Effect of BI 1358894 on Cholecystokinin-Tetrapeptide (CCK-4)-Induced Anxiety, Panic Symptoms, and Stress Biomarkers: A Phase I Randomized Trial in Healthy Males.

INTRODUCTION: Depression, anxiety, and/or panic disorder are often comorbid and have a complex etiology mediated through the same neuronal network. Cholecystokinin-tetrapeptide (CCK-4), a synthetic analog of the endogenous neuropeptide cholecystokinin (CCK), is thought to be implicated in this network. The CCK-4 challenge model is an accepted method of investigating the pathophysiology of panic and has been shown to mediate neuronal activation via the transient receptor potential canonical (TRPC) ion channels. OBJECTIVES: This study aimed to assess the pharmacodynamic effects of BI 1358894, a small-molecule inhibitor of TRPC ion channel members 4 and 5 (TRPC4/5), on CCK-4-induced anxiety/panic-like symptoms and evaluate circuit engagement. METHODS: Twenty healthy male CCK-4-sensitive volunteers entered a Phase I, double blind, randomized, two-way cross-over, single dose, placebo-controlled trial. Randomization was to oral BI 1358894 100 mg in the fed state followed by oral placebo in the fed state, or vice versa. Treatments were administered 5 h prior to intravenous CCK-4 50 µg. The primary endpoint was maximum change from baseline of the Panic Symptom Scale (PSS) sum intensity score after CCK-4 injection. Further endpoints included the emotional faces visual analog score (EVAS), the Spielberger State-Trait Anxiety Inventory (STAI), plasma adrenocorticotropic hormone (ACTH), and serum cortisol values. The safety and tolerability of BI 1358894 was assessed based on a number of parameters including occurrence of adverse events (AEs). All pharmacodynamic, pharmacokinetic, and safety endpoints were analyzed using descriptive statistics. RESULTS: Single oral doses of BI 1358894 were generally well tolerated by the healthy male volunteers included in this study. Adjusted mean maximum change from baseline in PSS sum intensity score was 24.4 % lower in volunteers treated with BI 1358894 versus placebo, while adjusted mean maximum change from baseline of EVAS was reduced by 19.2 % (BI 1358894 vs placebo). The STAI total score before CCK-4 injection was similar in both groups (placebo: 25.1; BI 1358894: 24.3). Relative to placebo, BI 1358894 reduced CCK-4-induced mean maximum plasma ACTH and serum cortisol values by 58.6 % and 27.3 %, respectively. Investigator-assessed drug-related AEs were reported for 13/20 participants (65.0 %). There were no serious or severe AEs, AEs of special interest, AEs leading to discontinuation of trial medication, or deaths. CONCLUSIONS: Overall, BI 1358894 reduced psychological and physiological responses to CCK-4 compared with placebo, as measured by PSS, subjective EVAS and objectively measured stress biomarkers. BI 1358894 had a positive safety profile, and single oral doses were well tolerated by the healthy volunteers. This trial (NCT03904576/1402-0005) was registered on Clinicaltrials.gov on 05.04.19.

Experimental panic provocation in healthy man-a translational role in anti-panic drug development?

Experimental neurochemical provocation of panic attacks in susceptible human subjects has considerably expanded our knowledge of the pathophysiology and psychopharmacology of panic disorder. Some panicogens also elicit short-lived panic-like states in healthy man. This offers the opportunity to assess the anti-panic action of drugs in proof-of-concept studies. However, from current data it is still unclear whether experimental panic in healthy man is a valid translational model. Most such studies in healthy volunteers have been performed using a cholecystokinin tetrapeptide (CCK-4) challenge. While CCK-4 panic was blocked by alprazolam pretreatment, escitalopram showed negative results in healthy man. Preliminary findings on novel investigational drugs and a few problematic results will be reviewed. Small sample sizes in many panic provocation studies, lack of dose-response aspects, and still-insufficient knowledge about the biological underpinning of experimental and spontaneous panic limit the interpretation of existing findings and should inspire further research.

Copeptin in CCK-4-induced panic in healthy man: Sexual dimorphisms in secretion pattern and panic response, but no correlation of copeptin with panic symptoms.

Copeptin, the C-terminal part of the hypothalamic arginine vaspopressin (AVP) precursor, closely mirrors the production of AVP and was proposed as an easily measured novel marker of the individual stress level in man. First data in male volunteers proposed copeptin as a potential endocrine surrogate marker of cholecystokinin-tetrapeptide (CCK-4)-induced panic. We tried to replicate these pilot data and to extend them to the other sex. 46 healthy human subjects (29 men, 17 women) were given an intravenous bolus of 50 µg CCK-4. Basal and stimulated plasma copeptin was measured and panic symptoms were assessed using the Acute Panic Inventory (API). Basal copeptin was significantly lower in women vs. men, while men showed a significantly higher CCK-4-induced increase of copeptin. In contrast, female subjects displayed a signifcantly higher increase of API ratings by CCK-4. No significant correlations of panic symptoms and copeptin release induced by CCK-4 could be found, neither in man, nor in women, nor in the total sample. A sexual dimorphism in copeptin secretion and in panic response was demonstrated. Prior unexpected findings of copeptin release as an objective read-out of panic could not be replicated. The role of the vasopressinergic system in panic anxiety needs further study in panic patients and in healthy man, using also other panic provocation paradigms.

Association testing of panic disorder candidate genes using CCK-4 challenge in healthy volunteers.

Despite continuing efforts to determine genetic vulnerability to panic disorder (PD), the studies of candidate genes in this disorder have produced inconsistent or negative, results. Laboratory panic induction may have a potential in testing genetic substrate of PD. In this study we aimed to explore the effects of several genetic polymorphisms previously implicated in PD on the susceptibility to cholecystokinin-tetrapeptide (CCK-4) challenge in healthy subjects. The study sample consisted of 110 healthy volunteers (47 males and 63 females, mean age 22.2 +/- 5.2) who participated in CCK-4 challenge test. Nine gene-candidates, including 5-HTTLPR, MAO-A VNTR, TPH2 rs1386494, 5-HTR1A -1019C-G, 5-HTR2A 102T-C, CCKR1 246G-A, CCKR2 -215C-A, DRD1 -94G-A and COMT Val158Met, were selected for genotyping based on previous positive findings from genetic association studies in PD. After CCK-4 challenge, 39 (35.5%) subjects experienced a panic attack, while 71 subjects were defined as non-panickers. We detected significant differences for both genotypic and allelic frequencies of 1386494A/G polymorphism in TPH2 gene between panic and non-panic groups with the frequencies of G/G genotype and G allele significantly higher in panickers. None of the other candidate loci were significantly associated with CCK-4-induced panic attacks in healthy subjects. In line with our previous association study in patients with PD, we detected a possible association between TPH2 rs1386494 polymorphism and susceptibility to panic attacks. Other polymorphisms previously associated with PD were unrelated to CCK-4-induced panic attacks, probably due to the differences between complex nature of PD and laboratory panic model.

Tryptophan depletion does not modify response to CCK-4 challenge in patients with panic disorder after treatment with citalopram.

RATIONALE: Data by [Bell et al. J Psychopharmacol (2002) 16:5-14] suggest that a decrease in 5-HT neurotransmission predisposes to panic attacks and that the antipanic effect of SSRIs depends upon the availability of 5-HT in the brain. OBJECTIVES: Our aim was to assess the effect of acute tryptophan depletion (TD) on cholecystokinin-tetrapeptide (CCK-4)- induced symptoms in patients with panic disorder (PD) who had responded to a 10-week treatment with a selective serotonin (5-HT) reuptake inhibitor (SSRI), citalopram. MATERIALS AND METHODS: A total of 18 patients (6 males and 12 females, mean age 34.5 years) received a tryptophan-free amino acid drink and a control drink, each followed by a CCK-4 challenge (25 microg), 1 week apart in a double-blind crossover design. RESULTS: The results showed no significant differences in response to the CCK-4 challenge between the TD and the control conditions. Panic rate after the CCK-4 challenge was 27.8% after depletion and 33.3% after control drink (chi2=0.13, p=0.72). No significant effects of TD were observed in panic intensity scores, subjective anxiety, or cardiovascular indices. CONCLUSIONS: This study demonstrates that an acute lowering of brain 5-HT availability with TD does not affect response to a CCK-4 challenge in PD patients successfully treated with citalopram. Thus, the reduction of CCK-4 sensitivity following SSRI-treatment in patients with PD may be related to mechanisms other than 5-HT availability in the brain, possibly to a reduction in brain cholecystokinin receptor sensitivity.

Sensitivity to CCK-4 in women with and without premenstrual dysphoric disorder (PMDD) during their follicular and luteal phases.

The authors determined whether women with premenstrual dysphoric disorder (PMDD) exhibit a heightened sensitivity to the panicogenic effects of CCK-4 administration and whether this enhanced sensitivity to CCK-4 would vary with the phase of the menstrual cycle at the time of CCK-4 injection. Twenty-one normal controls and 18 PMDD women were randomly assigned to receive the first and second CCK-4 injection during the follicular phase and the luteal phase or vice versa. PMDD women showed a greater anxiety and panic response to CCK-4. These preliminary results suggest that the CCK-B system may play a role in the pathophysiology of PMDD.

Influence of clonidine on psychopathological, endocrine and respiratory effects of cholecystokinin tetrapeptide in patients with panic disorder.

The influence of clonidine pretreatment on psychopathological, endocrine and respiratory effects of cholecystokinin tetrapeptide (CCK-4) was characterized. Patients with panic disorder (DSM-III-R) were given 50 micrograms CCK-4 i.v. at 1100 hours on 2 separate study days. In a randomized double-blind design they were additionally infused with 150 micrograms clonidine or placebo from 1040 to 1110 hours. After CCK-4 all patients experienced symptom attacks. No effects of clonidine on panic psychopathology or blood gas parameters were observed. After CCK-4, in the clonidine condition the pituitary release of adrenocorticotropin (ACTH) and prolactin was seemingly enhanced compared to placebo. Our results suggest that CCK-4-induced panic attacks are not suppressible by presynaptic alpha-2 receptor stimulation. Moreover, they point to a synergistic postsynaptic action of clonidine to CCK-4 upon pituitary hormone secretion. The diverging sites of action might possibly explain the discrepancies of psychopathological alterations and stress hormone secretion.

Sensitivity to cholecystokinin-tetrapeptide in major depression.

BACKGROUND: Sensitivity to the panicogenic effects of cholecystokinin-tetrapeptide (CCK-4) is enhanced in panic disorder patients relative to normal controls (NC). In the present study, we determined whether sensitivity to CCK-4 is enhanced in patients with major depressive disorder (MDD) with no history of panic attacks. We also determined whether CCK-4 would exacerbate depressive symptoms. METHODS: The study used a double-blind, randomized, placebo-controlled design. Behavioral and cardiovascular response to a submaximal dose (20 microg) of CCK-4 was studied in seven patients with MDD and 12 NC subjects. RESULTS: None of the subjects panicked with placebo, whereas 29% of MDD and 17% of NC subjects panicked with CCK-4. There was no significant difference between groups on the frequency of CCK-4-induced panic or the number and intensity of panic symptoms. No significant difference was detected for cardiovascular response to the CCK-4 challenge. CCK-4 did not worsen depressive symptoms in MDD patients. LIMITATIONS: Small number of study subjects. CONCLUSIONS: These data indicate that MDD patients show a response to CCK-4 that is comparable to NC. The lack of effect of CCK-4 on depressive symptoms suggest that central CCK receptors may not play an important role in the pathophysiology of MDD.

The effect of 5-hydroxytryptophan on cholecystokinin-4-induced panic attacks in healthy volunteers.

Previous studies suggest a modulatory role of serotonin (5-HT) in experimentally-induced panic attacks. In the current study, we investigated the acute effects of 5-HT precursor l-5-hydroxytryptophan (5-HTP) on the response to panicogenic challenge with cholecystokinin-tetrapeptide (CCK-4) in healthy volunteers. Thirty-two subjects were randomized to receive either 200 mg of 5-HTP or placebo with the CCK-4 challenge following in 90 min in a double-blind, parallel-group design. The results showed a nonsignificant difference between the groups in panic rate (19% after 5-HTP and 44% after placebo, p = 0.13) with a trend for lower intensity of symptoms after 5-HTP (p = 0.08). Further analysis by gender revealed that females in the 5-HTP group had a significantly lower panic rate and intensity of cognitive symptoms whereas, in males, the effect of 5-HTP was limited to lowering the intensity of somatic panic symptoms. Thus, an increased availability of 5-HT may have a gender-dependent protective effect in CCK-4-induced panic.

The influence of Type A behavior pattern on the response to the panicogenic agent CCK-4.

OBJECTIVES: Review of the literature equivocally suggests that subjects with Type A behavioral pattern (TABP) compared to subjects with Type B behavioral pattern display an increased sympathetic activity, a condition associated with sudden cardiac death. The objective of this study was to determine whether healthy subjects classified as Type A or Type B differed in their reactivity to the beta 1 and beta 2 receptor agonist isoproterenol and to the panicogenic agent cholecystokinin-tetrapeptide (CCK-4). By comparing reactivity to CCK-4 after pretreatment with placebo or propranolol, a beta 1 and beta 2 receptor antagonist, the role of the beta adrenergic system in the hypothesized increased response of Type A subjects to CCK-4 was also assessed. METHODS: The study used a randomized, double-blind, placebo-controlled design. Twenty-seven Type A or B subjects were included in the study. The reactivity to isoproterenol was assessed with the CD25 of isoproterenol (i.e., the intravenous dose of isoproterenol necessary to increase the heart rate of 25 bpm). The panic symptom response and the cardiovascular response to bolus injection of 50 microg of CCK-4 was assessed in subjects pretreated with either propranolol or placebo infusions prior to the CCK-4 challenge. An additional group of subjects was recruited and these subjects received a placebo infusion pretreatment before an injection of placebo. RESULTS: The CD25 was significantly greater in Type A subjects than in Type B subjects. No difference was found among the groups on behavioral sensitivity to the CCK-4 challenge. However, CCK-4-induced maximum increase in heart rate was greater in Type A subjects. CONCLUSION: Our finding that Type A subjects exhibited greater CD25 of isoproterenol and greater increases in heart rate following CCK-4 administration compared to Type B subjects suggests that peripheral beta-receptor sensitivity may be increased in individuals with TABP.

Trait dissociation affects the behavioral response to cholecystokinin tetrapeptide in healthy man.

Trait dissociation might influence the response to panicogens in normal controls. The behavioral effects of 25 microg of cholecystokinin tetrapeptide (CCK-4) were studied in 18 healthy men, nine each with high or low trait dissociation. Subjects with high trait dissociation showed a significantly lower increase of acute dissociative, anxiety and panic symptoms compared with subjects with low trait dissociation. Trait dissociation should be assessed in further behavioral challenge studies as a potentially important covariate.

Emotional and cognitive factors connected with response to cholecystokinin tetrapeptide in healthy volunteers.

This article examines the effect of baseline anxiety, anxiety sensitivity and dysfunctional attitudes on the response to cholecystokinin tetrapeptide (CCK-4) in healthy volunteers. CCK-4 and placebo were administered to 14 subjects in a double-blind manner. Four volunteers experienced a panic attack after CCK-4 administration. Those subjects who panicked had significantly higher baseline scores on dysfunctional attitudes. Dysfunctional thought patterns appeared also to predict number of symptoms and experience of cognitive and affective symptoms during injection. Baseline anxiety as well as anxiety sensitivity predicted reactions to placebo but not panic responses to CCK-4. Results suggest that a general tendency towards erroneous interpretation of information has some role in mediating the panicogenic effects of CCK-4, and also interpersonal sensitivity may constitute a vulnerability factor for panic. Psychological factors that have been considered more specific to panic disorder, namely high state and trait anxiety as well as anxiety sensitivity, appeared mainly to determine general reactions to a threatening situation.

Ames Salmonella/mammalian-microsome testing of peptides and peptide synthesis reagents.

The Ames Salmonella/mammalian-microsome assay was used to evaluate the bacterial mutagenicity of 6 bioactive peptides and of 11 chemical reagents used in peptide synthesis. Samples of 2 reagents, bis(2-oxo-3-oxazolidinyl)phosphinic chloride and fluoren-9-ylmethyl chloroformate, showed mutagenic activity with strains TA100 and TA1535, and with TA1537, respectively. No mutagenic activity was found with the bioactive peptides or with the other 9 peptide synthesis reagents.

One milligram of lorazepam does not decrease anxiety induced by CCK-4 in healthy volunteers: investigation of neural correlates with BOLD MRI.

Benzodiazepine effects on cholecystokinin tetrapeptide (CCK-4)-induced panic attack (PA) in humans are incompletely characterized, in particular on the neurofunctional level. This work explores the effects of lorazepam on brain activity and behavioral and physiological symptoms related to CCK-4-induced PA in healthy volunteers. Twenty-one male volunteers received 1 mg of lorazepam or placebo orally, 2 hours before an injection of 0.9% saline solution followed by 50 µg of CCK-4 during functional magnetic resonance imaging (fMRI) and heart rate recording. Panic attacks were defined using the panic symptom scale (PSS). In addition, the Y1-STAI (state anxiety) and the Bond & Lader Visual Analogue Scale (VAS) were used. Eleven subjects were classified as panickers. CCK-4 induced behavioral anxiety and cardiovascular effects along with cerebral activation in anxiety-related brain regions. Overall, lorazepam did not significantly modify the anxiogenic and cardiovascular effects of CCK-4. Regarding CCK-4-induced brain activation, lorazepam did not reduce activity in the insulae and cingulate gyrus of panickers. One milligram of lorazepam was not sufficient to reverse strong panicogenic effects, but decreased brain activity in the case of mild anxiety.

CCK-4: Psychophysiological conditioning elicits features of spontaneous panic attacks.

INTRODUCTION: Cholecystokinin-tetrapeptide (CCK-4) is an established model to generate subjective panic anxiety. CCK-4 injection also results in consistent and dose-dependent rise of stress hormones. Effects other than upon subjective panic and stress hormone activity have barely been examined. The purpose of the study was to investigate CCK-4 effects on emotional facial expression and especially on fear relevant facial muscles establishing therewith a more objective method to measure subjective panic anxiety. METHODS: 20 healthy male subjects were randomly and double-blindedly assigned in two groups (dose groups), each of which was investigated three times once with placebo and twice with 25 µg or 50 µg CCK-4 respectively. Subjects of each group were randomly assigned in two different balanced orders of investigations: CCK-CCK-Placebo vs. Placebo-CCK-CCK. Facial muscle and hypothalamo-pituitary-adrenocortical (HPA)-axis activity were recorded. RESULTS: CCK-4 led dose-dependently to an increase of panic anxiety, an activation of fear relevant facial muscles and a rise of stress hormones. Whereas placebo administration before CCK-4 revealed no significant panic and stress response, during placebo following CCK-4 stimulations a psychophysiological conditioning effect could be observed without rise in HPA-axis activity. DISCUSSION: Our findings indicate the possibility to measure different intensities of panic anxiety and conditioning effects with a facial EMG method. Dissociation of HPA-activity and fear relevant facial muscle activity is in accordance with former results about spontaneous panic attacks.

Independent component analysis applied to pharmacological magnetic resonance imaging (phMRI): new insights into the functional networks underlying panic attacks as induced by CCK-4.

Pharmacological magnetic resonance imaging (phMRI) is a method to study effects of psychopharmacological agents on neural activation. Changes of the blood oxygen level dependent (BOLD), the basis of functional MRI (fMRI), are typically obtained at relatively high sampling frequencies. This has more recently been exploited in the field of fMRI by applying independent component analysis (ICA), an explorative data analysis method decomposing activation into distinct neural networks. While already successfully used to investigate resting network and task-induced activity, its use in phMRI is new. Further extension of this method to tensorial probabilistic ICA (tensor PICA) allows to group similar brain activation across the anatomical, temporal, subject or session domain. This approach is useful for pharmacological experiments when no pharmacokinetic model exists. We exemplify this method using data from a placebo-controlled cholecystokinine-4 (CCK-4) injection experiment performed on 16 neuropsychiatrically and medically healthy males (age 25.6 +/- 4.2 years). Tensor PICA identified strong increases in activity in 12 networks. Comparison with results gained from the standard approach (voxelwise regression analysis) revealed good reproduction of areas previously associated with CCK-4 action, such as the anterior cingulate, orbitofrontal cortex, cerebellum, temporolateral, left parietal and insular areas, striatum, and precuneus. Several other components such as the dorsal anterior cingulate and medial prefrontal cortex were identified, suggesting higher sensitivity of the method. Exploration of the time courses of each activated network revealed differences, that might be lost when a fixed time course is modeled, e. g. neuronal responses to an acoustic warning signal prior to injection. Comparison of placebo and CCK-4 runs further showed that a proportion of networks are newly elicited by CCK-4 whereas other components are significantly active in the placebo conditions but further enhanced by CCK-4. In conclusion, group ICA is a promising tool for phMRI studies that allows quantifying and visualizing the modulation of neural networks by pharmacological interventions.

Effects of tiagabine on cholecystokinin-tetrapeptide (CCK-4)-induced anxiety in healthy volunteers.

There is increasing evidence that a dysregulation of the gamma-aminobutyric acid (GABA) system plays a role in the pathophysiology of panic disorder. Selective enhancement of GABAergic neurotransmission has been shown to improve anxiety in experimental animals and in patients with panic disorder. Tiagabine is an antiepileptic drug, which increases GABA via selective blockade of GABA reuptake. Apart from its anticonvulsant activity anxiolytic properties could therefore be suggested. To investigate the putative anxiolytic properties of the GABA reuptake blocker tiagabine, we studied the impact of tiagabine treatment on cholecystokinin tetrapeptide (CCK-4)-induced panic. Fifteen healthy volunteers received 15 mg tiagabine daily for 1 week. A CCK-4 challenge was performed before and after treatment. Panic was assessed using the API- and PSS-score. There was a marked improvement of CCK-4-induced panic after 1 week of treatment. Both API- and PSS-scores showed a significant reduction. Our results suggest anxiolytic properties of tiagabine in humans, which provide sufficient rationale to assess its putative anxiolytic effects in patients with panic disorder under controlled conditions.