Review
Current gene therapy trials for vascular diseases Paavo J Halonen, Jussi Nurro, Antti Kuivanen & Seppo Yla¨-Herttuala† Introduction
2.
Gene therapy trials currently in the clinic for cardiovascular diseases
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†
1.
3.
Expert opinion
University of Eastern Finland, A. I. Virtanen Institute, Department of Biotechnology and Molecular Medicine, Kuopio, Finland
Introduction: In the previous gene therapy trials for vascular diseases, safety of the therapies has been demonstrated with some evidence for clinical benefits. In the future, it will be important to also test the potential clinical benefits of the treatments in randomized and controlled trials with sufficient numbers of patients. Areas covered: This review covers 15 currently ongoing cardiovascular gene therapy trials that aim to treat coronary artery disease, heart failure and peripheral artery disease. This review summarizes current trials and their main features in the cardiovascular field. Expert opinion: In the gene therapy trials for vascular diseases, some limiting factors are still present. The trials have enrolled mainly elderly and severely affected patients who might not have the capacity to respond optimally to the therapies. Also, major cardiac adverse events, major amputations, mortality and other very demanding hard clinical end points have been used in relatively small patient populations. Therefore, there is an urgent need to enroll less severely affected patients and to use more informative surrogate end points in the forthcoming clinical trials. Keywords: clinical trial, coronary artery disease, gene therapy, heart failure, peripheral artery disease, vascular disease Expert Opin. Biol. Ther. (2014) 14(3):327-336
1.
Introduction
Cardiovascular diseases have long been the most common cause of death worldwide [1]. This group of diseases consists of the pathologies of the circulatory system, including atherosclerosis in its different forms and heart failure (HF). Atherosclerosis is responsible for most of the vascular diseases in the form of coronary artery disease (CAD) and peripheral artery disease (PAD). Noninvasive treatments for CAD and PAD include different prescription medications and lifestyle changes. In patients with advanced disease, these conditions are conventionally treated invasively using balloon angioplasty and stenting or bypass surgery to increase blood flow to the ischemic tissues. However, due to the highly invasive nature of these procedures, they are not suitable for every patient. Gene therapy has been proposed as a new way to treat ischemic diseases with a plethora of promising approaches demonstrating their efficacy in preclinical studies [2-6]. Many vascular gene therapy clinical trials have already been performed and published, demonstrating the safety of the treatments [3,7,8]. This review covers the gene therapy trials for vascular diseases that are currently ongoing in the clinic (Tables 1 and 2) and tries to put them in the context of previous trials and points out the future direction for gene therapy aimed to treat vascular diseases. Gene therapy is a fairly new paradigm in modern medicine and it holds a great promise. The number of clinical gene therapy trials has quickly risen since the first trial by Rosenberg et al. in 1989 [9] to over 1800 clinical gene therapy trials that were ongoing, completed or approved worldwide by 2012 [10]. However, the field has not 10.1517/14712598.2014.872237 © 2014 Informa UK, Ltd. ISSN 1471-2598, e-ISSN 1744-7682 All rights reserved: reproduction in whole or in part not permitted
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P. J. Halonen et al.
Article highlights. . .
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Previous gene therapy trials for cardiovascular diseases have mainly demonstrated the safety of the treatments. There are currently 15 ongoing clinical gene therapy trials for cardiovascular diseases that are aimed to treat CAD, HF or PAD. In the current clinical trials, the induction of therapeutic angiogenesis is the most common treatment strategy for ischemic conditions. Most of the current trials enroll ‘no-option’ patients who are not candidates for conventional treatments such as percutaneous coronary intervention or bypass grafting in the case of coronary or PAD. Forthcoming trials ought to enroll less severely affected patients and use more informative functional and metabolic surrogate end points to provide more accurate information about the effectiveness of the treatments.
This box summarizes key points contained in the article.
experienced a smooth transition from initial preclinical studies to clinical trials but instead has faced many challenges. The death of Jesse Gelsinger in 1999 due to complications caused by the vector administration in a gene therapy trial for ornithine transcarbamylase deficiency left the entire field in crisis for many years [11]. However, many lessons were learnt as the field recovered from the tragedy [12]. Recently, a new milestone has been achieved in gene therapy research as Glybera received its marketing authorization from the European Commission in November 2012, and thus, became the first commercially approved gene therapy product in the EU [13]. Most of the ongoing, completed or approved gene therapy trials are in the field of cancer research (64.4% of the total trials), with cardiovascular diseases being the third most common disease group (8.4%), just behind monogenic diseases (8.7%) [10]. The goals of cardiovascular gene therapy vary greatly, but in all approaches, the main concept is the gene transfer-induced overexpression of a therapeutic protein. Gene therapy has been studied, for instance, in the induction of myocardial protection, myocardial regeneration and prevention of restenosis or bypass failure. However, the most common approach in cardiovascular gene therapy trials has been to induce therapeutic angiogenesis to increase blood flow in the ischemic myocardium or lower limbs [10]. In most of the trials with this approach, the different members of either VEGF or fibroblast growth factor (FGF) family have been used [4,10]. 2. Gene therapy trials currently in the clinic for cardiovascular diseases
This review examines the gene therapy trials for cardiovascular diseases that are currently in the clinic. In addition to the 328
published literature, information has been collected from ClinicalTrials.gov (http://clinicaltrials.gov), Gene Therapy Clinical Trials Worldwide databases (http://www.abedia. com/wiley/index.html) and European Clinical Trials Database (EudraCT, https://eudract.ema.europa.eu).
Gene therapy trials for coronary heart disease and myocardial ischemia
2.1
There are currently five ongoing clinical gene therapy trials that aim to treat patients with coronary heart disease (Table 1). Ad5FGF-4 Two of the trials are continuations of the previous Angiogenic Gene Therapy (AGENT) clinical trials [14,15], and are known as ASPIRE [16] and AWARE [17]. In both of these Phase III trials, adenovirus serotype 5 (Ad5)-mediated FGF-4 gene transfer is used to relieve myocardial ischemia by inducing therapeutic angiogenesis. The gene transfer is delivered as a one-time intracoronary infusion. In a pooled analysis of the previous AGENT-3 and AGENT-4 trials, a significant gender-specific beneficial effect of the Ad5FGF-4 treatment was found on total exercise tolerance time, time to 1 mm ST-segment depression and time to angina and also the Canadian Cardiovascular Society (CCS) grading of angina pectoris in women [15]. Both ASPIRE and AWARE recruit patients who are not candidates for standard revascularization procedures. In the ASPIRE trial, the gene transfer is performed during induced transient ischemia, and a total viral dose of 6 109 viral particles (vp) is used [16]. The study enrolls patients with stable angina and confirmed CAD and is estimated to enroll 100 patients. The primary end point in this trial is the change in reversible perfusion defect size (RPDS) as measured using ATP single-photon emission computed tomography (SPECT) with technetium-99 m. These measurements are done at baseline and 8 weeks after the treatment. In addition, other secondary end points are measured at 8 weeks, and also long-term (12 months) safety of the treatment is assessed by serious adverse events. It should also be noted, that in the ASPIRE trial, gene therapy is compared to standard care, without a placebo group in an open-label design. The AWARE trial [17] standing for ‘Angiogenesis in Women with Angina pectoris who are not candidates for REvascularization’ uses the same treatment groups as the AGENT-3 and AGENT-4 trials. It is a randomized, double-blinded and placebo-controlled clinical trial, estimated to enroll a total of 300 patients with severe angina (CCS III -- IV). The difference in this trial as compared to the AGENT-3 and AGENT-4 trials is that this trial focuses only on women with stable angina, as women were the only subgroup to benefit from the treatment in the AGENT trials, where Ad5FGF-4 was administered on low dose (109 vp) in one group and on high dose (1010 vp) in another [15]. 2.1.1
Expert Opin. Biol. Ther. (2014) 14(3)
CT.gov NCT00787059
AC6 Gene Transfer for CHF
USA
USA, SWE, DEU, BEL, NLD, UK, DNK, POL FRA
USA
HF
I/II
II
IIb
HF
HF
II
I/II
I/II
III
III
I
Phase
HF
CAD
CAD
CAD
CAD
CAD
Indication
AC6
SERCA2a
SERCA2a
SDF-1
VEGF, AdVEGFAII6A+
HGF-X7 (VM202)
FGF-4
FGF-4
VEGF-DdNdD
Gene
Ad
AAV
AAV
Plasmid
Ad
Plasmid
Ad
Ad
Ad
Vector
Placebo
Placebo
Placebo
Placebo
1013 DRP
1013 DRP
3.2 109 -3.2 1011 in 5 dose groups
AdNull
108 pu, 109 pu, 1010 pu
15, 30 mg
No control
1, 2, 3 mg
Placebo
Standard care
6 109 vp one-time infusion
na
Electroanatomical mapping
Control
109, 1010, 1011 vp, 10 injections, 200 µL per infection
Dose
IC infusion
IC infusion
Endomyocardial injections, helical infusion catheter IC infusion
Intramyocardial injections, thoracotomy
Endocardial injections, NOGA
IC infusion
IC infusion, transient-induced ischemia
Endocardial injections, NOGA
Administration
56
44
ETT, rate of LV pressure and decline before and during dobutamine stress
LVEDV with CT
HF-related hospitalizations
6MWD, QOL
90
200
Time to 1 mm ST depression during ETT
Safety and tolerability
Ischemic ECG changes during ETT
Safety and tolerability, AEs, laboratory parameters, VBD, aAd-ABs, VEGF-levels RPDS with ATP-SPECT
Primary end points
41
12
300
100
30
Enrollment
Time to terminal event (all-cause death, heart transplant, LVAD implantation) Cardiac hemodynamics, VO2Max, QOL, NT-ProBNP, time to terminal events (all-cause death, heart transplant, LVAD implantation), hospitalizations Symptoms, hemodynamics, ICD discharge frequency
QOL, NYHA, time to first HF decompensation, MACE, AEs, SAEs
CCS
Angina frequency and NTG use, QOL, CCS, AEs and clinical laboratory testing, SAEs Total ETT time, Ischemic ECG changes during ETT, angina during ETT, angina frequency and NTG use, CCS, QOL, AEs and clinical laboratory testing, long-term safety ETT, SPECT, cardiac MRI and use of anti-anginal medications
Arrhythmias in 24 h Holter, improved symptoms, QOL and medication
Secondary end points
Secondary end
LVF with ECHO before and during dobutamine stress
LV measurements with CT, LVF with ECHO
LVEF with ECHO
Exercise-stress ECHO, cardiac MRI with and without adenosine stress
SPECT, cardiac MRI
RPDS with SPECT, LVEF with gated SPECT
Perfusion with MRI and PET, LVF with ECHO
point (imaging)
6MWD: 6-Min walking distance; AAV: Adeno-associated virus; AB: Antibody; AD: Adenovirus; AE: Adverse effect; CAD: Coronary artery disease; CT: Computer tomography; CT.gov: ClinicalTrials.gov; ECG: Electrocardiogram; ECHO: Echocardiography; ETT: Exercise tolerance test; HF: Heart failure; IC: Intracoronary; ICD: Implantable cardioverter defibrillator; LVEF: Left ventricular ejection fraction; LVAD: Left ventricular assisting devise; LVEDV: Left ventricular end diastolic volume; MACE: Major adverse cardiac events; MRI: Magnetic resonance imaging; NTG: Nitroglycerin medication; NT-ProBNP: N-terminal prohormone of brain natriuretic peptide; QOL: Quality of life; RPDS: Reversible perfusion defect size; SAE: Serious adverse effect; SPECT: Single-photon emission computed tomography; VBD: Vector biodistribution; VO2Max: Maximal oxygen consumption.
CT.gov NCT01966887
CT.gov NCT01643590
AGENT-HF
USA
CT.gov NCT01757223
CT.gov NCT01643330
USA
CT.gov NCT01002495
Gene Therapy for the Treatment of Chronic Stable Angina Administration of AdVEGF-All6A+ to Myocardium of Individuals With Diffuse CAD Via Minimally Invasive Surgery STOP-HF
CUPID-2b
USA
CT.gov NCT00438867
AWARE
RUS
CT.gov NCT01550614
ASPIRE
FIN
Country
CT.gov NCT01002430
Identifier
KAT301
or name
Abbreviation
Table 1. Ongoing clinical trials for CAD and HF.
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2.1.2
AdVEGFs
VEGFs are used in two currently ongoing trials for the treatment of myocardial ischemia with the purpose of inducing therapeutic angiogenesis. The first one, Kuopio Angiogenesis Trial 301 (KAT301) is a Phase I/IIa randomized, singleblinded trial enrolling patients with significant angina (CCS II -- III) who are not candidates for revascularization [18]. Total enrollment will be 30 patients. Gene transfers with AdVEGF-D, as opposed to AdVEGF-A and VEGF-A plasmid liposomes used in the previous KAT-trial [7], are performed percutaneously using an endocardial injection system (NOGA) with increasing doses of 1 109, 1 1010 or 1 1011 vp injected into 10 sites in the ischemic myocardium. Control patients receive electroanatomic mapping but no gene transfer. The primary outcome measures will assess the safety and tolerability of the gene transfer, with the secondary end points measuring the efficacy of the treatment with MRI, positron emission tomography (PET) to measure myocardial perfusion, exercise tolerance test (ETT) to measure functional capacity and echocardiography to measure left ventricular function. Another clinical trial using VEGF to treat myocardial ischemia uses AdVEGF-AII6A+ to treat patients with diffuse CAD [19]. AdVEGF-AII6A+ expresses all three major isoforms of human VEGF-A [20]. This trial consists of two parts, the first part being an open-label, dose--escalation study and the second part being a randomized, double-blinded and placebo-controlled study. In the second part, an empty adenovirus (AdNull) is used as a control. The trial enrolls patients with CAD and demonstrable reversible myocardial ischemia. Gene transfer is performed using intramyocardial injections during thoracotomy. The primary end point of the trial is the time to 1 mm ST-depression during ETT. Human hepatocyte growth factor One current clinical trial uses plasmid encoding human hepatocyte growth factor (HGF-X7, also called VM202) for the treatment of chronic stable angina [21]. It is a Phase I/II, open-label, dose--escalation study, where three groups are given 1, 2 or 3 mg of the HGF-plasmid, respectively, as endocardial injections using the NOGA cardiac navigation system. The primary end point of the study is to evaluate the safety and tolerability of the treatment. Also, an ETT, SPECT and cardiac MRI are used to measure the potential effects of the treatment and the angiogenic potential of the HGF-X7-plasmid [21]. 2.1.3
Gene therapy trials for HF There are four ongoing clinical gene therapy trials aimed to treat HF known as STOP-HF, CUPID-2b, AGENT-HF and AC6 Gene Transfer for CHF (Table 1). STOP-HF is a Phase II randomized, double-blinded, placebo-controlled trial that evaluates the safety and efficacy of stromal cell-derived factor 1 (SDF-1) encoding plasmid for the treatment of ischemic 2.2
330
HF [22]. Gene transfers are done as endomyocardial injections using a Helical Infusion Catheter (BioCardia) with the SDF-1 plasmid (low dose 15 mg, high dose 30 mg) or placebo [22]. The primary outcome measures of the trial include 6-min walking distance and the quality of life (Minnesota Living with Heart Failure Questionnaire) 4 months after the treatment. Also, left ventricular ejection fraction (LVEF) using echocardiography, New York Heart Association (NYHA) functional classification, major adverse cardiac events (MACE) and other end points will be measured. The study is expected to enroll 90 patients [22]. CUPID-2b is a Phase II, randomized, double-blinded placebo-controlled trial in which HF patients are treated using adeno-associated virus serotype 1 (AAV1)-mediated sarcoplasmic reticulum calcium ATPase 2a (SERCA2a) or placebo [23]. The AAV1-SERCA2a gene transfer is used to normalize deficient sarcoplasmic reticulum calcium uptake and intracellular calcium cycling in the cardiomyocytes of a failing heart to improve left ventricular function [24]. CUPID-2b is a larger confirmatory trial of the previous CUPID-2 trial, which consisted of only 39 patients and where AAV1-SERCA2a was given at three different doses of 6 1011, 3 1012 or 1 1013 DNase-resistant particles (DRP) in addition to the placebo group [24]. Despite its low statistical power, CUPID-2 still suggested that the highest dose of AAV1-SERCA2a gene therapy could achieve clinically significant improvements in patients with advanced HF [24]. CUPID-2b enrolls patients with chronic systolic HF or non-ischemic cardiomyopathy, and a total enrollment of 200 is expected. Participants are screened for neutralizing AAV1 antibodies to exclude patients with immunity to the vector. Gene therapy is administered as a one-time intracoronary infusion of 1 1013 DRP (corresponding to the highest dose used in CUPID-2) of AAV1-SERCA2a or placebo. Primary outcome measures include HF-related hospitalizations that are related to terminal events, such as heart transplantation, left ventricular assist device implantation or all-cause mortality. AGENT-HF (AAV1-CMV-SERCA2a Gene Therapy trial in Heart Failure) is a Phase II, randomized, double-blinded, placebo-controlled trial with a similar treatment protocol to the CUPID-2b trial [23,25] and takes place in France. Intracoronary AAV1-SERCA2a gene therapy is used to treat congestive HF, and a total of 44 patients with severe ischemic or non-ischemic HF (NYHA III/IV) will be enrolled. Patients are also screened for neutralizing AAV1 antibodies. The primary end point in the trials is left ventricle end-diastolic volume measured using computer tomography. Time to terminal events (heart transplantation, left ventricular assist device implantation or all-cause mortality), maximal oxygen consumption, left ventricle function measured with echocardiography and other secondary outcome measures are also recorded. AC6 Gene Transfer for CHF is a Phase I/II, randomized, double-blinded, placebo-controlled trial where gene therapy with adenoviral adenylyl cyclase 6 (AdAC6) is used to treat
Expert Opin. Biol. Ther. (2014) 14(3)
EudraCT FIN 2012-001019-22
KAT-PAD 101
Expert Opin. Biol. Ther. (2014) 14(3) I/II
I/II
II
II
II
I
Vector
Plasmid
Plasmid
Plasmid
VEGF-DdNdC Ad
MGA: Ang-1, RV VEGF-A165
HGF-X7 (NL003)
HGF-X7 (VM202)
SDF-1
MGA: Ang-1, RV VEGF-A165
Gene
No control
Control
Saline
MGA low therapeutic dose, MGA high therapeutic dose 3 1010 vp, 3 1011 vp
28
30
2 -- 4 IM injections Saline
200
50
48
12
IA infusion
96 IM injections
64 IM injections
IM injection
IA infusion
Administration Enrollment
Two treatment groups
4, 6 or 8 mg Saline once at 3 time points
8 or 16 mg
4, 8 or 16 mg Placebo
Escalating doses
Dose
Safety, clinical status, vital signs, serum VEGF-D levels, AEs
AEs
Ulcer area, VAS
VAS
AEs, SAEs, laboratory assessments
AEs
Primary end points
Pain and ulcer assessment, ABI, clinical laboratory testing, medical therapy, symptoms
VAS, TcPO2, ABI and TBI, wound healing, VAS, QOL, amputation rate, mortality TcPO2, VAS, ulcer area, ABI and TBI, QOL, complete ulcer healing, ulcer healing after gangrene treatment, major amputation rate Critical limb ischemia symptoms
Major and minor amputations, survival, QOL, ulcer healing, pressure measurements
PAD symptoms
Secondary end points
MRA, MRI, contrastenhanced ultrasound, Doppler ultrasound
Perfusion with MRA
Secondary end point (imaging)
ABI: Ankle-brachial index; AE: Adverse effect; Ad: Adenovirus; CT.gov: ClinicalTrials.gov; ECHO: Echocardiography; IA: Intra-arterial; IM: Intramuscular; LVEF: Left ventricular ejection fraction; LVAD: Left ventricular assisting devise; MRA: Magnetic resonance angiography; MRI: Magnetic resonance imaging; PAD: Peripheral artery disease; QOL: Quality of life; RV: Retrovirus; SAE: Serious adverse effect; SPECT: Single-photon emission computed tomography; TBI: Tibial-brachia index; TcPO2: Transcutaneous oxygen tension; VAS: Visual analog scale; vp: Viral particle.
ISR
CT.gov NCT00956332
Safety Study of MultiGeneAngio in Patients with Chronic Critical Limb Ischemia
CHN
CT.gov NCT01548378
Safety and Efficacy Study Using Gene Therapy for Critical Limb Ischemia
USA, KOR
USA, IND
CT.gov NCT01410331
CT.gov NCT01064440
USA
CT.gov NCT00390767
Safety Study of MultiGeneAngio in Patients With Peripheral Arterial Disease Study to Evaluate the Safety and Efficacy of JVS-100 Administered to Adults with Critical Limb Ischemia Safety and Efficacy Study Using Gene Therapy for Critical Limb Ischemia
Country Phase
Identifier
Abbreviation or name
Table 2. Ongoing clinical trials for PAD.
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Current gene therapy trials for vascular diseases
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P. J. Halonen et al.
patients with congestive HF [26]. AdAC6 gene transfer is used to improve left ventricular contractility. This study will enroll a total of 56 patients with compensated congestive HF. Intracoronary delivery of AdAC6 or placebo is performed on escalating doses from 3.2 109 to 3.2 1011 vp at five different doses [26]. The primary end points in the trial include ETT, left ventricular function with echocardiography and the rate of left ventricular pressure development and decline, both during rest and dobutamine-induced stress. Congestive heart failure (CHF) symptoms, hemodynamics and implantable cardioverter defibrillator discharge frequency are collected as the secondary outcome measures. Gene therapy trials for peripheral ischemia There are currently six gene therapy trials in the clinic for PAD and peripheral ischemia (Table 2). 2.3
Human hepatocyte growth factor (HGF-X7) There are two clinical trials where HGF is used to treat critical limb ischemia. Both trials use the HGF-X7 plasmid (also known as VM202 or NL003 depending on the country where the trial takes place) that expresses two HGF isoforms, HGF-728 and HGF-723 [27,28]. The same plasmid is also used in another trial for the treatment of chronic stable angina [21]. The first trial takes place in the USA and Korea and is a Phase II, randomized, double-blinded, placebo-controlled study in which the HGF-X7 plasmid is used to treat critical limb ischemia [27]. The study enrolls patients with limb ischemia (Rutherford category 4 or 5) who are poor candidates for bypass grafting or percutaneous angioplasty with a total enrollment of 50 patients. Patients are randomized into three groups: low dose (8 mg HGF-X7), high dose (16 mg HGFX7) and placebo (saline). The plasmids are given as intramuscular injections at multiple time points, 4 mg per time point. In the high dose group, HGF-X7 is given at time points days 0, 14, 28 and 42, whereas in the low dose group HGF-X7 is given only at time points days 0 and 14 with saline given at time points days 28 and 42. In the placebo group, saline is given at all aforementioned time points. At all of the time points, the treatment is administered using 16 intramuscular injections. The primary end point of the trial is the change in pain level (visual analog scale [VAS]) from baseline after a 9-month follow up. Also, mortality, amputation rate, tissue oxygenation (TcPO2) and other secondary outcome measures will be recorded. The other clinical trial using HGF-X7 plasmid is a Phase II, randomized, placebo-controlled study taking place in China. This study enrolls a total of 200 patients with Rutherford 4- or 5-grade limb ischemia. The study consists of four groups: low dose (4 mg), intermediate dose (6 mg) and high dose (8 mg) of HGF-X7 and placebo group. The therapy is administered as 32 intramuscular injections at time points days 0, 14 and 28. The primary end points are the difference in ulcer area from baseline and the difference in pain level 2.3.1
332
using VAS after a 6-month follow up. Other secondary outcome measures, such as difference in TcPO2, amputation rate and ulcer healing will be recorded. SDF-1 The same SDF-1 encoding plasmid that is used in the STOPHF trial is also used in a trial for critical limb ischemia [29]. This trial is a Phase II, randomized, double-blinded, placebo-controlled dose--escalation study evaluating the safety and efficacy of the treatment. The study will enroll 48 patients with Rutherford category 4- or 5-grade limb ischemia and poor candidacy for bypass grafting or percutaneous balloon angioplasty. Patients are assigned into four cohorts. Patients in the first two cohorts receive 4 or 8 mg of the plasmid or placebo as 8 injections, and patients in the other two cohorts receive 8 or 16 mg of the plasmid or placebo as 16 injections. The primary end point of the study is to investigate the safety of the plasmid at different doses. As secondary outcome measures, amputations, survival, quality of life, ulcer healing and different pressure measurements will be assessed. 2.3.2
AdVEGF-D In addition to the KAT301-trial, where AdVEGF-D is used to treat myocardial ischemia, in the Kuopio Angiogenesis Trial in Peripheral Arterial Disease (KAT-PAD 101), AdVEGF-D is used to treat critical lower limb ischemia (personal communications with Dr Petra Korpisalo-Pirinen, EudraCT: 2012-001019-22 [30]). This is a Phase I/II, a singleblinded, placebo-controlled trial. Gene transfers are performed into 2 -- 4 sites as intramuscular injections. The KAT-PAD 101 trial enrolls patients with chronic limb ischemia (according to Rutherford and Fontaine classification) and to whom surgical bypass is considered the most suitable treatment. Gene transfers are performed a day prior to a scheduled bypass operation. The end points include safety, feasibility, quality of life, clinical status, anti-adenoviral antibodies, pain and ulcer assessment, magnetic resonance angiography (MRA), MRI, contrast-enhanced ultrasound, Doppler ultrasound, ankle-brachial index (ABI), adverse events and concomitant medical therapy. 2.3.3
MultiGeneAngio In addition to trials where gene therapy is performed using in vivo gene transfer, there are also two cell therapy trials where cells isolated from the patient are transduced ex-vivo and then reintroduced to the patient. MultiGeneAngio is a cell therapy product that is composed of a cell suspension of endothelial and smooth muscle cells that are isolated from a vein segment of the patient. After the isolation, modification by angiogenic gene transfer and ex-vivo expansion, the cell suspension is reintroduced back to the patient as an intra-arterial injection at the site of the blockage. There are currently two clinical trials investigating the use of MultiGeneAngio in treatment of PAD -- one in the USA [31] and another in Israel [32]. The first one is a Phase I, 2.3.4
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Current gene therapy trials for vascular diseases
single group, open-label safety study, where patients receive escalating doses of MultiGeneAngio. The current enrollment is 12 patients and the study does not enroll new patients. The study enrolled patients with exercise-limiting intermittent claudication and PAD symptoms in one or both legs. The patients are monitored up to 15 years after the treatment for adverse effects. As a secondary outcome, PAD symptoms will also be monitored. The Israeli trial is a Phase I/II, randomized, open-label study, with two dose levels, where patients are given MultiGeneAngio in low and intermediate doses. The current enrollment is 28 patients and this study is not recruiting patients. This study enrolled patients with ischemic rest pain (Rutherford 4) or nonhealing wounds (Rutherford 5).
Conclusion There are many common denominators that can be seen in the current clinical trials for vascular diseases. Myocardial and peripheral ischemia are both conditions that result from the same cause: the lack of proper blood flow into the tissue. This is also reflected in the current clinical trials, as multiple gene therapy products are under investigation for the treatment of both myocardial and peripheral ischemia. The HGF-X7 plasmid encoding for HGF and AdVEGF-D treatment are studied in the settings of both myocardial and critical limb ischemia. The SDF-1 plasmid is studied for both the treatment of ischemic HF (STOP-HF) and ischemia caused by PAD. Several trials enroll so-called ‘no-option patients’ who are not suitable for conventional treatments such as bypass grafting or percutaneous balloon angioplasty and stenting. This means that the patients enrolled in the gene therapy trials are elderly and have many concomitant diseases, such as type 2 diabetes. The only exception is the KAT-PAD 101 trial that specifically includes patients to whom surgical bypass is considered the best treatment, as opposed to enrolling patients who are poor candidates for revascularization. Primary outcome measures in most trials measure either safety or hard clinical end points, such as MACE, amputations or mortality. In these kinds of settings, it might be difficult to achieve significant results when using relatively small patient populations that consist mostly of ‘no-option’ patients [4]. In 8 of these 15 trials, either intramyocardial or intramuscular injections are used for the administration of gene therapy as opposed to an intracoronary or intra-arterial infusion. Ten trials use viral vectors instead of plasmid DNA to deliver the therapeutic gene. The concept of therapeutic angiogenesis has persisted as the main strategy for novel therapies to increase perfusion in the ischemic tissues. In gene therapy for non-ischemic vascular diseases, CUPID-2b, AGENT-HF and AC6 Gene Therapy for CHF use a different approach and is aimed at improving the contractility of the failing heart. 2.4
3.
Expert opinion
Inclusion criteria Previous cardiovascular gene therapy trials have mainly demonstrated the safety and feasibility of the therapies. The primary end points of previous Phase I/II studies have mostly concerned safety, with secondary outcome measures used to determine the actual effects of the therapy. ‘No-option’ patients with high comorbidity are enrolled in Phase I/II trials even though it is not clear whether these patients have already lost their regenerative capacity when it comes to treating ischemic conditions with gene therapy [3,4]. It has been proposed that these patients could lack the capacity to respond to angiogenic gene therapies [2,4]. A recent review by Dragneva et al. pointed out that many preclinical studies using rodent models are largely based on young and healthy animals, whereas clinical patients are very sick and suffer from multiple conditions in addition to the vascular diseases [2]. After the safety of the treatments has been confirmed, new trials should move toward treating less severely affected patients. Also, the use of gene therapy as an adjuvant treatment for conventional revascularization therapies as in the KAT-PAD 101 trial should be seriously considered. 3.1
Surrogate end points Forthcoming gene therapy trials will probably enroll relatively small patient populations when compared with large clinical trials with regular pharmaceutical compounds. Previous trials have often focused on major clinical end points, such as MACE, amputation rate and ulcer healing. These end points by themselves are very challenging and not optimal for measuring therapeutic effects with other variables affecting the outcome in limited patient populations of sick and elderly patients. The primary end points of future trials could move from MACE toward validated and more sensitive surrogate end points, enabling smaller clinical trials to provide important information about the effectiveness of the new treatments. Especially the use of functional and metabolic surrogate end points, such as LVEF and PET-perfusion imaging, glucose uptake, viability and fatty acid uptake, should be considered. Also, metabolic analyses and functional molecular imaging would provide more useful and mechanistic data that cannot be acquired by using traditional end points [2]. 3.2
Vectors The vectors used in the clinical trials should have appropriate expression profiles, tissue tropism and transduction efficiencies to enable clinically relevant transgene expression in target tissues. In many previous and current clinical trials, plasmidbased therapies are used, although the capacity of plasmid therapies to cause robust transduction in target tissues is still unclear even using large amounts of plasmid DNA [33]. It should be critically investigated if plasmid based-therapies 3.3
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can cause clinically relevant transduction efficiencies in human patients. Only vectors that have been demonstrated to achieve high enough transduction efficiencies in preclinical studies should be used in clinical studies. If robust transduction is not achieved, negative results might be caused simply by the lack of the expression of a therapeutic gene. Treatment genes Only well-established therapeutic genes should be used in future clinical trials. These would include at least VEGFs, FGFs and HGF for ischemic conditions [3]. In a Phase II study, AdVEGF was shown to induce significant growth of new vasculature in the legs of PAD patients [34]. However, this trial also failed to show any changes in amputation frequency or survival, although it demonstrated excellent longterm safety [8]. Regarding local administration, it should also be assessed whether intracellular or secreted factors will provide the best possible outcome. It could be argued that secreted factors will cover a larger and more uniform area in the target tissue, but on the other hand, for example, intracellular transcription factors might result in more physiologic effects by acting more upstream and regulating the expression of multiple genes and therefore having broader therapeutic effects. However, covering functionally relevant areas of human skeletal muscles or myocardium might prove difficult by using treatment gene coding for intracellular factors.
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3.4
Administration In spite of many successful preclinical studies using either systemic or intravascular gene therapy, the human biodistribution and pharmacokinetics of the vectors should also be taken into account. When treating local ischemia, intravascular delivery will unnecessarily increase systemic distribution of the vector when compared with local injections. The effectiveness of, for example, adenoviral VEGF gene therapy using local delivery in clinically relevant large animal models of myocardial ischemia has been demonstrated in several studies [4,35]. Although there has recently been positive reports using an intracoronary infusion of AAV1-mediated gene transfer for HF in the CUPID trial [24], there is preclinical work suggesting that intramuscular injections might be a more effective way to transduce muscle tissue than intravascular infusion [3]. It should be further investigated whether there are differences in optimal administration routes depending on the vector. The AAV1 vector used in CUPID-2b could perform better 3.5
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than adenoviral vectors if gene therapy is given as an intracoronary infusion. Studies have also suggested that gene delivery methods, such as retrograde injection, coronary sinus blockade and balloon occlusion causing higher perfusion pressure, higher coronary flow and a longer dwelling time for the vector in the vascular tree, result in a better transduction efficiency than a simple anterograde infusion [36]. These methods might prove essential in achieving robust global transduction of the target tissues, whereas focal expression induced by local gene delivery could be more suitable for other approaches, such as angiogenic gene therapy to the infarct border [36]. Also, local gene delivery methods limit the possible side effects, such as the increase in tissue permeability in angiogenic therapies, to the target sites. New developments in tissue specific vectors, such as cardiotropic AAV vectors, and tissuespecific promoters might alter the situation in the future [37]. However, local delivery would seem to be the most logical approach for most cardiovascular conditions at the moment. Ethical considerations Excellent safety and tolerability of cardiovascular gene therapy has been demonstrated in multiple previous clinical trials using mainly ‘no-option’ patients [7,8,15]. Without forgetting the case of Jesse Gelsinger and with the lessons learned from it [12], it seems very unlikely for gene therapy to cause serious adverse effects in future clinical trials. Cardiovascular diseases are the most common cause of death [1] and an enormous public health issue especially in the western world. Novel therapies such as gene therapy are, thus, needed to treat patients with these increasingly ubiquitous diseases in aging populations. 3.6
Expert opinion conclusion Forthcoming clinical trials should begin enrolling less severely affected patients, since the safety of many gene transfer modalities have already been well established. Broader emphasis should also be given on new imaging-based, metabolic and functional surrogate end points in addition to the conventional clinical outcome measures to better capture the effects of novel therapies in relatively small patient populations. 3.7
Declaration of interest The authors have no competing interests to declare. Authors acknowledge financial support from Finnish Academy, Kuopio University Hospital Research Unit, Sigrid Juselius Foundation, European Research Council (ERC) and Finnish Heart Foundation.
Expert Opin. Biol. Ther. (2014) 14(3)
Current gene therapy trials for vascular diseases
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Affiliation Paavo J Halonen1 BMed, Jussi Nurro1 BMed, Antti Kuivanen1 BMed & Seppo Yla¨-Herttuala†1,2 MD PhD FESC † Author for correspondence 1 University of Eastern Finland, A. I. Virtanen Institute, Department of Biotechnology and Molecular Medicine, P.O. Box 1627, FIN-70211, Kuopio, Finland Tel: +358 40 355 2076; E-mail: [email protected] 2 Kuopio University Hospital, Research Unit and Gene Therapy Unit, FIN-70211, Kuopio, Finland
Current gene therapy trials for vascular diseases Paavo J Halonen, Jussi Nurro, Antti Kuivanen & Seppo Yla¨-Herttuala† Introduction
2.
Gene therapy trials currently in the clinic for cardiovascular diseases
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†
1.
3.
Expert opinion
University of Eastern Finland, A. I. Virtanen Institute, Department of Biotechnology and Molecular Medicine, Kuopio, Finland
Introduction: In the previous gene therapy trials for vascular diseases, safety of the therapies has been demonstrated with some evidence for clinical benefits. In the future, it will be important to also test the potential clinical benefits of the treatments in randomized and controlled trials with sufficient numbers of patients. Areas covered: This review covers 15 currently ongoing cardiovascular gene therapy trials that aim to treat coronary artery disease, heart failure and peripheral artery disease. This review summarizes current trials and their main features in the cardiovascular field. Expert opinion: In the gene therapy trials for vascular diseases, some limiting factors are still present. The trials have enrolled mainly elderly and severely affected patients who might not have the capacity to respond optimally to the therapies. Also, major cardiac adverse events, major amputations, mortality and other very demanding hard clinical end points have been used in relatively small patient populations. Therefore, there is an urgent need to enroll less severely affected patients and to use more informative surrogate end points in the forthcoming clinical trials. Keywords: clinical trial, coronary artery disease, gene therapy, heart failure, peripheral artery disease, vascular disease Expert Opin. Biol. Ther. (2014) 14(3):327-336
1.
Introduction
Cardiovascular diseases have long been the most common cause of death worldwide [1]. This group of diseases consists of the pathologies of the circulatory system, including atherosclerosis in its different forms and heart failure (HF). Atherosclerosis is responsible for most of the vascular diseases in the form of coronary artery disease (CAD) and peripheral artery disease (PAD). Noninvasive treatments for CAD and PAD include different prescription medications and lifestyle changes. In patients with advanced disease, these conditions are conventionally treated invasively using balloon angioplasty and stenting or bypass surgery to increase blood flow to the ischemic tissues. However, due to the highly invasive nature of these procedures, they are not suitable for every patient. Gene therapy has been proposed as a new way to treat ischemic diseases with a plethora of promising approaches demonstrating their efficacy in preclinical studies [2-6]. Many vascular gene therapy clinical trials have already been performed and published, demonstrating the safety of the treatments [3,7,8]. This review covers the gene therapy trials for vascular diseases that are currently ongoing in the clinic (Tables 1 and 2) and tries to put them in the context of previous trials and points out the future direction for gene therapy aimed to treat vascular diseases. Gene therapy is a fairly new paradigm in modern medicine and it holds a great promise. The number of clinical gene therapy trials has quickly risen since the first trial by Rosenberg et al. in 1989 [9] to over 1800 clinical gene therapy trials that were ongoing, completed or approved worldwide by 2012 [10]. However, the field has not 10.1517/14712598.2014.872237 © 2014 Informa UK, Ltd. ISSN 1471-2598, e-ISSN 1744-7682 All rights reserved: reproduction in whole or in part not permitted
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Article highlights. . .
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.
Previous gene therapy trials for cardiovascular diseases have mainly demonstrated the safety of the treatments. There are currently 15 ongoing clinical gene therapy trials for cardiovascular diseases that are aimed to treat CAD, HF or PAD. In the current clinical trials, the induction of therapeutic angiogenesis is the most common treatment strategy for ischemic conditions. Most of the current trials enroll ‘no-option’ patients who are not candidates for conventional treatments such as percutaneous coronary intervention or bypass grafting in the case of coronary or PAD. Forthcoming trials ought to enroll less severely affected patients and use more informative functional and metabolic surrogate end points to provide more accurate information about the effectiveness of the treatments.
This box summarizes key points contained in the article.
experienced a smooth transition from initial preclinical studies to clinical trials but instead has faced many challenges. The death of Jesse Gelsinger in 1999 due to complications caused by the vector administration in a gene therapy trial for ornithine transcarbamylase deficiency left the entire field in crisis for many years [11]. However, many lessons were learnt as the field recovered from the tragedy [12]. Recently, a new milestone has been achieved in gene therapy research as Glybera received its marketing authorization from the European Commission in November 2012, and thus, became the first commercially approved gene therapy product in the EU [13]. Most of the ongoing, completed or approved gene therapy trials are in the field of cancer research (64.4% of the total trials), with cardiovascular diseases being the third most common disease group (8.4%), just behind monogenic diseases (8.7%) [10]. The goals of cardiovascular gene therapy vary greatly, but in all approaches, the main concept is the gene transfer-induced overexpression of a therapeutic protein. Gene therapy has been studied, for instance, in the induction of myocardial protection, myocardial regeneration and prevention of restenosis or bypass failure. However, the most common approach in cardiovascular gene therapy trials has been to induce therapeutic angiogenesis to increase blood flow in the ischemic myocardium or lower limbs [10]. In most of the trials with this approach, the different members of either VEGF or fibroblast growth factor (FGF) family have been used [4,10]. 2. Gene therapy trials currently in the clinic for cardiovascular diseases
This review examines the gene therapy trials for cardiovascular diseases that are currently in the clinic. In addition to the 328
published literature, information has been collected from ClinicalTrials.gov (http://clinicaltrials.gov), Gene Therapy Clinical Trials Worldwide databases (http://www.abedia. com/wiley/index.html) and European Clinical Trials Database (EudraCT, https://eudract.ema.europa.eu).
Gene therapy trials for coronary heart disease and myocardial ischemia
2.1
There are currently five ongoing clinical gene therapy trials that aim to treat patients with coronary heart disease (Table 1). Ad5FGF-4 Two of the trials are continuations of the previous Angiogenic Gene Therapy (AGENT) clinical trials [14,15], and are known as ASPIRE [16] and AWARE [17]. In both of these Phase III trials, adenovirus serotype 5 (Ad5)-mediated FGF-4 gene transfer is used to relieve myocardial ischemia by inducing therapeutic angiogenesis. The gene transfer is delivered as a one-time intracoronary infusion. In a pooled analysis of the previous AGENT-3 and AGENT-4 trials, a significant gender-specific beneficial effect of the Ad5FGF-4 treatment was found on total exercise tolerance time, time to 1 mm ST-segment depression and time to angina and also the Canadian Cardiovascular Society (CCS) grading of angina pectoris in women [15]. Both ASPIRE and AWARE recruit patients who are not candidates for standard revascularization procedures. In the ASPIRE trial, the gene transfer is performed during induced transient ischemia, and a total viral dose of 6 109 viral particles (vp) is used [16]. The study enrolls patients with stable angina and confirmed CAD and is estimated to enroll 100 patients. The primary end point in this trial is the change in reversible perfusion defect size (RPDS) as measured using ATP single-photon emission computed tomography (SPECT) with technetium-99 m. These measurements are done at baseline and 8 weeks after the treatment. In addition, other secondary end points are measured at 8 weeks, and also long-term (12 months) safety of the treatment is assessed by serious adverse events. It should also be noted, that in the ASPIRE trial, gene therapy is compared to standard care, without a placebo group in an open-label design. The AWARE trial [17] standing for ‘Angiogenesis in Women with Angina pectoris who are not candidates for REvascularization’ uses the same treatment groups as the AGENT-3 and AGENT-4 trials. It is a randomized, double-blinded and placebo-controlled clinical trial, estimated to enroll a total of 300 patients with severe angina (CCS III -- IV). The difference in this trial as compared to the AGENT-3 and AGENT-4 trials is that this trial focuses only on women with stable angina, as women were the only subgroup to benefit from the treatment in the AGENT trials, where Ad5FGF-4 was administered on low dose (109 vp) in one group and on high dose (1010 vp) in another [15]. 2.1.1
Expert Opin. Biol. Ther. (2014) 14(3)
CT.gov NCT00787059
AC6 Gene Transfer for CHF
USA
USA, SWE, DEU, BEL, NLD, UK, DNK, POL FRA
USA
HF
I/II
II
IIb
HF
HF
II
I/II
I/II
III
III
I
Phase
HF
CAD
CAD
CAD
CAD
CAD
Indication
AC6
SERCA2a
SERCA2a
SDF-1
VEGF, AdVEGFAII6A+
HGF-X7 (VM202)
FGF-4
FGF-4
VEGF-DdNdD
Gene
Ad
AAV
AAV
Plasmid
Ad
Plasmid
Ad
Ad
Ad
Vector
Placebo
Placebo
Placebo
Placebo
1013 DRP
1013 DRP
3.2 109 -3.2 1011 in 5 dose groups
AdNull
108 pu, 109 pu, 1010 pu
15, 30 mg
No control
1, 2, 3 mg
Placebo
Standard care
6 109 vp one-time infusion
na
Electroanatomical mapping
Control
109, 1010, 1011 vp, 10 injections, 200 µL per infection
Dose
IC infusion
IC infusion
Endomyocardial injections, helical infusion catheter IC infusion
Intramyocardial injections, thoracotomy
Endocardial injections, NOGA
IC infusion
IC infusion, transient-induced ischemia
Endocardial injections, NOGA
Administration
56
44
ETT, rate of LV pressure and decline before and during dobutamine stress
LVEDV with CT
HF-related hospitalizations
6MWD, QOL
90
200
Time to 1 mm ST depression during ETT
Safety and tolerability
Ischemic ECG changes during ETT
Safety and tolerability, AEs, laboratory parameters, VBD, aAd-ABs, VEGF-levels RPDS with ATP-SPECT
Primary end points
41
12
300
100
30
Enrollment
Time to terminal event (all-cause death, heart transplant, LVAD implantation) Cardiac hemodynamics, VO2Max, QOL, NT-ProBNP, time to terminal events (all-cause death, heart transplant, LVAD implantation), hospitalizations Symptoms, hemodynamics, ICD discharge frequency
QOL, NYHA, time to first HF decompensation, MACE, AEs, SAEs
CCS
Angina frequency and NTG use, QOL, CCS, AEs and clinical laboratory testing, SAEs Total ETT time, Ischemic ECG changes during ETT, angina during ETT, angina frequency and NTG use, CCS, QOL, AEs and clinical laboratory testing, long-term safety ETT, SPECT, cardiac MRI and use of anti-anginal medications
Arrhythmias in 24 h Holter, improved symptoms, QOL and medication
Secondary end points
Secondary end
LVF with ECHO before and during dobutamine stress
LV measurements with CT, LVF with ECHO
LVEF with ECHO
Exercise-stress ECHO, cardiac MRI with and without adenosine stress
SPECT, cardiac MRI
RPDS with SPECT, LVEF with gated SPECT
Perfusion with MRI and PET, LVF with ECHO
point (imaging)
6MWD: 6-Min walking distance; AAV: Adeno-associated virus; AB: Antibody; AD: Adenovirus; AE: Adverse effect; CAD: Coronary artery disease; CT: Computer tomography; CT.gov: ClinicalTrials.gov; ECG: Electrocardiogram; ECHO: Echocardiography; ETT: Exercise tolerance test; HF: Heart failure; IC: Intracoronary; ICD: Implantable cardioverter defibrillator; LVEF: Left ventricular ejection fraction; LVAD: Left ventricular assisting devise; LVEDV: Left ventricular end diastolic volume; MACE: Major adverse cardiac events; MRI: Magnetic resonance imaging; NTG: Nitroglycerin medication; NT-ProBNP: N-terminal prohormone of brain natriuretic peptide; QOL: Quality of life; RPDS: Reversible perfusion defect size; SAE: Serious adverse effect; SPECT: Single-photon emission computed tomography; VBD: Vector biodistribution; VO2Max: Maximal oxygen consumption.
CT.gov NCT01966887
CT.gov NCT01643590
AGENT-HF
USA
CT.gov NCT01757223
CT.gov NCT01643330
USA
CT.gov NCT01002495
Gene Therapy for the Treatment of Chronic Stable Angina Administration of AdVEGF-All6A+ to Myocardium of Individuals With Diffuse CAD Via Minimally Invasive Surgery STOP-HF
CUPID-2b
USA
CT.gov NCT00438867
AWARE
RUS
CT.gov NCT01550614
ASPIRE
FIN
Country
CT.gov NCT01002430
Identifier
KAT301
or name
Abbreviation
Table 1. Ongoing clinical trials for CAD and HF.
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2.1.2
AdVEGFs
VEGFs are used in two currently ongoing trials for the treatment of myocardial ischemia with the purpose of inducing therapeutic angiogenesis. The first one, Kuopio Angiogenesis Trial 301 (KAT301) is a Phase I/IIa randomized, singleblinded trial enrolling patients with significant angina (CCS II -- III) who are not candidates for revascularization [18]. Total enrollment will be 30 patients. Gene transfers with AdVEGF-D, as opposed to AdVEGF-A and VEGF-A plasmid liposomes used in the previous KAT-trial [7], are performed percutaneously using an endocardial injection system (NOGA) with increasing doses of 1 109, 1 1010 or 1 1011 vp injected into 10 sites in the ischemic myocardium. Control patients receive electroanatomic mapping but no gene transfer. The primary outcome measures will assess the safety and tolerability of the gene transfer, with the secondary end points measuring the efficacy of the treatment with MRI, positron emission tomography (PET) to measure myocardial perfusion, exercise tolerance test (ETT) to measure functional capacity and echocardiography to measure left ventricular function. Another clinical trial using VEGF to treat myocardial ischemia uses AdVEGF-AII6A+ to treat patients with diffuse CAD [19]. AdVEGF-AII6A+ expresses all three major isoforms of human VEGF-A [20]. This trial consists of two parts, the first part being an open-label, dose--escalation study and the second part being a randomized, double-blinded and placebo-controlled study. In the second part, an empty adenovirus (AdNull) is used as a control. The trial enrolls patients with CAD and demonstrable reversible myocardial ischemia. Gene transfer is performed using intramyocardial injections during thoracotomy. The primary end point of the trial is the time to 1 mm ST-depression during ETT. Human hepatocyte growth factor One current clinical trial uses plasmid encoding human hepatocyte growth factor (HGF-X7, also called VM202) for the treatment of chronic stable angina [21]. It is a Phase I/II, open-label, dose--escalation study, where three groups are given 1, 2 or 3 mg of the HGF-plasmid, respectively, as endocardial injections using the NOGA cardiac navigation system. The primary end point of the study is to evaluate the safety and tolerability of the treatment. Also, an ETT, SPECT and cardiac MRI are used to measure the potential effects of the treatment and the angiogenic potential of the HGF-X7-plasmid [21]. 2.1.3
Gene therapy trials for HF There are four ongoing clinical gene therapy trials aimed to treat HF known as STOP-HF, CUPID-2b, AGENT-HF and AC6 Gene Transfer for CHF (Table 1). STOP-HF is a Phase II randomized, double-blinded, placebo-controlled trial that evaluates the safety and efficacy of stromal cell-derived factor 1 (SDF-1) encoding plasmid for the treatment of ischemic 2.2
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HF [22]. Gene transfers are done as endomyocardial injections using a Helical Infusion Catheter (BioCardia) with the SDF-1 plasmid (low dose 15 mg, high dose 30 mg) or placebo [22]. The primary outcome measures of the trial include 6-min walking distance and the quality of life (Minnesota Living with Heart Failure Questionnaire) 4 months after the treatment. Also, left ventricular ejection fraction (LVEF) using echocardiography, New York Heart Association (NYHA) functional classification, major adverse cardiac events (MACE) and other end points will be measured. The study is expected to enroll 90 patients [22]. CUPID-2b is a Phase II, randomized, double-blinded placebo-controlled trial in which HF patients are treated using adeno-associated virus serotype 1 (AAV1)-mediated sarcoplasmic reticulum calcium ATPase 2a (SERCA2a) or placebo [23]. The AAV1-SERCA2a gene transfer is used to normalize deficient sarcoplasmic reticulum calcium uptake and intracellular calcium cycling in the cardiomyocytes of a failing heart to improve left ventricular function [24]. CUPID-2b is a larger confirmatory trial of the previous CUPID-2 trial, which consisted of only 39 patients and where AAV1-SERCA2a was given at three different doses of 6 1011, 3 1012 or 1 1013 DNase-resistant particles (DRP) in addition to the placebo group [24]. Despite its low statistical power, CUPID-2 still suggested that the highest dose of AAV1-SERCA2a gene therapy could achieve clinically significant improvements in patients with advanced HF [24]. CUPID-2b enrolls patients with chronic systolic HF or non-ischemic cardiomyopathy, and a total enrollment of 200 is expected. Participants are screened for neutralizing AAV1 antibodies to exclude patients with immunity to the vector. Gene therapy is administered as a one-time intracoronary infusion of 1 1013 DRP (corresponding to the highest dose used in CUPID-2) of AAV1-SERCA2a or placebo. Primary outcome measures include HF-related hospitalizations that are related to terminal events, such as heart transplantation, left ventricular assist device implantation or all-cause mortality. AGENT-HF (AAV1-CMV-SERCA2a Gene Therapy trial in Heart Failure) is a Phase II, randomized, double-blinded, placebo-controlled trial with a similar treatment protocol to the CUPID-2b trial [23,25] and takes place in France. Intracoronary AAV1-SERCA2a gene therapy is used to treat congestive HF, and a total of 44 patients with severe ischemic or non-ischemic HF (NYHA III/IV) will be enrolled. Patients are also screened for neutralizing AAV1 antibodies. The primary end point in the trials is left ventricle end-diastolic volume measured using computer tomography. Time to terminal events (heart transplantation, left ventricular assist device implantation or all-cause mortality), maximal oxygen consumption, left ventricle function measured with echocardiography and other secondary outcome measures are also recorded. AC6 Gene Transfer for CHF is a Phase I/II, randomized, double-blinded, placebo-controlled trial where gene therapy with adenoviral adenylyl cyclase 6 (AdAC6) is used to treat
Expert Opin. Biol. Ther. (2014) 14(3)
EudraCT FIN 2012-001019-22
KAT-PAD 101
Expert Opin. Biol. Ther. (2014) 14(3) I/II
I/II
II
II
II
I
Vector
Plasmid
Plasmid
Plasmid
VEGF-DdNdC Ad
MGA: Ang-1, RV VEGF-A165
HGF-X7 (NL003)
HGF-X7 (VM202)
SDF-1
MGA: Ang-1, RV VEGF-A165
Gene
No control
Control
Saline
MGA low therapeutic dose, MGA high therapeutic dose 3 1010 vp, 3 1011 vp
28
30
2 -- 4 IM injections Saline
200
50
48
12
IA infusion
96 IM injections
64 IM injections
IM injection
IA infusion
Administration Enrollment
Two treatment groups
4, 6 or 8 mg Saline once at 3 time points
8 or 16 mg
4, 8 or 16 mg Placebo
Escalating doses
Dose
Safety, clinical status, vital signs, serum VEGF-D levels, AEs
AEs
Ulcer area, VAS
VAS
AEs, SAEs, laboratory assessments
AEs
Primary end points
Pain and ulcer assessment, ABI, clinical laboratory testing, medical therapy, symptoms
VAS, TcPO2, ABI and TBI, wound healing, VAS, QOL, amputation rate, mortality TcPO2, VAS, ulcer area, ABI and TBI, QOL, complete ulcer healing, ulcer healing after gangrene treatment, major amputation rate Critical limb ischemia symptoms
Major and minor amputations, survival, QOL, ulcer healing, pressure measurements
PAD symptoms
Secondary end points
MRA, MRI, contrastenhanced ultrasound, Doppler ultrasound
Perfusion with MRA
Secondary end point (imaging)
ABI: Ankle-brachial index; AE: Adverse effect; Ad: Adenovirus; CT.gov: ClinicalTrials.gov; ECHO: Echocardiography; IA: Intra-arterial; IM: Intramuscular; LVEF: Left ventricular ejection fraction; LVAD: Left ventricular assisting devise; MRA: Magnetic resonance angiography; MRI: Magnetic resonance imaging; PAD: Peripheral artery disease; QOL: Quality of life; RV: Retrovirus; SAE: Serious adverse effect; SPECT: Single-photon emission computed tomography; TBI: Tibial-brachia index; TcPO2: Transcutaneous oxygen tension; VAS: Visual analog scale; vp: Viral particle.
ISR
CT.gov NCT00956332
Safety Study of MultiGeneAngio in Patients with Chronic Critical Limb Ischemia
CHN
CT.gov NCT01548378
Safety and Efficacy Study Using Gene Therapy for Critical Limb Ischemia
USA, KOR
USA, IND
CT.gov NCT01410331
CT.gov NCT01064440
USA
CT.gov NCT00390767
Safety Study of MultiGeneAngio in Patients With Peripheral Arterial Disease Study to Evaluate the Safety and Efficacy of JVS-100 Administered to Adults with Critical Limb Ischemia Safety and Efficacy Study Using Gene Therapy for Critical Limb Ischemia
Country Phase
Identifier
Abbreviation or name
Table 2. Ongoing clinical trials for PAD.
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P. J. Halonen et al.
patients with congestive HF [26]. AdAC6 gene transfer is used to improve left ventricular contractility. This study will enroll a total of 56 patients with compensated congestive HF. Intracoronary delivery of AdAC6 or placebo is performed on escalating doses from 3.2 109 to 3.2 1011 vp at five different doses [26]. The primary end points in the trial include ETT, left ventricular function with echocardiography and the rate of left ventricular pressure development and decline, both during rest and dobutamine-induced stress. Congestive heart failure (CHF) symptoms, hemodynamics and implantable cardioverter defibrillator discharge frequency are collected as the secondary outcome measures. Gene therapy trials for peripheral ischemia There are currently six gene therapy trials in the clinic for PAD and peripheral ischemia (Table 2). 2.3
Human hepatocyte growth factor (HGF-X7) There are two clinical trials where HGF is used to treat critical limb ischemia. Both trials use the HGF-X7 plasmid (also known as VM202 or NL003 depending on the country where the trial takes place) that expresses two HGF isoforms, HGF-728 and HGF-723 [27,28]. The same plasmid is also used in another trial for the treatment of chronic stable angina [21]. The first trial takes place in the USA and Korea and is a Phase II, randomized, double-blinded, placebo-controlled study in which the HGF-X7 plasmid is used to treat critical limb ischemia [27]. The study enrolls patients with limb ischemia (Rutherford category 4 or 5) who are poor candidates for bypass grafting or percutaneous angioplasty with a total enrollment of 50 patients. Patients are randomized into three groups: low dose (8 mg HGF-X7), high dose (16 mg HGFX7) and placebo (saline). The plasmids are given as intramuscular injections at multiple time points, 4 mg per time point. In the high dose group, HGF-X7 is given at time points days 0, 14, 28 and 42, whereas in the low dose group HGF-X7 is given only at time points days 0 and 14 with saline given at time points days 28 and 42. In the placebo group, saline is given at all aforementioned time points. At all of the time points, the treatment is administered using 16 intramuscular injections. The primary end point of the trial is the change in pain level (visual analog scale [VAS]) from baseline after a 9-month follow up. Also, mortality, amputation rate, tissue oxygenation (TcPO2) and other secondary outcome measures will be recorded. The other clinical trial using HGF-X7 plasmid is a Phase II, randomized, placebo-controlled study taking place in China. This study enrolls a total of 200 patients with Rutherford 4- or 5-grade limb ischemia. The study consists of four groups: low dose (4 mg), intermediate dose (6 mg) and high dose (8 mg) of HGF-X7 and placebo group. The therapy is administered as 32 intramuscular injections at time points days 0, 14 and 28. The primary end points are the difference in ulcer area from baseline and the difference in pain level 2.3.1
332
using VAS after a 6-month follow up. Other secondary outcome measures, such as difference in TcPO2, amputation rate and ulcer healing will be recorded. SDF-1 The same SDF-1 encoding plasmid that is used in the STOPHF trial is also used in a trial for critical limb ischemia [29]. This trial is a Phase II, randomized, double-blinded, placebo-controlled dose--escalation study evaluating the safety and efficacy of the treatment. The study will enroll 48 patients with Rutherford category 4- or 5-grade limb ischemia and poor candidacy for bypass grafting or percutaneous balloon angioplasty. Patients are assigned into four cohorts. Patients in the first two cohorts receive 4 or 8 mg of the plasmid or placebo as 8 injections, and patients in the other two cohorts receive 8 or 16 mg of the plasmid or placebo as 16 injections. The primary end point of the study is to investigate the safety of the plasmid at different doses. As secondary outcome measures, amputations, survival, quality of life, ulcer healing and different pressure measurements will be assessed. 2.3.2
AdVEGF-D In addition to the KAT301-trial, where AdVEGF-D is used to treat myocardial ischemia, in the Kuopio Angiogenesis Trial in Peripheral Arterial Disease (KAT-PAD 101), AdVEGF-D is used to treat critical lower limb ischemia (personal communications with Dr Petra Korpisalo-Pirinen, EudraCT: 2012-001019-22 [30]). This is a Phase I/II, a singleblinded, placebo-controlled trial. Gene transfers are performed into 2 -- 4 sites as intramuscular injections. The KAT-PAD 101 trial enrolls patients with chronic limb ischemia (according to Rutherford and Fontaine classification) and to whom surgical bypass is considered the most suitable treatment. Gene transfers are performed a day prior to a scheduled bypass operation. The end points include safety, feasibility, quality of life, clinical status, anti-adenoviral antibodies, pain and ulcer assessment, magnetic resonance angiography (MRA), MRI, contrast-enhanced ultrasound, Doppler ultrasound, ankle-brachial index (ABI), adverse events and concomitant medical therapy. 2.3.3
MultiGeneAngio In addition to trials where gene therapy is performed using in vivo gene transfer, there are also two cell therapy trials where cells isolated from the patient are transduced ex-vivo and then reintroduced to the patient. MultiGeneAngio is a cell therapy product that is composed of a cell suspension of endothelial and smooth muscle cells that are isolated from a vein segment of the patient. After the isolation, modification by angiogenic gene transfer and ex-vivo expansion, the cell suspension is reintroduced back to the patient as an intra-arterial injection at the site of the blockage. There are currently two clinical trials investigating the use of MultiGeneAngio in treatment of PAD -- one in the USA [31] and another in Israel [32]. The first one is a Phase I, 2.3.4
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Current gene therapy trials for vascular diseases
single group, open-label safety study, where patients receive escalating doses of MultiGeneAngio. The current enrollment is 12 patients and the study does not enroll new patients. The study enrolled patients with exercise-limiting intermittent claudication and PAD symptoms in one or both legs. The patients are monitored up to 15 years after the treatment for adverse effects. As a secondary outcome, PAD symptoms will also be monitored. The Israeli trial is a Phase I/II, randomized, open-label study, with two dose levels, where patients are given MultiGeneAngio in low and intermediate doses. The current enrollment is 28 patients and this study is not recruiting patients. This study enrolled patients with ischemic rest pain (Rutherford 4) or nonhealing wounds (Rutherford 5).
Conclusion There are many common denominators that can be seen in the current clinical trials for vascular diseases. Myocardial and peripheral ischemia are both conditions that result from the same cause: the lack of proper blood flow into the tissue. This is also reflected in the current clinical trials, as multiple gene therapy products are under investigation for the treatment of both myocardial and peripheral ischemia. The HGF-X7 plasmid encoding for HGF and AdVEGF-D treatment are studied in the settings of both myocardial and critical limb ischemia. The SDF-1 plasmid is studied for both the treatment of ischemic HF (STOP-HF) and ischemia caused by PAD. Several trials enroll so-called ‘no-option patients’ who are not suitable for conventional treatments such as bypass grafting or percutaneous balloon angioplasty and stenting. This means that the patients enrolled in the gene therapy trials are elderly and have many concomitant diseases, such as type 2 diabetes. The only exception is the KAT-PAD 101 trial that specifically includes patients to whom surgical bypass is considered the best treatment, as opposed to enrolling patients who are poor candidates for revascularization. Primary outcome measures in most trials measure either safety or hard clinical end points, such as MACE, amputations or mortality. In these kinds of settings, it might be difficult to achieve significant results when using relatively small patient populations that consist mostly of ‘no-option’ patients [4]. In 8 of these 15 trials, either intramyocardial or intramuscular injections are used for the administration of gene therapy as opposed to an intracoronary or intra-arterial infusion. Ten trials use viral vectors instead of plasmid DNA to deliver the therapeutic gene. The concept of therapeutic angiogenesis has persisted as the main strategy for novel therapies to increase perfusion in the ischemic tissues. In gene therapy for non-ischemic vascular diseases, CUPID-2b, AGENT-HF and AC6 Gene Therapy for CHF use a different approach and is aimed at improving the contractility of the failing heart. 2.4
3.
Expert opinion
Inclusion criteria Previous cardiovascular gene therapy trials have mainly demonstrated the safety and feasibility of the therapies. The primary end points of previous Phase I/II studies have mostly concerned safety, with secondary outcome measures used to determine the actual effects of the therapy. ‘No-option’ patients with high comorbidity are enrolled in Phase I/II trials even though it is not clear whether these patients have already lost their regenerative capacity when it comes to treating ischemic conditions with gene therapy [3,4]. It has been proposed that these patients could lack the capacity to respond to angiogenic gene therapies [2,4]. A recent review by Dragneva et al. pointed out that many preclinical studies using rodent models are largely based on young and healthy animals, whereas clinical patients are very sick and suffer from multiple conditions in addition to the vascular diseases [2]. After the safety of the treatments has been confirmed, new trials should move toward treating less severely affected patients. Also, the use of gene therapy as an adjuvant treatment for conventional revascularization therapies as in the KAT-PAD 101 trial should be seriously considered. 3.1
Surrogate end points Forthcoming gene therapy trials will probably enroll relatively small patient populations when compared with large clinical trials with regular pharmaceutical compounds. Previous trials have often focused on major clinical end points, such as MACE, amputation rate and ulcer healing. These end points by themselves are very challenging and not optimal for measuring therapeutic effects with other variables affecting the outcome in limited patient populations of sick and elderly patients. The primary end points of future trials could move from MACE toward validated and more sensitive surrogate end points, enabling smaller clinical trials to provide important information about the effectiveness of the new treatments. Especially the use of functional and metabolic surrogate end points, such as LVEF and PET-perfusion imaging, glucose uptake, viability and fatty acid uptake, should be considered. Also, metabolic analyses and functional molecular imaging would provide more useful and mechanistic data that cannot be acquired by using traditional end points [2]. 3.2
Vectors The vectors used in the clinical trials should have appropriate expression profiles, tissue tropism and transduction efficiencies to enable clinically relevant transgene expression in target tissues. In many previous and current clinical trials, plasmidbased therapies are used, although the capacity of plasmid therapies to cause robust transduction in target tissues is still unclear even using large amounts of plasmid DNA [33]. It should be critically investigated if plasmid based-therapies 3.3
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can cause clinically relevant transduction efficiencies in human patients. Only vectors that have been demonstrated to achieve high enough transduction efficiencies in preclinical studies should be used in clinical studies. If robust transduction is not achieved, negative results might be caused simply by the lack of the expression of a therapeutic gene. Treatment genes Only well-established therapeutic genes should be used in future clinical trials. These would include at least VEGFs, FGFs and HGF for ischemic conditions [3]. In a Phase II study, AdVEGF was shown to induce significant growth of new vasculature in the legs of PAD patients [34]. However, this trial also failed to show any changes in amputation frequency or survival, although it demonstrated excellent longterm safety [8]. Regarding local administration, it should also be assessed whether intracellular or secreted factors will provide the best possible outcome. It could be argued that secreted factors will cover a larger and more uniform area in the target tissue, but on the other hand, for example, intracellular transcription factors might result in more physiologic effects by acting more upstream and regulating the expression of multiple genes and therefore having broader therapeutic effects. However, covering functionally relevant areas of human skeletal muscles or myocardium might prove difficult by using treatment gene coding for intracellular factors.
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3.4
Administration In spite of many successful preclinical studies using either systemic or intravascular gene therapy, the human biodistribution and pharmacokinetics of the vectors should also be taken into account. When treating local ischemia, intravascular delivery will unnecessarily increase systemic distribution of the vector when compared with local injections. The effectiveness of, for example, adenoviral VEGF gene therapy using local delivery in clinically relevant large animal models of myocardial ischemia has been demonstrated in several studies [4,35]. Although there has recently been positive reports using an intracoronary infusion of AAV1-mediated gene transfer for HF in the CUPID trial [24], there is preclinical work suggesting that intramuscular injections might be a more effective way to transduce muscle tissue than intravascular infusion [3]. It should be further investigated whether there are differences in optimal administration routes depending on the vector. The AAV1 vector used in CUPID-2b could perform better 3.5
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than adenoviral vectors if gene therapy is given as an intracoronary infusion. Studies have also suggested that gene delivery methods, such as retrograde injection, coronary sinus blockade and balloon occlusion causing higher perfusion pressure, higher coronary flow and a longer dwelling time for the vector in the vascular tree, result in a better transduction efficiency than a simple anterograde infusion [36]. These methods might prove essential in achieving robust global transduction of the target tissues, whereas focal expression induced by local gene delivery could be more suitable for other approaches, such as angiogenic gene therapy to the infarct border [36]. Also, local gene delivery methods limit the possible side effects, such as the increase in tissue permeability in angiogenic therapies, to the target sites. New developments in tissue specific vectors, such as cardiotropic AAV vectors, and tissuespecific promoters might alter the situation in the future [37]. However, local delivery would seem to be the most logical approach for most cardiovascular conditions at the moment. Ethical considerations Excellent safety and tolerability of cardiovascular gene therapy has been demonstrated in multiple previous clinical trials using mainly ‘no-option’ patients [7,8,15]. Without forgetting the case of Jesse Gelsinger and with the lessons learned from it [12], it seems very unlikely for gene therapy to cause serious adverse effects in future clinical trials. Cardiovascular diseases are the most common cause of death [1] and an enormous public health issue especially in the western world. Novel therapies such as gene therapy are, thus, needed to treat patients with these increasingly ubiquitous diseases in aging populations. 3.6
Expert opinion conclusion Forthcoming clinical trials should begin enrolling less severely affected patients, since the safety of many gene transfer modalities have already been well established. Broader emphasis should also be given on new imaging-based, metabolic and functional surrogate end points in addition to the conventional clinical outcome measures to better capture the effects of novel therapies in relatively small patient populations. 3.7
Declaration of interest The authors have no competing interests to declare. Authors acknowledge financial support from Finnish Academy, Kuopio University Hospital Research Unit, Sigrid Juselius Foundation, European Research Council (ERC) and Finnish Heart Foundation.
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Current gene therapy trials for vascular diseases
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Affiliation Paavo J Halonen1 BMed, Jussi Nurro1 BMed, Antti Kuivanen1 BMed & Seppo Yla¨-Herttuala†1,2 MD PhD FESC † Author for correspondence 1 University of Eastern Finland, A. I. Virtanen Institute, Department of Biotechnology and Molecular Medicine, P.O. Box 1627, FIN-70211, Kuopio, Finland Tel: +358 40 355 2076; E-mail: [email protected] 2 Kuopio University Hospital, Research Unit and Gene Therapy Unit, FIN-70211, Kuopio, Finland
