CCK Receptor Research Articles

Deoxynivalenol (Vomitoxin)-Induced Anorexia Is Induced by the Release of Intestinal Hormones in Mice.

Deoxynivalenol (DON), also known as vomitoxin, is a mycotoxin that can cause antifeeding and vomiting in animals. However, the mechanism of DON inducing anorexia is complicated. Studies have shown that intestinal hormones play a significant part in the anorexia caused by DON. We adopted the “modeling of acute antifeeding in mice” as the basic experimental model, and used two methods of gavage and intraperitoneal injection to explore the effect of intestinal hormones on the antifeedant response induced by DON in mice. We found that 1 and 2.5 mg/kg·bw of DON can acutely induce anorexia and increase the plasma intestinal hormones CCK, PYY, GIP, and GLP-1 in mice within 3 h. Direct injection of exogenous intestinal hormones CCK, PYY, GIP, and GLP-1 can trigger anorexia behavior in mice. Furthermore, the PYY receptor antagonist JNJ-31020028, GLP-1 receptor antagonist Exendin(9-39), CCK receptor antagonist Proglumide, GIP receptor antagonist GIP(3-30)NH2 attenuated both intestinal hormone and DON-induced anorectic responses. These results indicate that intestinal hormones play a critical role in the anorexia response induced by DON.

The Characteristic of Critical Genes in Neuroendocrine System and Their Regulation on Food Habit Transition and Metamorphosis of Veined Rapa Whelk Rapana venosa (Valenciennes, 1846)

Metamorphosis is a critical developmental event in mollusks, and neuroendocrine system plays an essential role in this process. Rapana venosa is an economically important shellfish in China, but the artificial technology of R. venosa aquaculture is limited by metamorphosis. As a carnivorous gastropod, food habit transition makes the mechanism of R. venosa metamorphosis more complex. To investigate the changes in the neuroendocrine system and to reveal its role in regulating the food habit transition and metamorphosis of R. venosa, we cloned the cDNA sequences encoding 5-hydroxytryptamine receptor (Rv-5HTR), nitric oxide synthetase (Rv-NOS) and cholecystokinin receptor (Rv-CCKR), and investigated their expression by quantitative real-time PCR analysis, and explore the spatio-temporal changes of 5-HT protein expression using Immunohistochemical (IHC) analysis. The expression of the three geens was significantly increased in the early intramembrane veliger stage, which indicates that the three genes are related to the development of digestive system. Additionally, expression of the three genes was decreased after metamorphosis, while Rv-NOS and Rv-CCKR were increasingly expressed in competent larvae, which may help the larvae find suitable environments and promote digestive system development for metamorphosis, and the result of 5-HT IHC analysis also reflects the development of neuroendocrine system. Furthermore, results show that CCK can effect the expression of digestive enzyme, NOS and 5-HT receptor. Finally, based on the present results, we hypothesized that CCK and CCK receptor may be critical regulatory factors of food habit transition and metamorphosis. These results might provide information on the development of neuroendocrine system of R. venosa, and new insight into the regulation of the food habit transition and metamorphosis of gastropods.

The Insight into Insulin-Like Growth Factors and Insulin-Like Growth-Factor-Binding Proteins and Metabolic Profile in Pediatric Obesity.

Insulin-like growth factors (IGFs) and insulin-like growth-factor-binding proteins (IGFBPs) regulate cell proliferation and differentiation and may be of importance in obesity development. The aim of the study was to analyze the expression of chosen IGF-axis genes and the concentration of their protein products in 28 obese children (OB) and 34 healthy control (HC), and their correlation with essential parameters associated with childhood obesity. The gene expression of IGFBP7 was higher, and the expression of IGF2 and IGFBP1 genes was lower in the OB. The expression of IGFBP6 tended to be lower in OB. IGFBP4 concentration was significantly higher, and IGFBP3 tended to be higher in the OB compared to the HC, while IGFBP1, IGFBP2, and IGFBP6 were significantly lower, and IGFBP7 tended to be lower in OB. We found numerous correlations between IGFs and IGFBP concentration and obesity metabolic parameters. IGFBP6 correlated positively with apelin, cholecystokinin, glucagone-like peptide-1, and leptin receptor. These peptides were also significantly lower in obese children in our study. The biological role of decreased levels of IGFBP6 in obese children needs further investigation.

Abstract 1931: Evaluation of cholecystokinin receptor expression and function in non-small cell lung cancer

Abstract Introduction: Lung cancer is the leading cause of cancer-related death in the United States, with nearly 85% of cases histologically classified as non-small cell lung cancer (NSCLC). A recent epidemiological study provides evidence of an increased risk ratio for NSCLC in association with adherence to a Western dietary pattern, which has a higher dietary fat content, compared to vegetarian or Tex-Mex dietary patterns. Our laboratory and others have measured increased tumor burden in murine models of NSCLC when feeding diets are high in fat compared to a standard rodent cereal diet. In our laboratory, increased tumor burden is associated with increased tumor multiplicity, but no change in proliferative markers (Ki-67 and pS6) were observed. Dietary fat increases both tumorigenesis in mouse models of pancreatic cancer and circulating levels of the gastrointestinal hormone cholecystokinin (CCK). Inhibition of CCK receptors antagonizes progression of precursor lesions in murine models of pancreatic cancer. Although CCK receptor expression has not been detected in normal human pulmonary tissue, the CCK-B receptor has been identified immunohistologically in human lung tumors. Further, the CCK-A receptor is expressed in normal murine pulmonary epithelium. Hypothesis: At concentrations approximating circulating levels in mice fed a high fat diet, CCK reduces apoptosis of NSCLC cells in vitro. Materials and Methods: The murine NSCLC cell line IO33 was grown in serum-replete RPMI 1640 or serum-deprived RPM 1640 with 0 to 1000 pM exogenous CCK-8 peptide added. Expression of CCK-A and CCK-B receptors were measured in IO33 and human NSCLC cell lines by qPCR. Apoptosis was assessed using a Caspase-3/7 peptide cleavage assay. Results: Expression of CCK-A and CCK-B receptors are reported in IO33 cells and 11 human lung cancer cell lines. Apoptosis of IO33 cells was significantly decreased by greater than 30% at CCK concentrations greater than 100pM (P < 0.03), which is the circulating concentration of CCK in mice fed a high fat diet. Conclusions: CCK receptors are expressed in human and murine NSCLC cells and circulating CCK may reduce NSCLC cell apoptosis. Citation Format: Alexander C. Miller, Shelby J. Subry, Manijeh M. Assar, Krista Pearman, Jeffrey William Norris. Evaluation of cholecystokinin receptor expression and function in non-small cell lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1931.

Radiopharmaceutical Formulation and Preclinical Testing of 68Ga-Labeled DOTA-MGS5 for the Regulatory Approval of a First Exploratory Clinical Trial.

The new minigastrin analog DOTA-MGS5 is a promising new candidate for targeting cholecystokinin-2 receptor (CCK2R)-expressing tumors. To enable the clinical translation of PET/CT imaging using 68Ga-labeled DOTA-MGS5, different quality and safety aspects need to be considered to comply with the regulatory framework for clinical trial application. The preparation of the radiopharmaceutical was established using a cassette-based automated synthesis unit. Product specifications, including analytical procedures and acceptance criteria, were adopted from Ph. Eur. monographs for other 68Ga-labeled radiopharmaceuticals. Non-clinical studies included receptor affinity and cell uptake studies using two different CCK2R-expressing cell lines, as well as pharmacokinetic biodistribution studies in BALB/c mice for dosimetry calculations and toxicological studies in Wistar rats. The produced masterbatches fulfilled the defined acceptance criteria. DOTA-MGS5, with confirmed affinity to the CCK2R, showed a high specific cell uptake and no interaction with other receptors in vitro when radiolabeled with gallium-68. Favorable in vivo properties were observed in biodistribution and dosimetry studies. An effective dose of ~0.01 mSv/MBq was estimated for humans utilizing OLINDA/EXM software. A maximum peptide dose of 50 µg was established for the initial clinical dose based on the toxicity study in rats. The standardized production of [68Ga]Ga-DOTA-MGS5 using an automated synthesis module and the performed non-clinical safety studies support a first exploratory clinical trial with this new PET imaging agent.

Roles of Cholecystokinin in the Nutritional Continuum. Physiology and Potential Therapeutics

Cholecystokinin is a gastrointestinal peptide hormone with important roles in metabolic physiology and the maintenance of normal nutritional status, as well as potential roles in the prevention and management of obesity, currently one of the dominant causes of direct or indirect morbidity and mortality. In this review, we discuss the roles of this hormone and its receptors in maintaining nutritional homeostasis, with a particular focus on appetite control. Targeting this action led to the development of full agonists of the type 1 cholecystokinin receptor that have so far failed in clinical trials for obesity. The possible reasons for clinical failure are discussed, along with alternative pharmacologic strategies to target this receptor for prevention and management of obesity, including development of biased agonists and allosteric modulators. Cellular cholesterol is a natural modulator of the type 1 cholecystokinin receptor, with elevated levels disrupting normal stimulus-activity coupling. The molecular basis for this is discussed, along with strategies to overcome this challenge with a corrective positive allosteric modulator. There remains substantial scope for development of drugs to target the type 1 cholecystokinin receptor with these new pharmacologic strategies and such drugs may provide new approaches for treatment of obesity.

Changes in the interstitial cells of Cajal in the gallbladder of guinea pigs fed a lithogenic diet.

Cholesterol cholelithiasis is a common disease and gallbladder hypomotility may underlie its pathogenesis. Interstitial cells of Cajal (ICCs) in the gallbladder serve vital roles in regulating gallbladder motility. The aim of the present study was to investigate changes in gallbladder ICCs during the development of cholesterol cholelithiasis. A total of 40 male guinea pigs were randomly assigned to four groups and fed a standard diet (SD) or lithogenic diet (LD) for 2 or 8 weeks. The LD significantly increased the total cholesterol levels in the serum and bile, as well as the serum levels of high-density lipoprotein-cholesterol and low-density lipoprotein-cholesterol after 2 and 8 weeks. The LD also significantly increased and decreased the number of gallbladder ICCs at 2 and 8 weeks, respectively, by regulating the stem cell factor/C-kit pathway. Moreover, the ultrastructure of gallbladder ICCs was significantly altered after 8 weeks, and the protein expression levels of connexin 43 in the gallbladder were differentially altered after 2 and 8 weeks. Finally, cholecystokinin receptor type A (CCK1R) expression in the gallbladder was assessed. In gallbladder ICCs, its expression was significantly increased and decreased after 2 and 8 weeks, respectively. In conclusion, these results demonstrate that the density, ultrastructure and CCK1R expression levels of gallbladder ICCs are differentially altered at various stages of cholesterol cholelithiasis progression, indicating that gallbladder ICCs may be considered a potential therapeutic target for treatment of cholesterol cholelithiasis.

Improved Tumor-Targeting with Peptidomimetic Analogs of Minigastrin 177Lu-PP-F11N

Simple SummarySeveral radiolabeled peptides targeting CCK2R-positive types of cancer (such as medullary thyroid cancer and small cell lung cancer) have been reported in the last 25 years, some of which have entered clinical trials. In an effort to improve its tumor-targeting properties, we applied chemical modifications to the backbone of the peptide 177Lu-PP-F11N, an analog of minigastrin in clinical trials. The generated radiolabeled peptidomimetics showed significantly improved characteristics in mice bearing CCK2R-positive tumor xenografts, such as higher tumor uptake, slower tumor washout, and increased tumor-to-kidney ratios. These properties make the novel compounds promising candidates for the imaging and therapy of CCK2R-positive tumors and metastases.The cholecystokinin-2 receptor (CCK2R) is an attractive target in nuclear medicine due to its overexpression by different tumors. Several radiolabeled peptidic ligands targeting the CCK2R have been investigated in the past; however, their low stability against proteases can limit their uptake in tumors and metastases. Substitution of single or multiple amide bonds with metabolically stable 1,4-disubstituted 1,2,3-triazoles as amide bond bioisosteres proved a promising strategy for improving the tumor-targeting properties of a truncated analog of minigastrin. In this study, we applied the previously studied structural modifications to improve the pharmacokinetic and pharmacodynamic properties of PP-F11N, a minigastrin analog currently in clinical trials. Novel minigastrins (NMGs) as analogs of PP-F11N with one or two amide bonds substituted by 1,2,3-triazoles were synthesized, radiolabeled with 177Lu3+, and subjected to full evaluation in vitro (cell internalization, receptor affinity, stability in blood plasma) and in vivo (stability, biodistribution, SPECT/CT imaging). NMGs with triazoles inserted between the amino acids DGlu10-Ala11 and/or Tyr12-Gly13 showed a significantly increased cellular uptake and affinity toward the CCK2R in vitro. Resistance against the metabolic degradation of the NMGs was comparable to those of the clinical candidate PP-F11N. Imaging by SPECT/CT and biodistribution studies demonstrated a higher uptake in CCK2R-positive tumors but also in the CCK2R-positive stomach. The peptidomimetic compounds showed a slow tumor washout and high tumor-to-kidney ratios. The structural modifications led to the identification of analogs with promising properties for progression to clinical applications in the diagnosis and therapy of CCK2R-positive neoplasms.

Expression of Cholecystokinin and its Receptors in the Intestinal Tract of Type 2 Diabetes Patients and Healthy Controls.

Cholecystokinin (CCK) is a gut hormone originally known for its effects on gallbladder contraction and release of digestive enzymes. CCK, however, also mediates satiety and stimulate insulin secretion. Knowledge of the distribution of CCK-producing enteroendocrine cells (I cells) in humans is sparse. The general notion, based on animal data, is that I cells are present mainly in the proximal small intestine. We examined the occurrence of I cells (immunohistochemically) and the expression of CCK messenger RNA (mRNA) as well as CCK1 and CCK2 receptor mRNA along the intestines in healthy individuals and patients with type 2 diabetes. Mucosal biopsies collected with 30-cm intervals in the small intestine and from seven anatomical locations in the large intestine (using double-balloon enteroscopy) from 12 patients with type 2 diabetes and 12 gender-, age-, and body mass index-matched healthy individuals were analyzed using mRNA sequencing and immunohistochemical staining. We observed a gradual decrease in CCK mRNA expression and density of CCK-immunoreactive cells from duodenum to ileum. Very few CCK-immunoreactive cells and nearly undetectable CCK mRNA expression were found in the large intestine. No significant differences were seen between the groups. Expression of CCK receptors was observed in the duodenum of both groups. Both density of CCK cells and expression of CCK mRNA decreased through the small intestine in both groups with low levels in the large intestine. Patients with type 2 diabetes did not have altered density of CCK cells or expression of CCK mRNA in intestinal mucosa.

Pseudomonas Exotoxin A-Based Immunotherapy Targeting CCK2R-Expressing Colorectal Malignancies: An In Vitro and In Vivo Evaluation.

Cholecystokinin-2 receptor (CCK2R) has been proven to be a specific biomarker for colorectal malignancies. Immunotoxins are a valuable class of immunotherapy agents consisting of a targeting element and a bacterial or plant toxin. Previous work demonstrated that targeting CCK2R is a good therapeutic strategy for the treatment of colorectal cancer (CRC). In the present study, we developed a new version of CCK2R-targeting immunotoxin GD9P using a targeted peptide, GD9, as the binding motif and a truncated Pseudomonas exotoxin A (PE38) as the cytokiller. BALB/c nude mice were treated with different doses of GD9P, and pharmacodynamics, pharmacokinetic, and toxicological data were obtained throughout this study. Compared to the parental immunotoxin rCCK8PE38, GD9P exhibited about 1.5-fold yield, higher fluorescence intensity, and increased antitumor activity against human CRC in vitro and in vivo. The IC50 values of GD9P in vitro ranged from 1.61 to 4.55 nM. Pharmacokinetic studies were conducted in mice with a T1/2 of 69.315 min. When tumor-bearing nude mice were treated with GD9P at doses ≥2 mg/kg for five doses, a rapid shrinkage in tumor volume and, in some cases, complete remission was observed. A preliminary safety evaluation demonstrated a good safety profile of GD9P as a Pseudomonas exotoxin A-based immunotherapy. The therapy in combination with oxaliplatin can increase the antitumor efficacy and reduce the toxic side effects caused by chemotherapy. In conclusion, the data support the use of GD9P as a promising immunotherapy targeting CCK2R-expressing colorectal malignancies.

Benefits of Sustained Upregulated Unimolecular GLP-1 and CCK Receptor Signalling in Obesity-Diabetes

Combined activation of GLP-1 and CCK1 receptors has potential to synergistically augment the appetite-suppressive and glucose homeostatic actions of the individual parent peptides. In the current study, pancreatic beta-cell benefits of combined GLP-1 and CCK1 receptor upregulation were established, before characterising bioactivity and antidiabetic efficacy of an acylated dual-acting GLP-1/CCK hybrid peptide, namely [Lys12Pal]Ex-4/CCK. Both exendin-4 and CCK exhibited (p<0.001) proliferative and anti-apoptotic effects in BRIN BD11 beta-cells. Proliferative benefits were significantly (p<0.01) augmented by combined peptide treatment when compared to either parent peptide alone. These effects were linked to increases (p<0.001) in GLUT2 and glucokinase beta-cell gene expression, with decreased (p<0.05-p<0.001) expression of NFκB and BAX. [Lys12Pal]Ex-4/CCK exhibited prominent insulinotropic actions in vitro, coupled with beneficial (p<0.001) satiety and glucose homeostatic effects in the mice, with bioactivity evident 24 h after administration. Following twice daily injection of [Lys12Pal]Ex-4/CCK for 28 days in diabetic high fat fed (HFF) mice with streptozotocin (STZ)-induced compromised beta-cells, there were clear reductions (p<0.05-p<0.001) in energy intake and body weight. Circulating glucose was returned to lean control concentrations, with associated increases (p<0.001) in plasma and pancreatic insulin levels. Glucose tolerance and insulin secretory responsiveness were significantly (p<0.05-p<0.001) improved by hybrid peptide therapy. In keeping with this, evaluation of pancreatic histology revealed restoration of normal islet alpha- to beta-cell ratios and reduction (p<0.01) in centralised islet glucagon staining. Improvements in pancreatic islet morphology were associated with increased (p<0.05) proliferation and reduced (p<0.001) apoptosis of beta-cells. Together, these data highlight the effectiveness of sustained dual GLP-1 and CCK1 receptor activation by [Lys12Pal]Ex-4/CCK for the treatment of obesity-related diabetes.

The Effect of Proton Pump Inhibitors on Insulin-Glucose Homeostasis in Patients With Type 2 Diabetes Mellitus

Introduction: Gastrin release from G cells stimulates cholecystokinin (CCK2) receptors throughout the body, most of which promote gastric acid secretion. However, gastrin also stimulates CCK2 receptors located elsewhere, including the islet of the pancreas. In turn, gastrin increases insulin secretion1. Gastrin also promotes pancreatic β cell neogenesis and replication. Proton pump inhibitors (PPIs) decrease the pH of the stomach and stimulate gastrin secretion, which may indirectly promote insulin secretion and improve hemoglobin A1c (HbA1c). Objective: To understand the effect of PPIs on insulin-glucose homeostasis (c-peptide, HbA1c, and glucose) in patients with type 2 diabetes mellitus (T2DM). Methodology: We retrospectively reviewed the charts of patients with T2DM at least 18 years of age who received care at AnMed Health facilities from Jan. 1, 2018 through Dec. 31, 2018 to compare HbA1c, C‐peptide, and glucose levels in patients with and without active PPI therapy. Slicer-dicer software was used to identify study population with diagnosis of T2DM and labs including both HbA1c and C-peptide. Out of total 215 patients satisfying inclusion criteria, 71 patients were on PPI. Statistical analyses were performed using SPSS version 20.0 (SPSS, Armonk, NY: IBM Corp). All values are presented as means ± SD. A p value of < 0.05 was considered to be significant. Independent T-test and chi-square test were performed to compare parameters in between groups. Results: The PPI and non‐PPI groups had no statistical difference regarding age, sex, race and BMI. There was no significant difference in HbA1c levels between PPI and non‐PPI groups (8.6% ± 2.1 vs 8.3% ± 2.0, respectively; p value = 0.37). However, we found a significant increase in C‐peptide levels (3.1 ng/mL ± 2.4 vs 2.4 ng/mL ± 2.3; p value = 0.037) and decrease in LDL levels (79.6 mg/dL ± 34.0 vs 89.73 mg/dL ± 32.9; p value = 0.046) in the PPI group compared to non-PPI group. In addition, there was a significantly greater prevalence of coronary artery disease in the PPI group (p = 0.01). Conclusion: PPI therapy in patients with T2DM was not associated with improved glycemic control. However, C-peptide levels were significantly higher in patients with T2DM who were on PPI therapy suggesting higher insulin secretion. The lack of difference in HbA1c levels may be a result of aggressive diabetic management by treating clinicians to achieve similar goal HbA1c in both groups. Further research is needed to understand the gastrin pathway as a potential option for improving glycemic control.

SP-128 - Structure-activity relationship study of peptide Gastramide-01 (GA-01), a potent binder of cholecystokinin-2 (CCK-2) receptor and theranostic candidate

SP-128 - Structure-activity relationship study of peptide Gastramide-01 (GA-01), a potent binder of cholecystokinin-2 (CCK-2) receptor and theranostic candidate

Rapid Increase of Gastrointestinal P-Glycoprotein Functional Activity in Response to Etoposide Stimulation.

We previously reported that exposure of human colon adenocarcinoma (Caco-2) cells to the bitter substance phenylthiocarbamide (PTC) rapidly enhanced the transport function of P-glycoprotein (P-gp). In this study, we investigated the short-term effect of etoposide, another bitter-tasting P-gp substrate, on P-gp transport function in the same cell line. We found that etoposide exposure significantly increased both the P-gp protein level in the plasma membrane fraction and the efflux rate of rhodamine123 (Rho123) in Caco-2 cells within 10 min. The efflux ratio (ratio of the apparent permeability coefficient in the basal-to-apical direction to that in the apical-to-basal direction) of Rho123 in etoposide-treated cells was also significantly increased compared with the control. These results indicated that etoposide rapidly enhances P-gp function in Caco-2 cells. In contrast, P-gp expression in whole cells at both the mRNA and protein level was unchanged by etoposide exposure, compared with the levels in non-treated cells. Furthermore, etoposide increased the level of phosphorylated ezrin, radixin and moesin (P-ERM) proteins in the plasma membrane fraction of Caco-2 cells within 10 min. P-gp functional changes were blocked by YM022, an inhibitor of cholecystokinin (CCK) receptor. These results suggest that etoposide induces release of CCK, causing activation of the CCK receptor followed by phosphorylation of ERM proteins, which recruit intracellular P-gp for trafficking to the gastrointestinal membrane, thereby increasing the functional activity of P-gp.

SSP-05 - Next generation therapeutic radiopharmaceuticals targeting cholecystokinin-2 receptor positive cancers

SSP-05 - Next generation therapeutic radiopharmaceuticals targeting cholecystokinin-2 receptor positive cancers

SSP-06 - Foldamers afford improved theranostics targeting cholecystokinin-2 receptor positive cancers

SSP-06 - Foldamers afford improved theranostics targeting cholecystokinin-2 receptor positive cancers

Hypoxia transactivates cholecystokinin gene expression in 3D-engineered muscle

At high altitudes, the hypoxic atmosphere decreases the oxygen partial pressure in the body, inducing several metabolic changes in tissues and cells. Furthermore, it exerts potent anorectic effects, thus causing an energy deficit. Two decades ago, a marked increase in the resting level of plasma cholecystokinin (CCK) was observed in humans at the Mt. Kanchenjunga basecamp, located at 5100m above the sea level, compared to sea-level control values. Interestingly, acute exercise also raises plasma CCK and exerts potent anorectic effects under normoxic conditions. However, the transcriptional regulations of Cck gene underlying these effects have not yet been established. Here, we employed acute electrical pulse stimulation (EPS) followed by microarray analysis to discover novel myokines in 3D-engineered muscle. Acute EPS affects the contractile function, inducing a decline in the contractile force. Surprisingly, microarray analysis revealed an EPS-induced activation of cholecystokinin receptor (CCKR)-mediated signaling. Furthermore, Cck was constitutively upregulated in 3D-engineered muscle, and its expression increased under hypoxic conditions. Notably, a hypoxia-responsive element was detected in the Cck promoters of mice and humans. Our results suggested that hypoxia transactivated Cck expression in 3D-engineered muscle. Furthermore, the elevation in plasma CCK levels following acute exercise or at high altitude might be partly attributed to myogenic cells.

A New Turn in Peptide-Based Imaging Agents: Foldamers Afford Improved Theranostics Targeting Cholecystokinin-2 Receptor-Positive Cancer.

Proteins adopt unique folded secondary and tertiary structures that are responsible for their remarkable biological properties. This structural complexity is key in designing efficacious peptides that can mimic the three-dimensional structure needed for biological function. In this study, we employ different chemical strategies to induce and stabilize a β-hairpin fold of peptides targeting cholecystokinin-2 receptors for theranostic application (combination of a targeted therapeutic and a diagnostic companion). The newly developed peptides exhibited enhanced folding capacity as demonstrated by circular dichroism (CD) spectroscopy, ion-mobility spectrometry-mass spectrometry, and two-dimensional (2D) NMR experiments. Enhanced folding characteristics of the peptides led to increased biological potency, affording four optimal Ga-68 labeled radiotracers ([68Ga]Ga-4b, [68Ga]Ga-11b-13b) targeting CCK-2R. In particular, [68Ga]Ga-12b and [68Ga]Ga-13b presented improved metabolic stability, enhanced cell internalization, and up to 6 fold increase in tumor uptake. These peptides hold great promise as next-generation theranostic radiopharmaceuticals.

Pain, Pathophysiological Mechanisms, and New Therapeutic Options for Alternative Analgesic Agents in Sheep: A Review and Investigation.

Simple SummaryThe purpose of this review is to provide current data on the definition and types of pain, describe its neuropharmacological and pathological mechanisms, and present a comparative analysis of the results obtained after intracerebroventricular (ICV) infusion of voltage-gated calcium channel inhibitors (VGCCIs) such as diltiazem, nifedipine, and verapamil, cholecystokinin receptor antagonists (PD, proglumide), and glutamatergic receptor antagonists (L-AP3, DL-AP3) during experimental distension of the duodenal and/or descending colonic wall. This method was used as a model for suppressing pain in sheep based on viscero-visceral inhibitory reflex, measured by the inhibition of behavioral symptoms of stress, degree of reticulo-ruminal motility, and changes in plasma cortisol and catecholamine concentration. After ICV infusion, all tested substances suppressed, to varying degrees, the viscero-visceral inhibitory reflex, tachycardia, hyperventilation, bleating, and gnashing of the teeth, whereas they increased the levels of cortisol and plasma catecholamines in sheep. These substances could be potential non-narcotic agents for the treatment of visceral intestinal pain (intestinal colic) in sheep, but clinical confirmation of the substances’ efficacy for treating intestinal colic is needed.Relief from suffering is the guiding principle of medical and veterinary ethics. Medical care for animals should be carried out to meet all welfare conditions. The need for pain management is demonstrated by recent monographs devoting attention to this urgent ethical need. Little data, however, are available on the prevention and attenuation of pain in sheep. After administration of narcotic analgesics used for severe visceral pain, sheep react with a state of excitement. Therefore, it was decided to experimentally investigate the usefulness of potential non-narcotic drugs to relieve pain in sheep with intestinal colic caused by 10 min of mechanical distension of their duodenal and/or descending colonic wall. The results indicate the potential usefulness of VGCCIs (diltiazem, nifedipine, verapamil), cholecystokinin receptor antagonists (PD, proglumide), and metabotropic glutaminergic receptor antagonists (mGluRAs), such as L-AP3, DL-AP3. As a premedication, these substances prevented the occurrence of symptoms of acute intestinal pain including atony of reticulo-rumen, tachycardia, hyperventilation, moaning, gnashing of teeth, hypercortisolemia, and catecholaminemia; hence, these substances are considered potential agents in the treatment of sheep visceral pain.

1,5-Disubstituted 1,2,3-Triazoles as Amide Bond Isosteres Yield Novel Tumor-Targeting Minigastrin Analogs.

1,5-Disubstituted 1,2,3-triazoles (1,5-Tz) are considered bioisosteres of cis-amide bonds. However, their use for enhancing the pharmacological properties of peptides or proteins is not yet well established. Aiming to illustrate their utility, we chose the peptide conjugate [Nle15]MG11 (DOTA-dGlu-Ala-Tyr-Gly-Trp-Nle-Asp-Phe-NH2) as a model compound since it is known that the cholecystokinin-2 receptor (CCK2R) is able to accommodate turn conformations. Analogs of [Nle15]MG11 incorporating 1,5-Tz in the backbone were synthesized and radiolabeled with lutetium-177, and their pharmacological properties (cell internalization, receptor binding affinity and specificity, plasma stability, and biodistribution) were evaluated and compared with [Nle15]MG11 as well as their previously reported analogs bearing 1,4-disubstituted 1,2,3-triazoles. Our investigations led to the discovery of novel triazole-modified analogs of [Nle15]MG11 with nanomolar CCK2R-binding affinity and 2-fold increased tumor uptake. This study illustrates that substitution of amides by 1,5-disubstituted 1,2,3-triazoles is an effective strategy to enhance the pharmacological properties of biologically active peptides.