Research

Normalization of sinus node pacemaker activity by GIRK4 (IKACh) inhibition. Partners will test whether pharmacologic inhibition of GIRK4 normalizes in vivo heart rate in a mouse model of HCN4-mediated SND secondary to HF and in coronary-perfused human sinus node preparations.

Characterization of miRs in the sinus node human hearts. Collaborative work has profiled miRs of the healthy human sinus node versus its surrounding atrial muscle. Certain miRs are upregulated in the human sinus node from hearts with history of HF. During year 1, further analysis of the normal human sinus node to identify miRs differentially expressed in subregions of the sinus node (central, caudal, transitional and paranodal).

Validation of the PLiPS-CM as a model of native sinus node myocytes. During the first year, partner laboratories have combined their experience in stem cells with expertise of PJM lab to explore the use of hiPS-CM as cellular models of human sinus node cells.

Establishment of genetic analysis protocol. Collaborative efforts have: (i) set-up multi-gene panel analysis and bioinformatic pipeline for idiopathic (primary) SND and (ii) establish criteria for recruitment of patients that will be used during the funding period.

• Aim 1: To normalize heart rate in HCN4-mediated SND by pharmacologic blockade of GIRK4.

  • Background and objective: We aim to test the effectiveness of pharmacologic inhibition of GIRK4 channels (IKACh) to normalize heart rate and rhythm in vivo and ex vivo. We will test inhibition of IKACh in models of secondary HCN4-mediated SND developed by partners MB, HD and IC. To extend testing in a translational setting, partners MM and PJM will work together with young fellows on isolated coronary-perfused human sinus nodes from hearts available in PJM’s team.

• Aim 2: To validate target miRs in HCN4-mediated SND mechanisms.

  • Background and objective: We have identified miRs significantly affecting SND. In addition, in vivo knockdown in mice of miRs targeting HCN4 channels prevents secondary HCN4-mediated SND due to training and HF respectively; raising the possibility that miR antagonism may be an effective therapy. We will study expression of miRs in animal secondary SND models and in the human sinus nodes with history of SND. Expression of miRs will be compared to that in control animals or in human sinus nodes with history of normal sinus rhythm, respectively. The objective is to identify new target miRs involved in ion channel regulation in a translational setting to prepare an anti-miR based SND reversal strategy (Aims 3 and 4).

• Aim 3: Model the mechanisms of onset and rescue of HCN4-mediated SND in vivo.

  • Background and objective: Partners have shown previously that it is possible to identify miRs involved in the regulation of HCN4 expression in sinus bradycardia induced by athletic training and HF. Importantly, it has been shown that by antagonizing specific miRs it is possible to improve HCN4 expression and reverse SND. We thus aim to extend the concept of SND reversal by manipulating the expression of identified miRs to induce SND in control mice, or repress SND in mouse models of secondary SND associated with training, HF and diabetes.

• Aim 4: Develop vector-based long-term reversal of HCN4-mediated SND.

  • Background and objective: We will pursue long-term reversal of the pathway leading to SND by antagonizing specific miRs in vivo. We plan to design vectors enabling long-term miR antagonism with potential future applicability to human SND. We will culture hiPS-CM as a cellular model to manipulate expression of miRs selected from human sinus nodes. We will use adeno-associated viral vectors (AAV) for long-term miR manipulation.

The overall goal of the FANTASY Leducq Network is to provide proof of concept knowledge to indicate new pharmacologic and molecular therapies to restore normal sinus node function, improve dysfunctional A-V conduction and control arrhythmias associated to bradycardia under a wide spectrum of cardiovascular diseases. At the beginning of the funding period, the teams of the two coordinators have jointly published a major review article discussing the clinical aspects of sinus node dysfunction (SND) and the perspective roles of pharmacologic approaches in managing this disease. In addition, another review article discusses the genetic complexity of primary SND forms, which is another important aim of our network. Generally, results of local and collaborative work over year 2020 and 2021 have largely validated our working hypotheses that selective targeting cardiac G protein activating K+ channels (GIRK, GIRK4) underlying I_KACh can constitute a new strategy to improve heart rate in SND and promote reversal of the disease phenotype.