/NK1R might induce oxidative stress in breast cancer-derived MCF-7 cells. Once SP interacts with NK1R, this signaling axis could disturb the balance between the expression of p53 and NF-κB, an event that leads to the accumulation of ROS within MCF-7 cells. The produced ROS, in turn, elevates the expression of pro-inflammatory cytokines (TNF-α and IL-6) and downregulates the expression of p21. On the other hand, aprepitant, an antagonist of NK1R, could reduce the survival of proliferative capacity of MCF-7 cells by decreasing the intracellular levels of ROS and p53-mediated up-regulation of p21. Along with the effect on p53, aprepitant could also reduce the expression of NF-κB and its related pro-inflammatory cytokines.

The anti-tumoral role of Hesperidin and Aprepitant on prostate cancer cells through redox modifications.

Prostate cancer is the second prevalent cancer in men. While the anti-cancer effect of Hesperidin and (Aprepitant) AP on prostate cancer cells is well documented, their combined effect and their mechanism of action are not fully investigated. Therefore, this study aimed to investigate the anti-cancer effects of Hesperidin and AP alone and in combination on prostate cancer cells. PC3 and LNCaP cell lines were treated with Hesperidin and AP alone and in combination. The Resazurin test was used for assessing cell viability. The ROS (reactive oxygen Species) level, P53, P21, Bcl-2, and Survivin gene expression were assessed. Also, a trypan blue assay was done. Hesperidin and AP reduced cell viability and increased apoptosis in PC3 and LNCaP cells. The ROS level reduced after treating the PC3 and LNCaP cells with AP with or without Hesperidin. P53 and P21 gene expression increased after treatment with Hesperidin with or without AP compared to the untreated group in the PC3 cell line. Bcl-2 and Survivin gene expression decreased with AP with or without Hesperidin in the PC3 and LNCaP cells. The current study showed the synergic anti-cancer effect of Hesperidin and AP in both PC3 and LNCaP cell lines.

Aprepitant inhibits the progression of esophageal squamous cancer by blocking the truncated neurokinin­1 receptor.

Increasing evidence showed that the substance P (SP)/neurokinin­1 receptor (NK1R) complex is involved in the development of several cancers. However, little is known about the mechanisms by which SP/NK1R complex plays a role in esophageal squamous cell carcinoma (ESCC) progression. RT­qPCR, CCK­8, Transwell, western blotting, immunohistochemical, immunofluorescence, ELISA and analysis of apoptosis were employed in the present study. It was aimed to investigate the function and therapeutic potential of the SP/tr­NK1R system in human ESCC progression. The results revealed that both SP and tr­NK1R were highly expressed in ESCC cell lines and specimens. In ESCC tissues, SP was mainly derived from ESCC cells and M2 macrophages. The NK1R antagonist aprepitant inhibited the SP­induced proliferation of human ESCC cell lines. Aprepitant inhibited cell migration and invasion and induced apoptosis of ESCC cells by downregulating the PI3K/AKT/mTOR signaling pathways. Animal experiments revealed that aprepitant inhibited tumor progression of ESCC in xenograft mice. In conclusion, high expression of SP plus tr­NK1R indicated poor prognosis in ESCC, suggesting that aprepitant has a potential application in ESCC. To the best of our knowledge, high SP and tr­NK1R expression in ESCC cell lines was reported for the first time in the present study. These findings provided evidence for a novel therapeutic strategy for patients with ESCC.

A pharmacological overview of aprepitant for the prevention of postoperative nausea and vomiting.

INTRODUCTION: Post-operative nausea and vomiting (PONV) affects 30% of all patients undergoing surgery and up to 80% of high-risk patients. Antiemetics for PONV prophylaxis target a variety of receptor systems, with varying degrees of efficacy and side effect profile. Neurokinin -1 receptor antagonists are the most recent class of compounds investigated for PONV prophylaxis, with aprepitant being the only one currently approved for this indication. AREAS COVERED: This review covers the pathophysiology of PONV, current recommendations for PONV prophylaxis, pharmacokinetics, and pharmacodynamics of aprepitant, and the evidence for its efficacy in the management of PONV as a single agent and in combination therapy. EXPERT OPINION: Aprepitant is effective for PONV prophylaxis. It has superior antivomiting efficacy, long half-life, and favorable side effect profile. Data on antiemetic combinations involving aprepitant are limited, and it is not clear if the addition of other antiemetics to aprepitant results in improved PONV prophylaxis. The oral route of administration of aprepitant is a potential limitation in a busy clinical practice. However, the recent approval of an intravenous formulation could provide a more convenient route of administration. Aprepitant remains more expensive than other antiemetics, and there are no studies assessing the cost effectiveness of its use.

Therapeutic antagonism of the neurokinin 1 receptor in endosomes provides sustained pain relief.

The hypothesis that sustained G protein-coupled receptor (GPCR) signaling from endosomes mediates pain is based on studies with endocytosis inhibitors and lipid-conjugated or nanoparticle-encapsulated antagonists targeted to endosomes. GPCR antagonists that reverse sustained endosomal signaling and nociception are needed. However, the criteria for rational design of such compounds are ill-defined. Moreover, the role of natural GPCR variants, which exhibit aberrant signaling and endosomal trafficking, in maintaining pain is unknown. Herein, substance P (SP) was found to evoke clathrin-mediated assembly of endosomal signaling complexes comprising neurokinin 1 receptor (NK1R), Gαq/i, and ßarrestin-2. Whereas the FDA-approved NK1R antagonist aprepitant induced a transient disruption of endosomal signals, analogs of netupitant designed to penetrate membranes and persist in acidic endosomes through altered lipophilicity and pKa caused sustained inhibition of endosomal signals. When injected intrathecally to target spinal NK1R+ve neurons in knockin mice expressing human NK1R, aprepitant transiently inhibited nociceptive responses to intraplantar injection of capsaicin. Conversely, netupitant analogs had more potent, efficacious, and sustained antinociceptive effects. Mice expressing C-terminally truncated human NK1R, corresponding to a natural variant with aberrant signaling and trafficking, displayed attenuated SP-evoked excitation of spinal neurons and blunted nociceptive responses to SP. Thus, sustained antagonism of the NK1R in endosomes correlates with long-lasting antinociception, and domains within the C-terminus of the NK1R are necessary for the full pronociceptive actions of SP. The results support the hypothesis that endosomal signaling of GPCRs mediates nociception and provides insight into strategies for antagonizing GPCRs in intracellular locations for the treatment of diverse diseases.

The in vitro anti-cancer synergy of neurokinin-1 receptor antagonist, aprepitant, and 5-aminolevulinic acid in glioblastoma.

Glioblastoma multiforme (GBM) is the most malignant type of cerebral neoplasm in adults with a poor prognosis. Currently, combination therapy with different anti-cancer agents is at the forefront of GBM research. Hence, this study aims to evaluate the potential anti-cancer synergy of a clinically approved neurokinin-1 receptor antagonist, aprepitant, and 5-aminolevulinic acid (5-ALA), a prodrug that elicits fluorescent porphyrins in gliomas on U-87 human GBM cells. We found that aprepitant and 5-ALA effectively inhibited GBM cell viability. The combinatorial treatment of these drugs exerted potent synergistic growth inhibitory effects on GBM cells. Moreover, aprepitant and 5-ALA induced apoptosis and altered the levels of apoptotic genes (up-regulation of Bax and P53 along with downregulation of Bcl-2). Furthermore, aprepitant and 5-ALA increased the accumulation of protoporphyrin IX, a highly pro-apoptotic and fluorescent photosensitizer. Aprepitant and 5-ALA significantly inhibited GBM cell migration and reduced matrix metalloproteinases (MMP-2 and MMP-9) activities. Importantly, all these effects were more prominent following aprepitant-5-ALA combination treatment than either drug alone. Collectively, the combination of aprepitant and 5-ALA leads to considerable synergistic anti-proliferative, pro-apoptotic, and anti-migratory effects on GBM cells and provides a firm basis for further evaluation of this combination as a novel therapeutic approach for GBM.

The In Vitro Pro-inflammatory Functions of the SP/NK1R System in Prostate Cancer: a Focus on Nuclear Factor-Kappa B (NF-κB) and Its Pro-inflammatory Target Genes.

Prostate cancer is one of the main global health threats for men which is in close association with chronic inflammation. Neuropeptide substance P (SP), acting through neurokinin receptor (NK-1R), induces various pro-inflammatory responses which are strongly involved in the pathogenesis of several diseases as well as cancer. Therefore, we aimed to investigate the pro-inflammatory functions of the SP/NK1R complex in prostate cancer and the therapeutic effects of its inhibition by NK-1R antagonist, aprepitant, in vitro. MTT assay was conducted for the cytotoxicity assessment of aprepitant in prostate cancer cells. The protein expression levels were evaluated by Western blot assay. Quantitative real-time PCR (qRT-PCR) was applied to measure mRNA expression levels of pro-inflammatory cytokines. Concurrently, the protein concentrations of pro-inflammatory cytokines were also analyzed by enzyme-linked immunosorbent assay. We observed that SP increased the levels of pro-inflammatory cytokines (IL-1ß, IL-6, and TNF-α), while treatment with aprepitant reduced the effects of SP. We also indicated that SP increased the protein levels of nuclear factor-kappa B (NF-κB), as the main regulator of inflammatory processes, and also an NF-κB target gene, cyclooxygenase 2 (COX-2) in prostate cancer cells, while treatment with aprepitant reversed these effects. Taken together, our findings highlight the importance of the SP/NK1R system in the modulation of pro-inflammatory responses in prostate cancer cells and suggest that aprepitant may be developed as a novel anti-inflammatory agent for the management of cancer-associated inflammation.

Virtual screening reveals aprepitant to be a potent inhibitor of neutral sphingomyelinase 2: implications in blockade of exosome release in cancer therapy.

PURPOSE: Exosomes are membrane-derived nano-vesicles upregulated in pathological conditions like cancer. Therefore, inhibiting their release is a potential strategy for the development of more efficient combination therapies. Neutral sphingomyelinase 2 (nSMase2) is a key component in exosome release; however, a clinically safe yet efficient nSMase2 inhibitor remains to be used discovered. Accordingly, we made an effort to identify potential nSMase2 inhibitor(s) among the approved drugs. METHODS: Virtual screening was performed and aprepitant was selected for further investigation. To evaluate the reliability of the complex, molecular dynamics were performed. Finally, using the CCK-8 assay in HCT116 cells, the highest non-toxic concentrations of aprepitant were identified and the nSMase2 activity assay was performed to measure the inhibitory activity of aprepitant, in vitro. RESULTS: To validate the screening results, molecular docking was performed, and the retrieved scores were in line with the screening results. The root-mean-square deviation (RMSD) plot of aprepitant-nSMase2 showed proper convergence. Following treatment with different concentrations of aprepitant in both cell-free and cell-dependent assays, nSMase2 activity was remarkably decreased. CONCLUSION: Aprepitant, at a concentration as low as 15 µM, was able to inhibit nSmase2 activity in HCT116 cells without any significant effects on their viability. Aprepitant is therefore suggested to be a potentially safe exosome release inhibitor.

Cardioprotective action of aprepitant in a rat model of ischemia-reperfusioninduced myocardial injury: role of PI3K-AkT-GSK-3ß-HIF-1α signaling pathway.

PURPOSE: The present study explored the role and mechanism involved in aprepitant-induced cardioprotective effects in rat model of ischemia-reperfusion injury. METHODS: The isolated hearts of Wistar male albino rats were subjected to ischemia-reperfusion injury on Langendorff apparatus. The extent of myocardial injury was assessed by measuring lactate dehydrogenase 1 and CK-MB release in the coronary effluent. The rats were treated with aprepitant (5, 10 and 20 mg/kg) before isolating hearts. After injury, the levels of HIF-1α, p-AkT, p-GSK-3ß/GSK-3ß were measured in heart homogenates. LY294002 was employed as PI3K inhibitor. RESULTS: Ischemia-reperfusion led to significant myocardial injury and decreased the levels of HIF-1α, p-AkT and ratio of p-GSK-3ß/GSK-3ß. Aprepitant attenuated myocardial injury and restored the biochemical changes in a dose-dependent manner. Pre-treatment with LY294002 (10 and 20 mg/kg) abolished aprepitant-mediated cardioprotective effects and restored the biochemical parameters in the heart homogenate. CONCLUSIONS: Aprepitant may be effective in preventing ischemia-reperfusion-induced myocardial injury, which may be due to activation of PI3K-AkT-GSK-3ß and HIF-1α signaling pathway.

The Effect of Blocking Neurokinin-1 Receptor by Aprepitant on the Inflammatory and Apoptosis Pathways in Human Ovarian Cancer Cells.

Ovarian cancer is the seventh most common cancer globally, and the second most common cancer among women with significant mortality. Toward this end, it is shown that substance P (SP) is involved in tumor initiation and progression through the neurokinin-1 receptor (NK1R). However, the exact molecular mechanism of the SP/NK1R system in ovarian cancer is not yet fully clarified. In this in vitro study, we decided to investigate the effect of the SP/NK1R system and blockage of NK1R by its specific antagonist (Aprepitant) on the proliferation of ovarian cancer cells as well as the alteration of inflammatory pathways. Our results revealed that Aprepitant stimulated apoptotic cell death and attenuated inflammation of ovarian cancer cells through the NF-kB and P53 signaling pathways. After treatment with Aprepitant, the expression of downstream anti-apoptotic genes related to the NF-kB pathway (survivine and bcl2) was decreased. However, we indicated the positive effect of SP on the proliferation of ovarian cancer cells by inducing the expression of NF-kB protein and NF-kB anti-apoptotic target genes. Moreover, Pro-apoptotic p53 target genes (P21 and Bax) were increased through aprepitant treatment, while SP attenuated these genes' expression. Besides, ROS generation in ovarian cancer cells after treatment with SP induced, while blocking of NK1R with Aprepitant reduced the level of ROS generation. Given this, our data suggest that this NK1R might be used as an important therapeutic target in ovarian cancer and Aprepitant could be considered a new drug in ovarian cancer therapy.

The Potential In Vitro Inhibitory Effects of Neurokinin-1 Receptor (NK-1R) Antagonist, Aprepitant, in Osteosarcoma Cell Migration and Metastasis.

Background: Osteosarcoma, the most frequent osteogenic malignancy, has become a serious public health challenge due to its high morbidity rates and metastatic potential. Recently, the neurokinin-1 receptor (NK-1R) is proved to be a promising target in cancer therapy. This study is aimed at determining the effect of aprepitant, a safe and Food and Drug Administration (FDA) approved NK-1R antagonist, on osteosarcoma cell migration and metastasis, and to explore its underlying mechanism of action. Methods: Colorimetric MTT assay was employed to assess cell viability and cytotoxicity. A wound-healing assay was used to examine migration ability. The desired genes' protein and mRNA expression levels were measured by western blot assay and quantitative real-time PCR (qRT-PCR), respectively. Gelatinase activity was also measured by zymography. Results: We found that aprepitant inhibited MG-63 osteosarcoma cell viability in a dose-dependent manner. We also observed that aprepitant inhibited the migrative phenotype of osteosarcoma cells and reduced the expression levels and activities of matrix metalloproteinases (MMP-2 and MMP-9). Aprepitant also reduced the expression of an angiogenic factor, VEGF protein, and NF-κB as an important transcriptional regulator of metastasis-related genes. Conclusion: Collectively, our observations indicate that aprepitant modulates the metastatic behavior of human osteosarcoma cells, which may be applied to an effective therapeutic approach for patients with metastatic osteosarcoma.

The effect of substance P and its specific antagonist (aprepitant) on the expression of MMP-2, MMP-9, VEGF, and VEGFR in ovarian cancer cells.

BACKGROUND: Substance P (SP) has a crucial role in cancer initiation and progression via binding to its specific receptor (NK1R). Various evidence confirmed the overexpression of NK1R and SP in the tissue of multiple cancers, including ovarian cancer. Despite numerous studies, the mechanism of the SP/NK1R system on migration and angiogenesis of ovarian cancer cells has not yet been deciphered. In this study, considering the critical factors in cell migration (MMP-2, MMP-9) and angiogenesis (VEGF, VEGFR), we investigated the possible mechanism of this system in inducing migration and angiogenesis of ovarian cancer cells. METHODS AND RESULTS: First, the resazurin assay was conducted to evaluate the cytotoxic effect of aprepitant (NK1R antagonist) on the viability of A2780 ovarian cancer cells. After that, the impact of this system and aprepitant on the mRNA expression of the factors mentioned above were studied using RT-PCR. Besides, the scratch assay was performed to confirm the effect of the SP/NK-1R system and aprepitant on cell migration. Our results implied that this system induced cell migration and angiogenesis by increasing the mRNA expression of MMP-2, MMP-9, VEGF, and VEGFR. The obtained results from the scratch assay also confirmed the positive effect of this system on cell migration. Meanwhile, the blocking of NK1R by aprepitant suppresses the SP effects on cell migration and angiogenesis. CONCLUSIONS: Overall, the SP/NK1R system plays a vital role in ovarian cancer progression, and the inhibition of NK1Rusing aprepitant could inhibit the spread of ovarian cancer cells through metastasis and angiogenesis.

Sustained endosomal release of a neurokinin-1 receptor antagonist from nanostars provides long-lasting relief of chronic pain.

Soft polymer nanoparticles designed to disassemble and release an antagonist of the neurokinin 1 receptor (NK1R) in endosomes provide efficacious yet transient relief from chronic pain. These micellar nanoparticles are unstable and rapidly release cargo, which may limit the duration of analgesia. We examined the efficacy of stable star polymer nanostars containing the NK1R antagonist aprepitant-amine for the treatment of chronic pain in mice. Nanostars continually released cargo for 24 h, trafficked through the endosomal system, and disrupted NK1R endosomal signaling. After intrathecal injection, nanostars accumulated in endosomes of spinal neurons. Nanostar-aprepitant reversed mechanical, thermal and cold allodynia and normalized nociceptive behavior more efficaciously than free aprepitant in preclinical models of neuropathic and inflammatory pain. Analgesia was maintained for >10 h. The sustained endosomal delivery of antagonists from slow-release nanostars provides effective and long-lasting reversal of chronic pain.

GPCR large-amplitude dynamics by 19F-NMR of aprepitant bound to the neurokinin 1 receptor.

Comparisons of G protein-coupled receptor (GPCR) complexes with agonists and antagonists based on X-ray crystallography and cryo-electron microscopy structure determinations show differences in the width of the orthosteric ligand binding groove over the range from 0.3 to 2.9 Å. Here, we show that there are transient structure fluctuations with amplitudes up to at least 6 Å. The experiments were performed with the neurokinin 1 receptor (NK1R), a GPCR of class A that is involved in inflammation, pain, and cancer. We used 19F-NMR observation of aprepitant, which is an approved drug that targets NK1R for the treatment of chemotherapy-induced nausea and vomiting. Aprepitant includes a bis-trifluoromethyl-phenyl ring attached with a single bond to the core of the molecule; 19F-NMR revealed 180° flipping motions of this ring about this bond. In the picture emerging from the 19F-NMR data, the GPCR transmembrane helices undergo large-scale floating motions in the lipid bilayer. The functional implication is of extensive promiscuity of initial ligand binding, primarily determined by size and shape of the ligand, with subsequent selection by unique interactions between atom groups of the ligand and the GPCR within the binding groove. This second step ensures the wide range of different efficacies documented for GPCR-targeting drugs. The NK1R data also provide a rationale for the observation that diffracting GPCR crystals are obtained for complexes with only very few of the ligands from libraries of approved drugs and lead compounds that bind to the receptors.

The substance P/ neurokinin-1 receptor signaling pathway mediates metastasis in human colorectal SW480 cancer cells.

BACKGROUND: The substance P (SP)/neurokinin-1 receptor (NK1R) system, a critical metastatic signaling pathway, can be targeted by substance P antagonists to prevent its cancer-progressive impacts. In the current study, we aimed to investigate the carcinogenic activity of the SP/NK1R system in human SW480 colorectal cancer cells and study the antagonistic impact of aprepitant (AP) by measuring MMP-2 and MMP-9 enzymatic activity. METHODS: Different concentrations of SP, alone or mixed by AP, were utilized to treat SW480 cells to investigate the cells' viability and metastasis by applying Resazurin and Gelatin Zymography methods, respectively. The cells' metastatic response was analyzed by measuring the MMP-2 and MMP-9 in transcriptional and translational levels. Finally, the Scratch assay was carried out to evaluate the cells' metastatic response following the SP/AP treatment. RESULTS: A significant metastatic activity was observed in SW480 cells following incubation with the increasing SP doses by detecting MMP-2/MMP-9 enzyme activity, genes overexpression, and enhanced cell migration. This is while the AP treatment meaningfully diminished all the SP-mediated metastatic effects (p-Value < 0.001). CONCLUSIONS: According to the results, the SP/NKR1 signaling pathway can be considered one of the main metastatic effectors in human colorectal cancer. Therefore, AP might be suggested to be used as the SP antagonist and an efficient anti-metastatic drug.

The Therapeutic Potential of Aprepitant in Glioblastoma Cancer Cells through Redox Modification.

Although there is no doubt regarding the involvement of oxidative stress in the development of glioblastoma, many questions remained unanswered about signaling cascades that regulate the redox status. Given the importance of the substance P (SP)/neurokinin 1 receptor (NK1R) system in different cancers, it was of particular interest to evaluate whether the stimulation of this cascade in glioblastoma-derived U87 cells is associated with the induction of oxidative stress. Our results showed that SP-mediated activation of NK1R not only increased the intracellular levels of malondialdehyde (MDA) and reactive oxygen species (ROS) but also reduced the concentration of thiol in U87 cells. We also found that upon SP addition, there was a significant reduction in the cells' total antioxidant capacity (TAC), revealing that the SP/NK1R axis may be involved in the regulation of oxidative stress in glioblastoma cells. The significant role of SP/NK1R in triggering oxidative stress in glioblastoma has become more evident when we found that the abrogation of the axis using aprepitant reduced cell survival, probably through exerting antioxidant effects. The results showed that both MDA and ROS concentrations were significantly reduced in the presence of aprepitant, and the number of antioxidant components of the redox system increased. Overall, these findings suggest that aprepitant might exert its anticancer effect on U87 cells through shifting the balance of oxidant and antioxidant components of the redox system.