Genetics of Heart Failure Clinical Stages
Study Details
Study Description
Brief Summary
Heart failure is one of the most severely disabling cardiovascular diseases and is a leading cause of death worldwide; especially in developing countries like Egypt, where it is considered one of the most significant health and economic burdens. Heart failure is a chronic long-term clinical syndrome that gets worse with time. There are 4 stages of heart failure (Stage A, B, C, and D). The stages range from "high risk of developing heart failure" to "advanced heart failure". The pathogenesis of heart failure involves a complex interaction between environmental and genetic factors. Genetic factors may affect the susceptibility to the underlying etiology of heart failure, the rapidity of disease progression, or the response to pharmacologic therapy. Ischemic heart diseases (IHD) and dilated cardiomyopathy (DCM) are the most common structural heart diseases in stage B heart failure that progress to stage C and stage D. Many studies investigated the most important, gene expression alteration associated with DCM and IHD. However, there are no sufficient data regarding gene expression profiles and prognostic biomarkers specific to patients with DCM or IHD showing no symptoms of heart failure cascade (stage B heart failure), Patients with typical heart failure symptoms and underlying IHD and DCM (stage C heart failure) and patients with decompensated heart failure( stage D heart failure). This project aims to use RNA sequencing for gene expression profiling of different clinical stages of heart failure, the discovery of novel transcripts, identification of alternatively spliced genes, detection of alternative splicing, gene regulation, discoveries and detection of allele-specific expression allowing subtle changes in gene functioning to be identified in different stages of heart failure. The main goal of the project is to explore novel down-regulated, up-regulated genes, transcripts and allele-specific expressions that may contribute to heart failure progression from stage II(structural cardiac abnormalities with no significant symptoms) to stage IV (recurrent hospital admission due to heart failure exacerbations).)
| Condition or Disease | Intervention/Treatment | Phase |
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Detailed Description
B.3. Outcomes and Impact:
It's a great honor that sir Magdy yacoub revised this project more than once and it's a great honor to me and all the project 's team that he sir Magdy yaq. Accepted to be the co- investigator o this project, And this truly reflects his patriotism and his concern for everything that is in the interest of the health of Egyptians Finding out genetic biomarkers and signatures that typify ischaemic cardiomyopathy patients and dilated cardiomyopathy patients at the preclinical stage of heart failure, symptomatic clinical stage of heart failure and decompensated end-stage heart failure. These specific biomarkers may be greatly used as diagnostic biomarkers for IHD and DCM patients at extremely high risk for progression to later severely symptomatic stages of heart failure. The early screening of those patients may help to prevent their progression to later stages of heart failure by close monitoring and therapeutic and non-therapeutic strategies. We put into consideration that the traditional pharmacologic agents used to slow down or prevent the progression of typically symptomatic heart failure patients- mainly beta blockers and angiotensin converting enzyme inhibitors- are already prescribed to all IHD patients with no proven efficacy to prevent the progression of those patients to typical heart failure patients (left ventricular dysfunction patients). Based on the above hypothesis, there is a great need for novel therapies that may prevent the progression of structural heart disease patients(stage B heart failure) to typical and end-stage heart failure patients ( stage C and D heart failure).
The expected results may explain novel genetic pathways of the clinical progression of asymptomatic structural cardiac disorder (stage B heart failure) to typically symptomatic heart failure(stage C) and decompensated heart failure(stage D).
The expected results of this project may lead us to the novel foundation of the first Egyptian database of the genetic biomarkers of different heart failure stages with underlying IHD or DCM.
The novelty of the idea and our plan to use the most recent gene expression profiling techniques and the most updated bioinformatics methodologies may pave the way for publishing the results of this work in high impact factor journal both in molecular genetics field and cardiology field.
The expected result of this project may be a reference for many candidates of master degrees and doctorate degrees in the field of heart failure and cardiac molecular genetics in Egypt and around the world. The novelty of the idea, given that- for the most of our knowledge- there are no previous studies that investigated the specific gene expression profiling in different stages of heart failure, may attract high score of citation by specialized scientists in the field.
Early screening of asymptomatic structural heart disease patients at high risk of progressive heart failure stages development may help to prevent their progression by pharmacologic and non-pharmacologic strategies. This may contribute to saving a huge amount of government money spent on the care and the treatment of those patients at severely symptomatic decompensated stages of heart failure.
The plan of the investigators is to further use the information that will be revealed in this project to apply the most recently investigated techniques of RNA activation and repression to treat with gene overexpression and gene dysregulation in patients suffering progressive stages of heart failure VS. those at primary asymptomatic stages of heart failure to pave the way for the innovative novel gene therapy for prevention of progression from primary stages to progressive stages of heart failure and the recovery of end-stage heart failure patients to the stable asymptomatic normal lifestyle stages of the disease. This novel therapy will not only contribute to mitigating the misery of such huge number of Egyptians suffering from this greatly disabling disease but also it will be a scientific revolutionary breakthrough in the field of gene therapy all over the world. We are full of faith that we are the Egyptian can make miracles by patience, determination, fight and belief in our dream.
B.4. State-of-the-Art and Excellence:
Heart failure (HF) affects an estimated 6.5 million adult Americans [1]. Although survival rates have raised by 10% between 1979 and 2000, the current 5-year mortality rate is still about 50% [1, 2]. A long-standing paradigm is that end-stage heart failure with reduced ejection fraction (HFrEF) evolves via a "final common pathway" despite having a wide variety of etiologies and genetic contributions [3, 4]. Clinical trial results and current guidelines for the management of HFrEF confirm this viewpoint and direct therapy based mainly on the degree of left ventricular ejection fraction, and clinical severity using the New York Heart Association (NYHA) classification [5].
Ischaemic heart disease (IHD) is the most common cause of death globally according to WHO reports and is the leading cause of heart failure in the developed countries [6-9]. Heart failure secondary to IHD has been found to be independently associated with higher mortality rates compared with a non-ischemic etiology [10, 11]. The increasing incidence has been attributed to the high efficacy of the thrombolytic and primary percutaneous coronary intervention in the treatment of acute myocardial infarctions, leading to improved patient survival, however often leading to increased morbidity due to left ventricular (LV) remodeling and chronic left ventricular dysfunction. The term ischaemic cardiomyopathy (ICM) has been defined as LV systolic dysfunction with one or more of the following: a history of prior myocardial revascularization or myocardial infarction, more than 75% stenosis in the left main stem or left anterior descending artery, or two vessels or more with a greater than 75% stenosis [12].
One of the most common causes of heart failure is cardiomyopathies. The American Heart Association described cardiomyopathies as a heterogeneous group of myocardial diseases that are predominantly genetic and associated with mechanical and/or electrical dysfunction [13]. DCM is characterized by the presence of a left ventricular or biventricular dilatation and systolic dysfunction in the absence of abnormal loading conditions. It is the second most common cause of heart failure with reduced ejection fraction. DCM is a complex disorder with diverse genetic variants and environmental factors that determine the disease onset and the course of the disease [14, 15].
IHD and cardiomyopathies induce modification of the myocardium structure and may lead to l alterations of the heart function [16] including a reduction in the left ventricular ejection fraction (LVEF). Identification of patients at the "pre-heart failure" stage may prevent the development of HF through the implementation of adapted medical and non-medical strategies. This silent preclinical state (pre-heart failure) is referred to as asymptomatic left ventricular dysfunction (ALVD) and can only be discriminated by transthoracic echocardiography. ALVD, common in the general population, leads to a high risk of developing overt HF. Indeed, ALVD subjects have a 12-fold increase in the annual rate of hospitalization for first-event HF compared to individuals with normal LVEF [17] and a 4-fold increase in the risk of death over a 6-year period [16]. Effective large-scale screening for ALVD, representing a major unmet clinical challenge, requires a determination of ALVD biomarkers.
The screening for ALVD has started for over a decade [17], [18]. However, there are no ALVD biomarkers. Indeed, ALVD diagnosis warrants a complicated echocardiographic analysis, which is time-consuming, costly and not applicable to the large population of individuals at risk. The lack of biomarker(s) may be of high importance because ALVD is highly prevalent due to the general increase in cardiovascular risk factors [19]. ALVD has been established as a predictive early indicator of severe HF [20]. Follow-up studies have shown that patients with ALVD display an average annual chronic heart failure rate of 4.9 to 20%, with a mortality rate of 5.1 to 10.5% [21], [22]. These observations were recently confirmed in a 5-year survival rate analysis that demonstrated a death rate of 31% for subjects with from ALVD and of 47% for patients suffering systolic HF [23]. Finally, the SOLVD study demonstrated that the treatment of ALVD can significantly delay the occurrence of HF [24]. Therefore, it may be of utmost clinical importance to identify ALVD individuals in the general population before they develop overt HF.
There is a paucity of trials that investigated the genetic biomarkers specific for ischaemic heart disease (IHD) patients (with or without previous history of acute myocardial infarction) or DCM patients who sustained a normal left ventricular function (stage B), IHD or DCM patients with typical symptomatic heart failure (stage C) and IHD or DCM patients who progress to decompensated heart failure (stage D). Maciejak et al, [25] investigated the genetic biomarkers of IHD patients with acute myocardial infarction (AMI) who developed left ventricular dysfunction VS. acute myocardial infarction patients who retained normal ventricular ejection fraction at the same time duration. They demonstrated an altered gene expression profile in AMI who developed HF. Sweet et al, [26] analyzed whole transcriptome in ischaemic cardiomyopathy (ICM) patients and DCM patients VS healthy patients with no history of cardiovascular diseases. They found key etiology-specific signatures that typified ICM and DCM. Tijsen et al, [27] performed micro RNA array in healthy subjects, heart failure patients, patients with dyspnea due to heart failure and patients with dyspnea for causes other than heart failure. They identified 6 miRNAs that are elevated in patients with HF, among which miR423-5p was most strongly related to the clinical diagnosis of HF. Smih F et al, [28] investigated, by microarray analysis, specific gene expression profile for asymptomatic left ventricular dysfunction, namely, pre-heart failure patients. None of the above-mentioned trials investigated the gene expression profiles or genetic biomarker signatures specific for progressive symptomatic stages of heart failure VS. pre-clinical heart failure stage.
Prior microarray studies revealed distinct gene expression signatures between HF etiologies [28-33]; but others failed to find distinctions [34, 35]. Direct RNA-sequencing (RNA-seq) also provides accurate quantification of transcripts and has been used to identify expression signatures between HF and non-failing (NF) hearts [36], novel transcriptional regulators and perturbed miRNA networks in ICM or DCM [37, 38], pre- and post-LVAD transcriptomes [39, 40], common HF genes in pediatric cardiomyopathy [41], and splicing, eQTL, and allelic expression in DCM [42].
We hypothesize that the full understanding of the specific genetic variations such as gene overexpression or gene dysregulation in such cohorts of patients with IHD or DCM who progress to later stages of heart failure VS. IHD or DCM patients who retain normal systolic function may pave the way for finding out diagnostic genetic biomarkers of IHD patients and DCM patients with high risk for progression to later stages of heart failure. Moreover, finding out differential gene expression profiles in IHD and DCM subjects at different stages of heart failure may be the first step for the innovation of novel gene therapies for prevention of heart failure progression in IHD patients at stage B and the recovery of IHD patients at progressive stages of heart failure (C and D).
B.7. Approach and Methodology:
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Project management and consortium
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Patients recruitment and allocation The protocol of this study will be extensively designed and reviewed by the institutional review board of Asyut University educational cardiology hospital. All patients will be requested to sign a detailed informed consent before recruitment.
Inclusion criteria: IHD or DCM patients with no HF symptoms (stage B), with typical HF symptoms (stage C) and patients with decompensated HF (stage D).
Exclusion criteria: Patients with structural heart diseases other than IHD or DCM.
IHD patients or DCM patients followed up in Assyut University educational cardiology hospital will be recruited in this study.
All recruited patients will undergo full laboratory examinations and complete physical examination by at least two cardiologists at the hospital. All patients will undergo echocardiographic examination for structural heart disease detection and confirmation and accurate measurement of ejection fraction for heart failure staging.
Recruited patients will be allocated to one of the study groups according to the current clinical stage of heart failure.
Group A: IHD patients with no clinical symptoms of heart failure and normal left ventricular systolic function (pre-clinical stage or stage B heart failure) Group B: IHD patients with typical clinical symptoms of heart failure together with reduced ejection fraction < 35% (stage C heart failure) Group C: IHD patients with decompensated heart failure ( stage D heart failure) Group D: DCM patients with no clinical symptoms of heart failure and normal left ventricular systolic function (pre-clinical stage or stage B heart failure) Group E: DCM patients with typical clinical symptoms of heart failure together with reduced ejection fraction < 35% (stage C heart failure) Group F: DCM patients with decompensated heart failure ( stage D heart failure)
Patients will be recruited from the external cardiology clinics, internal cardiology department and intensive cardiac care unit of Assyut University teaching Cardiology Hospital.
Sample size calculation We calculate the sample size of IHD patients according to the equation for the sample size of descriptive study design (43) based on the prevalence of Total CHD prevalence is 6.7% in US adults ≥ 20 years of age and the prevalence of DCM is 5-8 per 100000 (44).
N = (Z1-α/2)2 P (1 - P)/ D2 N= Sample Size Z1-α/2= Represent the number of standard errors from the mean ( constant = 1.96) P= Represent the proportion of the best guess about the value of the proportion of interest. = prevalence D= he absolute precision required on either side of the proportion, or the distance; how close to the proportion of interest the estimate is desired to be. 0.05.
The sample size required for IHD patients based on the prevalence rate of 0.067 will be 96 patients.
The sample size required for DCM patients based on the prevalence rate of 0.008 will be 11 patients The total sample size( including control subjects) is 107 patients.
- Cardiac biopsy
Bioptome will be used for ventricular tissue extraction (biopsy). The extracted specimens will be reserved in liquid nitrogen in - 80 Celsius degree.
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Kits, chemicals, and consumables importation
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RNA extraction Total RNA will be extracted from tissue specimens of patients with different clinical heart failure stages and healthy people using Tempus TM spin RNA Isolation Kit in Tempus TM Blood RNA Tube recommended by the manufacturer's instruction (Life Technologies Invitrogen, USA). (45) (Appendix A)
7- RNA sequencing
(Thermofisher Ion Torrent platform): option A
Truly complete transcriptome isolation for next-generation sequencing analysis will be produced using The RiboMinus™ Eukaryote System v2 ( Appendix C). The Ion Total RNA-Seq Kit v2 will be used for simple RNA library construction (Appendix D). The Ion Chef System will be used for walk-away template preparation and chip loading (Appendix E). The Ion GeneStudio S5 systems will be utilized for sequencing of gene panels all the way to the transcriptome (Appendix F). Torrent Suite Software automatically provides the sequence reads in exportable FASTQ or BAM formats (Appendix G).
8- Bioinformatics and data analysis
The Torrent Suite Software "RNASeqAnalysis" Plugin will be employed for initial analysis including RNA transcript alignment and quantification. We will use hg19 as a human reference genome, where full and partial alignments will be detected using both STAR and Bowtie2 mappers. Picard tools will be applied to count reads aligned to each gene, whereas cufflinks software will be used to represent different isoforms for each gene. Comprehensive plots will be generated for individual samples, as well as plots comparing samples . The further specific analysis will be achieved using in-house developed scripts (in python or R) for fold change calculation, identifying up and down-regulated genes/isoforms and pathway enrichment analysis using Cytoscape software. Pathway enrichment analysis can identify pathways that may have a role in the disease on a time series data. We will apply an integrative Multi-Omics approach to integrate and correlate data we obtained from both types of data (RNA-Seq).
9- manuscript writing and publication
We expect the publication of this project results in at least two high impact factor journals in the molecular genetics field and cardiology field.
B.8. Qualifications and competencies (max 2 pages):
The most important qualification is the availability of professional cardiology and molecular genetic experts that will be able to expeditiously conduct the study. Also, Highly expensive fundamentally critical instruments required for both microarray gene expression profiling and whole transcriptome ion torrent RNA sequencing are available in the molecular labs of the national research center, Magdy Yaacoub foundation and many other private molecular labs in Egypt.
The most important competencies are the high cost of the kits, chemicals, consumables and some minor instruments that should be purchased for the complete conductance of the project.
Study Design
Arms and Interventions
| Arm | Intervention/Treatment |
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| Control subjects control cases who were diagonosed as healthy subjects after diagnostic catheterization |
Genetic: quantitative polymerase chain reaction(QPCR) for seven genes
quantitative polymerase chain reaction(QPCR) for seven genes
Procedure: Cardiac biopsy
All study subjects underwent cardiac biopsy after PCI in order to take two speciments from the left ventricle using judkin right seven french catheters for radial access and cardiac bioptome 2.3 mm wide. All cardiac specimens were collected in cryotubes immediately inserted in liquid nitrogen container and then stored in -80 refregrators.
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| Stable coronary artery disease patients with stable coronary artery disease undergoing elective PCI who were diagnozed with coronary lesion/s and underwent stent implantation without previous history of myocardial infarction |
Genetic: quantitative polymerase chain reaction(QPCR) for seven genes
quantitative polymerase chain reaction(QPCR) for seven genes
Procedure: Cardiac biopsy
All study subjects underwent cardiac biopsy after PCI in order to take two speciments from the left ventricle using judkin right seven french catheters for radial access and cardiac bioptome 2.3 mm wide. All cardiac specimens were collected in cryotubes immediately inserted in liquid nitrogen container and then stored in -80 refregrators.
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| Myocardial infarction patients patients with history of myocardial infarction or with acute myocardial infarction undergoing primary PCI |
Genetic: quantitative polymerase chain reaction(QPCR) for seven genes
quantitative polymerase chain reaction(QPCR) for seven genes
Procedure: Cardiac biopsy
All study subjects underwent cardiac biopsy after PCI in order to take two speciments from the left ventricle using judkin right seven french catheters for radial access and cardiac bioptome 2.3 mm wide. All cardiac specimens were collected in cryotubes immediately inserted in liquid nitrogen container and then stored in -80 refregrators.
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| Ischaemic cardiomyopathy patients patients with ischaemic cardiomyopathy diagnosed by ejection fraction less than 35% with history of coronary artery disease or myocardial infarction or those with acute myocardial infarction with reduced ejection fraction. |
Genetic: quantitative polymerase chain reaction(QPCR) for seven genes
quantitative polymerase chain reaction(QPCR) for seven genes
Procedure: Cardiac biopsy
All study subjects underwent cardiac biopsy after PCI in order to take two speciments from the left ventricle using judkin right seven french catheters for radial access and cardiac bioptome 2.3 mm wide. All cardiac specimens were collected in cryotubes immediately inserted in liquid nitrogen container and then stored in -80 refregrators.
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Outcome Measures
Primary Outcome Measures
- Genetic pathway of pathological progression from stable coronary artery disease to myocardial infarction and finally to ischaemic cardiomyopathy [2 years]
The expected results may explain novel genetic pathways of the clinical progression of asymptomatic structural cardiac disorder (Stable coronary artery disease) to typically symptomatic ischaemic cardiomyopathy. Gene expression profiling of seven genes in RNA of all cardiac specimens was measured using QPCR analysis.
Eligibility Criteria
Criteria
Inclusion criteria The enrolled patients aged 20-80 years with stable coronary artery disease undergoing elective PCI or patients with unstable angina or myocardial infarction undergoing rescue PCI and admitted to the cardiology hospital (Asyut University, Asyut, Egypt) were recruited in this study.
Exclusion criteria Patients with dilated cardiomyopathy were execluded from this study.
Contacts and Locations
Locations
| Site | City | State | Country | Postal Code | |
|---|---|---|---|---|---|
| 1 | cardiology hospital (Asyut University, Asyut, Egypt) | Asyūţ | Egypt |
Sponsors and Collaborators
- Cairo University
Investigators
None specified.Study Documents (Full-Text)
None provided.More Information
Publications
None provided.- RHFCS1