Large Animal Model for Evaluating the Efficacy of the Gene Therapy in Ischemic Heart

Summary

Myocardial gene therapy for ischemic heart disease holds great promise for future therapeutics. Here, we introduce a large animal model for evaluating the efficacy of gene therapy in the ischemic heart.

Abstract

Coronary artery disease is one of the significant causes of mortality and morbidity worldwide. Despite the progression of current therapeutics, a considerable proportion of coronary artery disease patients remain symptomatic. Gene therapy-mediated therapeutic angiogenesis offers a novel therapeutic method for improving myocardial perfusion and relieving symptoms. Gene therapy with different angiogenic factors has been studied in few clinical trials. Due to the novelty of the method, the progress of myocardial gene therapy is a continuous path from bench to bedside. Therefore, large animal models are needed for evaluating the safety and efficacy. The more the large animal model identifies the original disease and the endpoints used in clinics, the more predictable outcomes are from clinical trials. Here, we introduce a large animal model for evaluating the efficacy of the gene therapy in the ischemic porcine heart. We use clinically relevant imaging methods such as ultrasound imaging and ¹⁵H₂O-PET. For targeting the gene transfers into the desired area, electroanatomical mapping is used. The aim of this method is: (1) to mimic chronic coronary artery disease, (2) to induce therapeutic angiogenesis at hypoxic areas of the heart, and (3) to evaluate the safety and efficacy of the gene therapy by using relevant endpoints.

Introduction

Coronary artery disease is accountable for the vast proportion of mortality and disease burden worldwide¹. Current treatment strategies are percutaneous interventions, pharmacological treatment, and bypass surgery². However, despite the progression of these current therapeutics, many patients suffer from so-called refractory angina, underlining the unmet need for novel treatment approaches³. Gene therapy-mediated therapeutic angiogenesis could target this patient group.

Myocardial gene therapy is most often delivered by using different viral vectors, most commonly repli

Protocol

The experiments presented here are performed using about 10-week-old female domestic pigs and are approved by the Animal Experiment Board in Finland. Animals weigh 30-40 kg at the beginning of the protocol, allowing the same procedural equipment and imaging modalities as possible for humans. Chronic ischemia is induced 14 days before the gene transfer, and the follow-up time after the gene transfer depends on the viral vector used. The study protocol is shown in Figure 1. This protocol can be used to perform adenoviral or AAV-mediated gene therapy injections. The time of sample collection has to be adjusted to the transgene expression pea

Results

The success of the ischemia operation can be confirmed with this protocol by coronary angiogram and by determining the hypokinetic area by transthoracic ultrasound (Figure 1) before proceeding to the gene delivery. The state of the coronary occlusion can be evaluated by coronary angiogram, and the electroanatomical mapping ensures the ischemic and hibernating areas.

The efficacy of the gene therapy can be analyzed by measuring the circumferential strain, ejection ...

Discussion

The timepoints of this protocol may be modified according to the viral vector used. Also, the immunohistological analyses may be selected according to the therapeutic gene. It is also possible to add more timepoints and endpoints to the protocol if needed.

This protocol comprises stages, which are essential to succeed and impossible to correct afterward. First, if one fails to induce appropriate ischemia, the animal must be excluded from further procedures and analyses. Standardization of the ...

Materials

Name	Company	Catalog Number	Comments
1% PFA	VWR	VWRC28794.295	Prepared from paraformaldehyde powder
15 % sucrose	VWR	VWRC27480.294	Prepared from solid sucrose
4% PFA	VWR	VWRC28794.295	Prepared from paraformaldehyde powder
5 F pigtail catheter	Cordis	534-550S
6 F catheter AR2	Cordis	670-112-00
6 F introducer sheath	Cordis	504-606X
8 F introducer sheath	Cordis	504-608X
Acetylsalicylic acid			Varying producer
Amiodarone			Varying producer
Angiographic station	GE Healthcare
Angiolaboratory set	Mölnlycke		designed for the needs of our angiolaboratory, contains sterile drapes, cups and swabs
Bisoprolol			Varying producer
Cefuroxime			Varying producer
Clopidogrel			Varying producer
Coroflex Blue stent	B.Braun Medical	5029012	Catalog number depends on stent size
Crile forceps
Cyclotron	GE Healthcare
Dobutamine			Varying producer
Electroanatomical mapping system	Biologics Delivery Systems, Johnson & Johnson company
Enoxaparin			Varying producer
Fentanyl			Varying producer
Intramyocardial injection catheter	Johnson & Johnson
Iodine contrast agent	Iomeron
Kitchen knife			Varying producer
Lidocaine			Varying producer
Liquid nitrogen			Varying producer
MgSO4			Varying producer
Needle 18 G	Cordis	12-004943
Perfusion pump
PET-CT scanner	Siemens Healthcare
Polytetrafluoroethylene tube
Propofol			Varying producer
Scalpel no 11	VWR	SWAN0503
Sublingual dinitrate	Takeda
Ultrasound machine	Philips