Roland Lawson

New therapeutic opportunities for 5-HT2 receptor ligands.

ABSTRACT Serotonergic dysfunction is mainly associated with neuropsychiatric and cardiovascular disorders but has also been linked with many other pathological conditions. Serotonin (5‐hydroxytryptamine, 5‐HT) mediates numerous physiological functions in the brain and the periphery by activating a variety of receptors. 5‐HT receptors are divided into four classes, three of which belong to the G protein‐coupled receptor family. This review provides an overview of the recent pharmacological developments involving the Gq‐coupled 5‐HT2 receptor subfamily as well as the pathological implications of this receptor subfamily with regard to fibrosis, the central nervous system, cardiovascular disorders, and cancer. The final section highlights new therapeutic opportunities and emerging research revealing unexplored medical opportunities for this class of 5‐HT receptors. The development of biased 5‐HT2 receptor ligands appears to be an interesting topic in various areas. In light of recent discoveries, the need for the development of new and safer drugs should take into account the risk of cardiovascular side effects such as pulmonary hypertension and heart valve disease.

Serotonin 2B receptor slows disease progression and prevents degeneration of spinal cord mononuclear phagocytes in amyotrophic lateral sclerosis

Microglia are the resident mononuclear phagocytes of the central nervous system and have been implicated in the pathogenesis of neurodegenerative diseases such as amyotrophic lateral sclerosis (ALS). During neurodegeneration, microglial activation is accompanied by infiltration of circulating monocytes, leading to production of multiple inflammatory mediators in the spinal cord. Degenerative alterations in mononuclear phagocytes are commonly observed during neurodegenerative diseases, yet little is known concerning the mechanisms leading to their degeneration, or the consequences on disease progression. Here we observed that the serotonin 2B receptor (5-HT2B), a serotonin receptor expressed in microglia, is upregulated in the spinal cord of three different transgenic mouse models of ALS. In mutant SOD1 mice, this upregulation was restricted to cells positive for CD11b, a marker of mononuclear phagocytes. Ablation of 5-HT2B receptor in transgenic ALS mice expressing mutant SOD1 resulted in increased degeneration of mononuclear phagocytes, as evidenced by fragmentation of Iba1-positive cellular processes. This was accompanied by decreased expression of key neuroinflammatory genes but also loss of expression of homeostatic microglial genes. Importantly, the dramatic effect of 5-HT2B receptor ablation on mononuclear phagocytes was associated with acceleration of disease progression. To determine the translational relevance of these results, we studied polymorphisms in the human HTR2B gene, which encodes the 5-HT2B receptor, in a large cohort of ALS patients. In this cohort, the C allele of SNP rs10199752 in HTR2B was associated with longer survival. Moreover, patients carrying one copy of the C allele of SNP rs10199752 showed increased 5-HT2B mRNA in spinal cord and displayed less pronounced degeneration of Iba1 positive cells than patients carrying two copies of the more common A allele. Thus, the 5-HT2B receptor limits degeneration of spinal cord mononuclear phagocytes, most likely microglia, and slows disease progression in ALS. Targeting this receptor might be therapeutically useful.

Old spontaneously hypertensive rats gather together typical features of human chronic left-ventricular dysfunction with preserved ejection fraction

Objective: Heart failure with preserved left-ventricular ejection fraction (HF-PEF) is an entity leading to pulmonary congestion because of impaired diastolic filling. This syndrome usually strikes those who have experienced a long history of hypertension or metabolic risk factors. Pathophysiological mechanisms are not fully understood, and standard therapy is not established. Relevant preclinical models are still lacking. The aim of this work was to evaluate aging spontaneously hypertensive rats (SHRs) as a model of HF-PEF. Methods: Serial echocardiographic and blood pressure (BP) measurements were performed in 28, 36, 43, 47 and 51-week-old SHRs and their normotensive controls (Wistar–Kyoto rats). In 52–53-week-old animals, final investigations included ECG, invasive left-ventricular (LV) and aortic catheterization, brain natriuretic peptide (BNP) plasma concentrations, ventricular reverse transcription-qPCR evaluations (&bgr;-myosin heavy chain, atrial natriuretic peptide, BNP, sarco/endoplasmic reticulum calcium ATPase 2a and collagens 1a, 3a and 2a) and cardiac histology. Results: SHRs develop a progressive alteration of the early diastole, some of the echocardiographic parameters being not sensitive to BP reduction by the calcium blocker, nicardipine. The systolic function evaluated by echocardiography and invasive catheterization was preserved. When the observation period was over, an increase in collagen synthesis and deposits were identified in subendocardial layers. This attested a probable myocardial ischemia that was confirmed by ECG changes of the ST segment. BNP increased in the blood and at the mRNA level in the myocardium. Conclusion: When aging, SHRs progressively develop HF-PEF showed by impaired LV relaxation and hypertrophy, BNP increase but preserved contractility and fibrosis. This model seems pertinent for further pharmacological preclinical studies in the field.

Contribution of serotonin to cardiac remodeling associated with hypertensive diastolic ventricular dysfunction in rats.

Objective: Left-ventricular hypertrophy and interstitial fibrosis are the main pathophysiological factors of heart failure with preserved ejection fraction. Blockade of the serotonin 5-HT2B receptor (5-HT2BR) has been shown to reduce cardiac hypertrophy, oxidative stress, and extracellular cell matrix activation. In this study, we evaluated the effects of the 5-HT2BR blockade, on hemodynamic and cardiac remodeling, in spontaneously hypertensive rats (SHRs) that display a diastolic dysfunction with preserved ejection fraction. Method: Thirty-seven-week-old SHRs were randomized in four groups receiving either saline, the selective 5-HT2BR antagonist RS-127445 (1 mg/kg per day), a calcium channel blocker nicardipine (6 mg/kg per day), or RS-127445 + nicardipine. During the 14 weeks of treatment period, cardiac function and blood pressure were monitored by echocardiography and tail-cuff. Finally, electrocardiograms and invasive hemodynamics were obtained before blood collection. Heart was analyzed for morphology and mRNA expression. A complementary study evaluated the cardiac and vascular effects of serotonin on wild-type and mice knockout for the 5-HT2BR (Htr2B−/−) and/or the 5-HT2AR (Htr2A−/−). Results: Despite the left ventricular 5-HT2BR overexpression, 5-HT2BR blockade by RS-127445 did not affect left ventricular hypertrophy and fibrosis in SHRs. This antagonist did not improve diastolic dysfunction, neither alone nor in combination with nicardipine, although it induced plasma brain natriuretic peptide decrease. Moreover, RS-127445 amplified subendocardial fibrosis and favored left ventricular dilatation. Finally, a subendocardial left ventricular fibrosis was induced by chronic serotonin in wild-type mice, which was increased in Htr2B−/− animals, but prevented in Htr2A−/− and Htr2A/2B−/− mice, and could be explained by a contribution of the endothelial 5-HT2BRs to coronary vasodilatation. Conclusion: This work is the first to identify a cardioprotective function of the 5-HT2BR in an integrated model of diastolic dysfunction with preserved ejection fraction.

Mitral bioprosthesis hypertrophic scaring and native aortic valve fibrosis during benfluorex therapy

The authors describe the case of a simultaneous mitral bioprosthesis hypertrophic scaring and native aortic valve fibrosis during benfluorex therapy in a 40‐year‐old woman. Four years before, she underwent a mitral valve replacement after the diagnosis of mitral regurgitation during benfluorex treatment (150 mg/day). This drug was reintroduced postoperatively. She presented with exercise and sometimes resting dyspnoea. The bioprosthesis and aortic valves exhibited similar histopathological lesions. Thickening and plaque deposits made by smooth muscle alpha actin‐ and vimentin‐positive cells in a glycosaminoglycan matrix were observed. The study discusses the putative contribution of circulating progenitor cells activated by 5‐HT2B receptor agonists in the development of drug‐induced heart disease.

The role of 5‐HT2B receptors in mitral valvulopathy: bone marrow mobilization of endothelial progenitors

Valvular heart disease (VHD) is highly prevalent in industrialized countries. Chronic use of anorexigens, amphetamine or ergot derivatives targeting the 5‐HT system is associated with VHD. Here, we investigated the contribution of 5‐HT receptors in a model of valve degeneration induced by nordexfenfluramine, the main metabolite of the anorexigens, dexfenfluramine and benfluorex.

Serotonin contribution to cardiac valve degeneration: new insights for novel therapies?

Heart valve disease (HVD) is a complex entity made by different pathological processes that ultimately lead to the abnormal structure and disorganization of extracellular matrix proteins resulting to dysfunction of the leaflets. At its final evolutionary step, treatments are limited to the percutaneous or surgical valve replacement, whatever the original cause of the degeneration. Understanding early molecular mechanisms that regulate valve interstitial cells remodeling and disease progression is challenging and could pave the way for future drugs aiming to prevent and/or reverse the process. Some valve degenerative processes such as the carcinoid heart disease, drug-induced valvulopathy and degenerative mitral valve disease in small-breed dogs are clearly linked to serotonin. The carcinoid heart is typically characterized by a right-sided valve dysfunction, observed in patients with carcinoid tumors developed from serotonin-producing gut enterochromaffin cells. Fenfluramine or ergot derivatives were linked to mitral and aortic valve dysfunction and share in common the pharmacological property of being 5-HT2B receptor agonists. Finally, some small-breed dogs, such as the Cavalier King Charles Spaniel are highly prone to degenerative mitral valve disease with a prevalence of 40% at 4 years-old, 70% at 7 years-old and 100% in 10-year-old animals. This degeneration has been linked to high serum serotonin, 5-HT2B receptor overexpression and SERT downregulation. Through the comprehension of serotonergic mechanisms involved into these specific situations, new therapeutic approaches could be extended to HVD in general. More recently, a serotonin dependent/ receptor independent mechanism has been suggested in congenital mitral valve prolapse through the filamin-A serotonylation. This review summarizes clinical and molecular mechanisms linking the serotonergic system and heart valve disease, opening the way for future pharmacological research in the field.

0171 : Contribution of progenitor cells sharing some serotonergic proteins in the pathophysiology of human aortic and mitral valves degeneration

Many compounds (pergolide, fenfluramine, ectasy) were described as inducers of fibrotic valvular lesions. All these compounds share in common to activate the 5HT2B serotonergic receptor. This observation leads to the hypothesis that cardiac valves express a « serotonergic system » that could be activated by serotonin (5-HT) or 5-HT receptor (5-HTR) agonists. In this work, we characterized the expression pattern of 5-HT2A, 2B, 4 R and the 5- HT transporter (SERT) in whole and cell subpopulations of 30 human mitral and aortic valves collected at the time of surgical valve replacement (Aortic: 11 calcified, 5 sclerotic, 4 bicuspid; Mitral: 12 dystrophic). All samples express 5HT2A, 2B, 4 R and SERT, the amount of 5HT2B R mRNA being higher than the 5HT2A R whatever the valve and etiology. 5HT2BR expression is found in endothelial cells (CD31+) at the valve surface, but also inside valve lesions, expressed by interstitial cells (smooth muscle α-actin and vimentin positive cells) located in an abundant glycosaminoglycan matrix. In fact, fibromyxoid lesions and calcified aortic valves express a high amount of CD34+ cells. These cells are endothelial progenitors because they express VEGFR2 and eNOS together with 5-HT2R. After collagenase treatment, valve samples were labeled with CD31 and CD34 antibodies: 60.1 (± 11) % of all mitral valvular cells are CD34+ compared to 36.1 (±8) % of aortic valvular cells (FACS analysis and sorting). To summarize, 5HT2A, 2B, 4 receptors and SERT are expressed in aortic and mitral diseased valves. The amounts of 5HT2A, 2B R mRNA are equivalent between mitral and aortic valves. High amount of CD34+ endothelial progenitors, expressing 5HT2AR and 5HT2BR, are found in degenerated valves. The contribution of the two 5-HT2 receptors and endothelial progenitors in valve degeneration is now under investigation.

0122: The serotonergic system in pathological human cardiac valves. What is the role of progenitors cells expressing the 5-HT2B receptor?

Many compounds (pergolide, cabergoline, fenfluramine, ectasy) were described as inducers of fibrotic valvular lesions, a rare but severe drug reaction. All these drugs share in common the pharmacological property to activate a serotonergic receptor subtype, the 5HT2B. Together with the well known “carcinoid heart” that is a valvulopathy due to high amounts of circulating serotonin, these observations lead to the hypothesis that cardiac valves express a “serotonergic system” that could be activated by 5-HT or 5-HTR agonists. The aim of this work was to characterize the pattern of expression of 5-HT2A,2B,4 receptors, the serotonin transporter (SERT) and the biosynthesis peripheral enzyme (Tph1) in various valvulopathies. Thirty degenerated human valves were collected: 11 calcified aortic valves (CAV), 5 sclerotic aortic valves (SAV), 11 dystrophic mitral valves (DMV). They were analyzed by RT-qPCR and immunohistochemistery. All samples express 5HT2A,2B,4 receptors, SERT and Tph1. In these valve tissues, the amount of 5HT2B receptor (5HT2B R) mRNA is higher than the 5HT2A one (5HT2A R) : Δ Ct (5HT2B R -18S) = 12,53±1,12 vs Δ Ct (5HT2A R -18S) = 15,95±2,37 for CAV, Δ Ct (5HT2B R -18S) = 13,04±2,62 vs Δ Ct (5HT2A R - 18S)=16,00±1,46 for SAV, Δ Ct (5HT2B R -18S) = 12,34±0,77 vs Δ Ct (5HT2A R -18S) = 16,14±0,86 for DMV. The amounts of SERT, Tph1 and 5HT4 receptor mRNA are negligible whatever valve and etiology. At a topographical point of view, 5HT2BR expression is found in endothelial cells (at the valve surface) but also inside valve lesions, by interstitial cells (smooth muscle α-actin and vimentin positive cells) located in an abundant glycosaminoglycan matrix. Characterization of these cells is in progress. In particular, we characterize the high amount CD34+ hematopoietic progenitors that are highly present in fibromyxoid lesions. To summarize, 5HT2A,2B,4 receptors, SERT and Tph1 are expressed in aortic and mitral diseased valves. The amounts of 5HT2A,2B R mRNA are equal between mitral and aortic valves. The contribution of the two 5-HT2 receptors in valve degeneration is now under investigation whatever the pathological process considered.