White Paper on Gene Therapies

Gene Therapies for Diseases Other Than Cancers Vying for Commercial Success

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A gene therapy product has yet to be approved by the US FDA and despite an approval in the EU, Glybera has not yet launched.

Apart from clinical development and regulatory approval, several challenges need to be overcome for gene therapies to become commercially viable.

Over 2,000 gene therapy clinical studies have been

conducted across diverse therapeutic areas, with

80% of these studies conducted in various cancers.

Despite heroic efforts by basic researchers and clinical

investigators alike, no gene therapies have become

commercially available in the US or EU. The lack of

success of gene therapies to date highlights multiple

limitations still to be overcome, e.g. patient selection

TABLE 1: APPROVED GENE THERAPY PRODUCTS

Gene therapies for diseases other than cancers are

progressing through late stages of development,

indicating that some of these therapies are likely to

become available in the near future. Herein we have

discussed the characteristics of the diseases that

Gene Therapy

Gene/MoA Vector Disease/Indication Country of Approval

Year of Approval

Glybera Lipoprotein lipase (LPL)

Adeno Associated Virus

Familial LPLD (LPL deficiency) EU/EMA 2012

Oncorine Adenovirus V5 with E1B and E3B deleted

Adenovirus Head and neck squamous cell carcinoma, lung cancer, liver cancer, malignant pleural and peritoneal effusion, pancreatic cancer

China 2005

Rexin-G Cyclin G1 (cytocidal mutant)

Moloney murine leukemia virus Retrovirus

Solid tumors refractory to standard chemotherapy

Philippines 2007

Gendicine Wt p53 Adenovirus Head and neck squamous cell carcinoma

China 2003

for clinical trials, safer and more effective vectors, and

cost. The first gene therapy product approved in the

EU, Glybera, was for a rare genetic disease caused by a

mutation of the pancreatic enzyme lipoprotein lipase

that renders patients unable to break down fatty acids

effectively. The three other gene therapy products

now approved (Table 1) are for the treatment of

different cancers.

make them suitable for gene therapies, characteristics

of vectors for such modalities, changing industry

perceptions, and the possible future outlook for

these therapies.

Although 80% of global gene therapy clinical studies have been conducted for different cancers, the first and only approved gene therapy in the Western world – Glybera (alipogene tiparvovec) – was approved in the EU for a non-cancer monogenic condition (lipoprotein lipase (LPL) deficiency). Glybera, after several rounds of assessments by the Committee on Advanced Therapies (CAT) was authorized in 2012 under “exceptional circumstances.” Due to the small size of clinical trials supporting Glybera’s authorization, the EMA has required its maker to submit additional clinical data annually for 6 years and to establish a registry for long-term patient tracking. Despite approval, Glybera has yet to be marketed in EU countries due to pending pricing negotiations and other regulatory requirements - even though it made headlines in 2014 for a target €1.1M price tag in Germany.

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Disease-specific characteristics make certain

monogenic disorders tractable for gene-replacement

strategies

The original goal of gene therapies was to treat

monogenic diseases by replacing a non-functional

or defective gene with a healthy copy. Monogenic

hematological disorders such as hemophilias are

particularly amenable to gene therapy because even a

limited increase in expression of the defective gene is

sufficient to offer a phenotypic cure. This notion has

been strengthened with results from recent trials for

hemophilia B. A single intravenous infusion of an adeno-

associated virus serotype 8 (AAV-8) vector expressing

Factor IX in 10 patients with severe hemophilia B

resulted in a dose-dependent increase in its circulating

levels to 1-6% of the normal value over a median period

of 3.2 years. In patients who received the high dose

(2×1012 vector genomes per kilogram of body weight),

there was a consistent increase in Factor IX level to over

5%. Despite such low levels of circulating factor, there

was over a 90% reduction in bleeding episodes and the

use of prophylactic Factor IX concentrate. Hemophilias

also tend to be suitable for gene therapy since minimal

logistic issues need to be addressed with respect to

intravenous administration of viral vectors encoding

clotting factors. Production of such vectors is now

routine, and there are very few manipulations of the

product prior to administration.

Other monogenic disorders such as childhood cerebral

adrenoleukodystrophy (CCALD), while certainly tractable

vis-à-vis gene replacement, present additional logistic

issues. This disease is caused by deficiency in ALD

protein – an adenosine triphosphate-binding cassette

transporter encoded by the ABCD1 gene – which can

be corrected by transducing autologous CD34+ cells ex

vivo with a lentiviral vector encoding wild-type gene and

then re-infusing transduced cells into the patients after

they have received myeloablative treatment. These

additional steps are likely to make it expensive and

restricted to the most advanced medical centers.

Polygenic disorders can be amenable to gene therapy

strategies if specific targets are identified

As many disorders such as diabetes, coronary artery

disease, and rheumatoid arthritis are polygenic,

replacement of a single gene is unlikely to cure the

disease. However, if a specific therapeutic target

(common disease pathway) can be identified, then

delivery of a gene product that expresses sufficient

levels of the protein, in theory, can sufficiently

ameliorate symptoms of a polygenic disease and be

a useful therapy. A case in point is Generx® a Phase

III adenoviral FGF-4 gene therapy for patients with

myocardial ischemia due to coronary artery disease,

developed by Cardium Therapeutics. FGF-4 has been

shown to stimulate the growth of microvascular

circulation in the heart, enhancing cardiac perfusion.

Generx®, is designed to be administered only once, by

an interventional cardiologist. The company recently

reported that the benefits from this gene therapy are

similar in magnitude to the large vessel revascularization

procedures such as bypass surgery or angioplasty.

Tissue tropism and viral integration are important

considerations determining the choice of vector

While there has been significant progress in vector

development in the past two decades, it is also clear

that no vectors can be universally utilized for all gene

therapy applications. There are a few important

considerations in this regard. First, the tropism of

vectors used will determine whether the gene product

will be expressed in the tissue of interest. For instance,

in the case of AAV vectors, as compared to other

serotypes, AAV1 is suitable for expression in skeletal

muscle and retina, AAV5 transduces neuronal and lung

cells efficiently and AAV8 demonstrates high levels

of expression in liver cells. A second consideration is

whether the viral vector employed stably integrates into

the host genome or remains episomally in the nucleus.

Lentivirus vectors have the ability to stably integrate

into target cells, thus providing genetic modification

of the cell and all of its progeny. However, insertional

mutagenesis is a potential issue for vectors that

integrate foreign DNA into the genome. On the other

hand, in case of vectors based on the Herpes Simplex

Virus (HSV), the latent HSV genome is maintained as an

episome, and the potential risks of viral integration into

host cell chromatin are avoided.

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Immune responses need not be a barrier to their effective use in gene transfer While AAV and adenovirus-based vectors are popular gene therapy vectors, the human immune response to them might be considered problematic. AAV vectors often induce immune responses against the capsid or the transgene, especially when used in large doses required for clinical benefit, leading to a decrease in transgene expression. In addition to AAV, the efficacy of in vivo gene delivery with adenoviral vectors might be expected to be severely compromised owing to both innate and acquired immune responses as most immunocompetent people have been exposed to adenovirus. With up to 80% of the human population seropositive for AAV-2 (and vectors based on it), concerns over their immunogenicity are justified.

However, it is now recognized that although gene therapy vectors such as adenoviruses and AAV are highly immunogenic, this need not preclude successful gene expression, even long-term expression. The immune-privileged status of the eye makes AAV-mediated ocular gene transfer with low doses relatively efficient with low risk of immune-related events. This was seen in the trials that used AAV to deliver the RPE65 gene to retinal cells in patients of Leber Congenital Amaurosis (LCA), an inherited disorder that results in severe vision loss. As there are many gene therapies that do not target the eye, diverse strategies have been developed to overcome immunogenicity concerns when targeting other tissues. One broadly applicable strategy has been to transduce cells ex vivo and then transfer them into the body. Proof of principle for such a strategy has been obtained in preclinical models where myoblasts transduced with “gutless” adenoviral vectors expressing full-length dystrophin cDNA have been shown to fuse with mature myofibers ex vivo to successfully deliver the gene. Another strategy has been to use virus serotypes such as AAV-5 and AAV-8 which have low

sero-prevalence. Several other approaches to generate immunologically inert AAV vectors have been employed – including targeted mutagenesis, capsid shuffling, or directed evolution approaches – to alter the epitopes on the AAV capsid surface. Chemical modifications of immunogenic sites by strategies such as PEGylation or co-administration of immunosuppressive agents or plasmapharesis to reduce neutralizing antibodies have also been tried. All these approaches, while useful in obtaining a vector with immune-escape potential, are nevertheless fraught with potential problems. These include loss of infectivity, packaging ability, or tissue tropism. While not insurmountable, the obvious disadvantage of all these strategies is additional cost, as cell culture and lab-based engineering necessitates several additional manufacturing and quality control issues.

Large-scale industrialization of vector production makes “low cost of goods” possible for certain gene therapy modalitiesLimited economical access to large quantities of GMP grade vector product is another factor that has hampered progress in this arena. Table 2 lists the most commonly used vectors currently in gene therapy modalities along with their packaging capacity, immunogenicity, tissue tropism, and other features. In most cases, transient transfection into a human producer line was found to be an effective vector manufacturing strategy. With recent innovations, industrial production of both AAV and lentiviral vector platforms has now reached such a stage of maturity that robust scalable manufacturing processes that generate highly reproducible products are now a reality. In the case of gene therapy products (e.g. Glybera) that use AAV, this was facilitated by the introduction of baculovirus vectors, in combination with Sf9 insect cells, which are easily grown in animal-derived component-free media, and are amenable to further processing at any scale.

Parti

cle

Char

acte

risti

cs Adenovirus Adeno-associated virus Herpes virus Retrovirus/ Pox/vaccinia virus

Genome dsDNA ssDNA dsDNA ssRNA (+) dsDNA

Coat Naked Naked Enveloped Enveloped Enveloped

Genome size1 30-38 kb 5 kb 120-200 kb 3-9 kb 130-280 kb

GT

Prop

erti

es

Infection/tropism Dividing and non-dividing cells

Dividing and non-dividing cells

Dividing and non-dividing cells

Dividing cells2 Dividing and non-dividing cells

Host genome interaction Non-integrating Non-integrating3 Non-integrating Integrating Non-integrating

Transgene expression Transient Potential long lasting Potential long lasting

Long lasting Transient

Packaging capacity 7.5 kb 4.5 kb >30 kb 8 kb 25 kb

Immunogenicity High High High Low High

Other comments World’s first approved gene therapy ‘Gendicine’

EU’s first approved gene therapy ‘Glybera’

Toxicity related to lytic infection

X-linked SCID study where leukemia was reported

Potent immune response suitable for live recombinant vaccines

TABLE 2: COMMONLY USED VECTORS IN GENE THERAPY MODALITIES

1 A large genome can be engineered for insertion and simultaneous expression of multiple genes. 2 Lentiviruses also infect non-dividing cells. 3 Adeno-associated viruses are able to integrate with low frequency into chromosome 19.

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Company Gene Therapy/ Disease Status

1 Uniqure hF-IX gene/ Hemophilia B Collaborator: Chiesi Farmaceutici (licensed from St. Jude Children’s Research Hospital ), Phase I/II

NaGlu gene/ San Fillipo B Syndrome Collaborator: Institute Pasteur, Phase I/II

2 Oxford Biomedica StarGen™ (Sanofi) / Stargardt Disease Phase I/IIa trial ongoing

UshStat® (Sanofi) / Usher Syndrome Type 1B Phase I/IIa

EncorStat® / Corneal Graft Rejection Phase I/II trial preparation

OXB-102 / ProSavin® / Parkinson’s Disease Phase I/II trial completed

Retinostat®/ Wet AMD Phase 1 trial ongoing

3 Bluebird Bio Lenti-D/ Childhood Cerebral ALD Phase II/III global study initiated

LentiGlobin/ Beta Thalassemia, SCD Beta thalassemia: Phase I/ II study initiated SCD: Phase I in US

4 Sangamo SB-728/ HIV/AIDS Phase II

CERE-110/ Alzheimer’s Disease Phase II

5 AGTC AATD / Alpha-1 Antitrypsin Deficiency Phase IIb

RS1 gene/ X-linked Juvenile Retinoschisis (XLRS) IND filed; Phase I/II expected to start 2Q2015

6 Spark Therapeutics

SPK-RPE65/ Inherited Retinal Dystrophies due to RPE65 Gene Mutations Phase III

SPK-CHM / Choroideremia Phase I/II

SPK-FIX / Hemophilia B Collaborator: PfizerPhase I/II expected 1H2015

7 AnGes MG Collategene™ Licensed from Vical (Also Known as Beperminogene Perplasmid, AMG0001)/ Critical Limb Ischemia

Phase III

AMG0001/Primary Lymphedema Phase I/II (Japan)

AMG0001/ Ischemic Heart Disease Phase I completed (US)

8 Taxus Cardium Generx® (Alferminogene Tadenovec) [Ad5FGF-4]/Cardiac Microvascular Insufficiency (CMI) in patients with Myocardial Ischemia and Symptomatic Chronic Stable Angina Pectoris

Phase III

9 Genethon WAS ( Wiskott–Aldrich Syndrome) Phase I/II

X-Linked CGD Patients Phase I/II

10 GSK GSK2696273/ ADA Gene Transfer into Hematopoietic Stem/Progenitor Cells for the Treatment of ADA-SCID

Phase II

11 Viromed Critical Limb Ischemia Phase II completed

Chronic Granulomatous Disease Phase I/II (Korea)

Painful Diabetic Neuropathy Phase II

Chronic Stable Angina Phase I/ II

12 Voyager Therapeutics

VY-AADC01/ Parkinson’s Disease Collaborators: UCSF and Genzyme Phase I

13 Avalanche Biotechnologies

AVA101/ Wet AMD Phase IIa

14 Celladon MYDICAR® (SERCA 2A) / Systolic HF Phase II/III

MYDICAR® ( (SERCA 2A) /Advanced HF with LVAD Phase I/II

TABLE 3: KEY PLAYERS WITH GENE THERAPIES IN CLINICAL DEVELOPMENT

Changing industry perception on risks and benefits of gene therapies The pharmaceutical industry is generally risk averse and understandably so. In 1999, the fatal systemic inflammatory response syndrome generated in an 18-year-old patient with partial ornithine transcarbamylase deficiency – following transfer of a vector based on human adenovirus type 5 – and the tough regulatory climate that ensued, were cause for concern. For many years, this led to a decline in interest in furthering such therapies, especially for non-lethal diseases where such risks were unacceptable. Despite these setbacks, consistent data from various trials showing sustained clinical benefits has now led to a re-evaluation by industry experts, and an improved investor climate. Major pharmaceutical companies

have demonstrated renewed interest and have struck numerous deals with academic groups and smaller biotech companies.

In December 2014, Pfizer inked a deal with a privately owned US biotech firm Spark Therapeutics to develop a gene therapy for hemophilia B. Among other major pharmaceutical companies, Bayer AG has struck a gene therapy deal with Dimension Therapeutics, while Novartis AG recently established a new cell and gene therapies unit, and Sanofi has a long-standing tie-up with Oxford BioMedica. Genzyme recently announced a major strategic collaboration with Voyager Therapeutics for gene therapies for CNS disorders. Perceptions are also changing about accepting the risks associated with gene delivery for the potential benefits, as evident from the large number of clinical studies in these indications.

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Future Outlook Although the first gene therapy clinical trial began in 1990, progress has been slow. Recent developments, including the approval of Glybera and the increasing level of clinical trial activity, have indicated an upswing and a renewed interest in the role of gene therapy in diseases with high unmet need. For instance, in many hematological or metabolic monogenic disorders, where allogeneic hematopoeitic stem cell transplantation (HSCT) from donors has been successfully used to treat disease, gene therapy may correct the genetic defect and permit autologous HSCT. This approach will overcome the challenges associated with arranging a compatible donor and the consequences of an unmatched or unrelated transplantation as in case of graft-versus-host disease.

There is also an increased focus on the issues that would be relevant to maximizing commercial potential, including reimbursement, regulatory hurdles, manufacturing costs, and postmarketing surveillance. The approval of Glybera, while certainly encouraging, is a small step in an evolving field with enormous potential. Glybera’s approval in the EU was accompanied by requirements for a post-approval study, implementation of a disease registry, and a risk-management procedure. All of this can be costly and time-consuming, thereby hindering initial uptake.

Commercial success will depend on the ability of companies to develop gene therapy products with convincing clinical data to support the value proposition of potential long-term benefit or cure without any serious safety concern. They will also face reimbursement challenges as gene therapies may be a one-time or a short-term treatment, and the actual cost may be incurred upfront, unlike conventional therapies where the cost of treatment is incurred over a long period of time. This may be especially problematic in the US where commercial payers may balk at incurring the upfront cost with no certainty of the individual staying with the insurer to realize the long-term benefit. Further, specialist training, patient awareness, and dedicated infrastructure in the form of centers of excellence (to identify / diagnose patients early, to offer them gene therapy, and to manage them post treatment) may be required.

Overall, despite various challenges, gene therapies have the potential to change the future treatment paradigm or even become the standard-of-care (SoC) therapies for certain diseases. While the gene therapy market will largely depend on the regulatory environment, in view of the emerging encouraging data, there is a high likelihood that gene therapy will become a clinical reality in the near future.

Ophthalmic Diseases1. Lebers Congenital Amaurosis2. Stargardt Disease3. Age Related Macular Degenera�on4. Choroideremia5. Leber's Hereditary Op�c Atrophy

Neurological Disorders1. Parkinson’s Disease2. Spinal Muscle Atrophy3. Alzheimer’s disease4. Diabe�c Neuropathy

Cardiovascular Disorders1. Heart Failure2. Angina/Ischemic Heart Disease3. Cri�cal Limb Ischemia/ Intermi�ent Claudica�on

Musculoskeletal Disorders1. Limb Girdle Muscle Dystrophy 2C/2D2. Becker Muscle Dystrophy3. Duchenne Muscular Dystrophy

Other Diseases1. HIV2. Epidermolysis Bullosa3. Acute Intermi�ent Porphyria

Respiratory Diseases1. Cys�c Fibrosis2. Alpha-1 An�trypsin Deficiency

Metabolic Disorders1. Adrenoleukodystrophy2. Pompe3. Ba�en4. Metachtoma�c Leukodystrophy5. San Filippo A6. Hunter Syndrome

Blood Disorders1. Hemophilia2. ADA-SCID3. X Linked- SCID4. Chronic Granulomatous Disease 5. Wischo� Aldrich Syndrome6. Sickle Cell Anemia7. Beta Thalassemia8. Fanconi Anemia

FIGURE 1: CLINICAL APPLICATIONS OF GENE THERAPY

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