Inovio Pharmaceuticals, Inc. a leader in the development of therapeutic and preventive vaccines against cancers and infectious diseases, announced that significant T cell and antibody responses were generated in its phase I clinical study of VGX-3400X, a SynCon DNA vaccine for the prevention of avian H5N1 influenza delivered using intramuscular (IM) electroporation.

These results were presented at DNA Vaccines 2011, hosted in San Diego by the International Society of DNA Vaccines, by Dr Niranjan Sardesai, Inovio's Sr. VP, Research and Development. In conjunction with these results, Inovio has launched a second phase I clinical study as part of its universal influenza vaccine program. This trial will assess a multi-subtype SynCon vaccine for H1N1 and H5N1 influenza using its skin-targeted intradermal (ID) electroporator. Inovio's SynCon vaccine design process uses a proprietary method to achieve cross-strain protection against the natural and frequent mutations of influenza strains within subtypes.

Dr J Joseph Kim, Inovio's president and CEO, said: “We are encouraged by the immune responses generated in this proof-of-principle study of our first SynCon influenza vaccine, VGX-3400X, delivered using intramuscular electroporation. The second phase I study (INO-3510) builds on our universal influenza vaccine development programme by adding a second component targeting the H1N1 subtype and delivering vaccine formulations using our minimally invasive intradermal electroporation delivery system, which is designed to directly access the skin tissue that is most ideal for inducing preventive antibody responses. Inovio's ultimate goal is to develop a broadly cross-protective influenza vaccine simultaneously targeting multiple flu sub-types and unmatched strains within subtypes. We are approaching this goal in a modular manner and we look forward to generating data from this study using a multi-antigen vaccine, which we expect in the first quarter of 2012.”

Inovio's VGX-3400X targets the pandemic influenza subtype H5N1. The vaccine consists of three distinct DNA plasmids coded for a consensus hemagglutinin (HA) antigen derived from different H5N1 virus strains; a consensus neuraminidase (NA) antigen derived from different N1 sequences; and a consensus nucleoprotein (NP) fused to a small portion of the m2 protein (m2E) based on a broader cross-section of influenza viruses in addition to H5N1 and H1N1.

In this first proof of principle study of Inovio's universal flu vaccine program, VGX-3400X was delivered with IM electroporation using Inovio's Cellectra electroporation device. The primary objectives of this clinical trial were to assess safety and tolerability. The secondary objective was the measurement of antigen-specific T cell and antibody responses, including binding and hemagglutination inhibition (HI) responses.

The study assessed a total of 60 healthy volunteers, 30 in the US and 30 in Korea (in a separate, parallel clinical trial sponsored by Inovio affiliate VGX International). Three dose cohorts of 10 subjects were each given two injections of 0.2 mg, 0.67 mg, or 2.0 mg of each plasmid at months 0 and 1.

In the study, VGX-3400X was found to be generally safe and well tolerated at all dose levels. There were no vaccine-related serious adverse events. Reported adverse events and injection site reactions were mild to moderate and required no treatment.

Inovio tested for antibody responses against the target antigens and observed high levels of binding antibodies in 26 of 27 evaluated subjects (96%). Antibodies were generated against all three antigens, as tested by the enzyme-linked immunosorbent assay (ELISA). Positive antibody responses persisted to seven months, the latest time point tested.

In testing for HI responses against the Vietnam (A/H5N1/1203/04) strain, 3 of 27 subjects (11%) showed HI titers greater than 1:40, which is considered to be an indicator of protection against influenza in humans. Two of the three subjects with HI titers exceeding 1:40 against the Vietnam strain also demonstrated greater than 1:40 titers against the Indonesia (A/H5N1/5/2005) strain, demonstrating cross-reactive responses in these volunteers.

Significantly, antigen-specific Cytotoxic T-Lymphocyte (CTL) responses were also observed against all three antigens (HA, NA and NP). After two vaccinations, 13 of 18 vaccinated subjects (72%) from the first two cohorts developed strong CTL responses to at least one of the vaccine components. Cohort 3 samples have not yet been analysed. These positive T cell responses were measured up to seven months after the first vaccination.

Generation of influenza antigen-specific T cell responses is believed to be important for generating universal, long-lasting immunity against influenza as well as to generate a stronger immune response against flu in elderly people. Inovio previously reported the generation of strong, long-lasting and best-in-class T-cell responses in phase I studies of both VGX-3100 for cervical cancer/dysplasia and Pennvax-B for HIV via IM electroporation delivery of Inovio SynCon DNA vaccines.

Speaking at the DNA Vaccines Meeting, Dr Sardesai commented, “The high rates of vaccine-induced antibody and T-cell responses elicited by VGX-3400X delivered via IM electroporation are very promising for a DNA vaccine for flu. In our next phase I study, for INO-3510, we will be looking at further optimized vaccine formulation (monovalent versus multivalent), vaccine regimens, and route of delivery. This new study will introduce the first use of skin-targeted delivery to support prophylactic vaccine applications.”

With the encouraging proof of principle immune response data from VGX-3400X, Inovio has since initiated a booster study in which US study participants were vaccinated with a booster dose of 0.9 mg of just the H5 HA SynCon DNA vaccine component delivered with Inovio's new intradermal electroporation delivery technology.

This is the first ever flu study of ID electroporation delivery in humans. ID electroporation is focused on delivering a DNA vaccine into skin, which contains large amounts of immune cells such as dendritic cells and macrophages. Inovio's skin electroporation device includes a patented design that introduces a miniaturized array, which delivers the electrical pulses into the skin, and uniquely optimized electroporation conditions. A recent controlled study conducted in human volunteers showed that tolerability of this device was similar to conventional syringe-delivered vaccines. Inovio believes these important design outcomes could make such devices suitable for widely applicable preventive vaccine regimens against infectious diseases such as influenza, dengue and malaria.

Inovio's new minimally-invasive, skin-targeted ID electroporation technology has been used to deliver DNA vaccines in several preclinical animal models. In previously-published preclinical studies for flu and smallpox in monkeys, ID electroporation delivery of SynCon DNA vaccines generated greater neutralizing antibody titers than IM electroporation and protected monkeys in influenza and smallpox challenge models.

The goal of the booster study is to determine if ID vaccination with the monovalent H5HA construct can increase HI titers. Of the 27 available subjects, 18 have already received the ID booster vaccination. Safety and immune response data are expected in 4Q 2011.

In conjunction with completing the interim assessment of Inovio's first human influenza study with its single subtype SynCon vaccine, the company is advancing its universal influenza vaccine strategy with the initiation of a second phase I influenza vaccine study. This trial is assessing INO-3510, Inovio's multi-subtype SynCon DNA vaccine consisting of the H5N1 and H1N1 subtypes delivered exclusively with its new ID electroporator. This phase I for INO-3510 introduces three strategic components for its universal influenza program, which is funded in part by a recent grant from the National Institutes of Health. First, this vaccine adds the H1N1 seasonal/pandemic influenza subtype to the H5N1 pandemic subtype. Second, it will be Inovio's first clinical study solely using ID electroporation delivery. Third, this study will test the SynCon vaccine's ability to provide cross-strain protection against H1N1 and H5N1 viruses.

In this study, 90 subjects will receive two or three doses of the DNA vaccines over 1 to 6 months. The Investigational New Drug submission for this study has been approved by the FDA and all 90 subjects are currently being screened and randomized. Preliminary safety and immune response data are expected in 1Q 2012.

Of three broad types of influenza (A, B, C), influenza A is the most deadly and the causative agent of pandemics. Two of the eight proteins associated with influenza A are HA (hemagglutinin) and NA (neuraminidase). H1, H2, and H3 are the HAs and N1 and N2 are the NAs that have caused the majority of disease in man, with the specific combinations of concern to man being H1N1, H2N2, and H3N2. H5N1 is pathogenic and quite lethal but has not been shown to transmit from person to person.

The challenge to the development of effective vaccines against these influenza subtypes is that, within subtypes, there are hundreds or thousands of strains that may vary slightly and which naturally and frequently mutate to create new strains. Today's vaccines only provide protection by matching the virus strain of concern, and are often unable to provide protection because they do not match the strain that becomes prolific during the next flu season. Moreover, subtypes can "reassort" - for example, a current concern is that a strain of H5N1, which is very deadly but does not currently pass from person to person, may combine with a strain of H1N1 or other subtype that does pass from person to person.

Inovio's novel approach is to synthetically create consensus gene sequences derived from the HA antigens of multiple existing virus strains within a targeted influenza subtype. This results in patentable DNA vaccine constructs encoded to produce antigens that do not match any one existing strain but have been shown in multiple animal studies to protect against unmatched known and newly emergent strains (e.g. the 2009 swine-origin H1N1 virus). While providing cross-strain protection within each subtype, creating and combining multiple DNA vaccine constructs for different subtypes expands Inovio's vision for a truly universal influenza vaccine capable of providing broad protection against yearly seasonal influenza and the less frequent but inevitable virulent strains that rise to pandemic status.

In addition to creating SynCon vaccines based on HA and NA antigens, which are the proteins that evolve most frequently, Inovio has also created SynCon constructs for the NP and m2E proteins, which mutate much less frequently and are more conserved across different strains, adding additional antigenic targets to broaden the potential protective capability of its influenza vaccines.

Inovio has established a modular development plan for systematically combining vaccine components targeting the influenza A H1N1, H5N1, H3N2, H2N2 subtypes as well as influenza B with the goal of developing a universal influenza vaccine.

Inovio is developing a new generation of vaccines, called DNA vaccines, to treat and prevent cancers and infectious diseases. Its SynCon vaccines are designed to provide broad cross-strain protection against known as well as newly emergent strains of pathogens such as influenza.