Table of Contents

Volume 8, Issue 11

  November 2012

  Pages 1549 - 1738

Special Issue: DNA Vaccines

Guest Editors:Kenneth E. Ugen and David Weiner

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 Issue Catalogue (MARC XML)

EDITOR'S CORNER

Open Access Article

Letter from the Editor

Ronald Ellis and Eva M. Riedmann
Page 1549
http://dx.doi.org/10.4161/hv.22751
Full Text | PDF




NEWS, POLICY AND PROFILES

Open Access Article

Human Vaccines: News

Eva M. Riedmann
Pages 1550 - 1553
http://dx.doi.org/10.4161/hv.22753
Abstract | Full Text | PDF

High safety marks for Merck’s Gardasil

Cuba tests prostate cancer vaccine

HIV’s weak spot: V2

Unique anti-cancer agent ColoAd1 enters the clinic

Broadly neutralizing antibodies against influenza A and B discovered

Clinical trials initiated: Nexvax2 therapeutic vaccine for celiac disease

The 20 top-selling vaccines in the first half of 2012

Influenza vaccine safe for pregnant women





DNA Vaccines

EDITOR'S CORNER

Open Access Article

Highlights of the DNA Vaccines 2011 Meeting

Kenneth E. Ugen and David B. Weiner
Page 1554
http://dx.doi.org/10.4161/hv.22752
Full Text | PDF

RESEARCH PAPERS

IL-4 and IFN-γ induced by human immunodeficiency virus vaccine in a schistosome infection model

Jiangmei Yin, Anlan Dai, Tatiana Arango, Ravi S. Kasinathan, Robert M. Greenberg and Jean D. Boyer
Pages 1555 - 1563
http://dx.doi.org/10.4161/hv.22142
Abstract | Full Text | PDF

The co-infection of HIV and helminth parasites, such as Schistosoma spp, has increased in sub-Saharan Africa. Many HIV vaccine candidate studies have been completed or are in ongoing clinical trials, but it is not clear how HIV vaccines might affect the course of schistosome infections. In this study, we immunized S. mansoni-infected mice with an efficient DNA vaccine that included HIV gag. Using this model, we found that Th2 cytokines, such as IL-4 and IL-13, were highly induced after schistosome infection. Treatment of infected mice with the HIV DNA vaccine resulted in a significant attenuation of this rise in IL-13 expression and an increase in expression of the Th1 cytokine, TNF-α. However, vaccine administration did not significantly influence the expression of IL-4, or IFN-γ, and did not affect T cell proliferative capacity. Interestingly, the IL-4⁺IFN-γ⁺ phenotype appears in schistosome-infected mice that received HIV vaccination, and is associated with the expression of transcription factors GATA3⁺T-bet⁺ in these mice. These studies indicate that DNA vaccination can have an impact on ongoing chronic infection.

Open Access Article

Clinical trial in healthy malaria-naïve adults to evaluate the safety, tolerability, immunogenicity and efficacy of MuStDO5, a five-gene, sporozoite/hepatic stage Plasmodium falciparum DNA vaccine combined with escalating dose human GM-CSF DNA

Thomas L. Richie and 34 others
Pages 1564 - 1584
http://dx.doi.org/10.4161/hv.22129
Abstract | Full Text | PDF

When introduced in the 1990s, immunization with DNA plasmids was considered potentially revolutionary for vaccine development, particularly for vaccines intended to induce protective CD8 T cell responses against multiple antigens. We conducted, in 1997−1998, the first clinical trial in healthy humans of a DNA vaccine, a single plasmid encoding Plasmodium falciparum circumsporozoite protein (PfCSP), as an initial step toward developing a multi-antigen malaria vaccine targeting the liver stages of the parasite. As the next step, we conducted in 2000–2001 a clinical trial of a five-plasmid mixture called MuStDO5 encoding pre-erythrocytic antigens PfCSP, PfSSP2/TRAP, PfEXP1, PfLSA1 and PfLSA3. Thirty-two, malaria-naïve, adult volunteers were enrolled sequentially into four cohorts receiving a mixture of 500 μg of each plasmid plus escalating doses (0, 20, 100 or 500 μg) of a sixth plasmid encoding human granulocyte macrophage-colony stimulating factor (hGM-CSF). Three doses of each formulation were administered intramuscularly by needle-less jet injection at 0, 4 and 8 weeks, and each cohort had controlled human malaria infection administered by five mosquito bites 18 d later. The vaccine was safe and well-tolerated, inducing moderate antigen-specific, MHC-restricted T cell interferon-γ responses but no antibodies. Although no volunteers were protected, T cell responses were boosted post malaria challenge. This trial demonstrated the MuStDO5 DNA and hGM-CSF plasmids to be safe and modestly immunogenic for T cell responses. It also laid the foundation for priming with DNA plasmids and boosting with recombinant viruses, an approach known for nearly 15 y to enhance the immunogenicity and protective efficacy of DNA vaccines.

REVIEW

Open Access Article

Development of a multicomponent Staphylococcus aureus vaccine designed to counter multiple bacterial virulence factors

Annaliesa S. Anderson, Alita A. Miller, Robert G.K. Donald, Ingrid L. Scully, Jasdeep S. Nanra, David Cooper and Kathrin U. Jansen
Pages 1585 - 1594
http://dx.doi.org/10.4161/hv.21872
Abstract | Full Text | PDF

Staphylococcus aureus is a major cause of healthcare-associated infections and is responsible for a substantial burden of disease in hospitalized patients. Despite increasingly rigorous infection control guidelines, the prevalence and corresponding negative impact of S. aureus infections remain considerable. Difficulties in controlling S. aureus infections as well as the associated treatment costs are exacerbated by increasing rates of resistance to available antibiotics. Despite ongoing efforts over the past 20 years, no licensed S. aureus vaccine is currently available. However, learnings from past clinical failures of vaccine candidates and a better understanding of the immunopathology of S. aureus colonization and infection have aided in the design of new vaccine candidates based on multiple important bacterial pathogenesis mechanisms. This review outlines important considerations in designing a vaccine for the prevention of S. aureus disease in healthcare settings.

RESEARCH PAPERS

Preclinical evaluation of the immunogenicity and safety of plasmid DNA-based prophylactic vaccines for human cytomegalovirus

Jukka Hartikka, Vesselina Bozoukova, Jane Morrow, Denis Rusalov, Mark Shlapobersky, Qun Wei, Sou Boutsaboualoy, Ming Ye, Mary K. Wloch, John Doukas, Sean M. Sullivan, Alain Rolland and Larry R. Smith
Pages 1595 - 1606
http://dx.doi.org/10.4161/hv.21225
Abstract | Full Text | PDF

Human cytomegalovirus (CMV) establishes a lifelong persistent infection characterized by periods of latency and sporadic viral replication and is a major infectious cause of birth defects following congenital infection. Currently, no licensed vaccine is available that would prevent CMV infection. In an effort to develop a prophylactic CMV vaccine, the effects of different formulations, immunization routes and delivery devices on the immunogenicity of plasmid DNA (pDNA)-based vaccines were evaluated in rabbits and mice. Compared with PBS- and poloxamer-based formulations, significantly higher antibody responses were obtained with pDNA formulated with Vaxfectin^®, a cationic lipid-based adjuvant. With low vaccine doses, the intradermal (ID) route resulted in higher antibody responses than obtained when the same dose was administered intramuscularly (IM). Since the IM route allowed injection of larger volumes and higher doses than could be administered at a single ID site, better antibody responses were obtained using the IM route. The needle-free injection system Biojector^® 2000 and electroporation devices enhanced antibody responses only marginally compared with responses obtained with Vaxfectin^®-formulated pDNA injected IM with a needle. A single-vial Vaxfectin^® formulation was developed in a dosage form ready for use after thawing at room temperature. Finally, in a GLP-compliant repeat-dose toxicology study conducted in rabbits, single-vial Vaxfectin^®-formulated vaccines, containing pDNA and Vaxfectin^® up to 4.5 mg and 2 mg/injection, respectively, showed a favorable safety profile and were judged as well-tolerated. The results support further development of a Vaxfectin^®-formulated pDNA vaccine to target congenital CMV infection.

Generation of antigen-specific immunity following systemic immunization with DNA vaccine encoding CCL25 chemokine immunoadjuvant

Noshin Kathuria, Kimberly A. Kraynyak, Diane Carnathan, Michael Betts, David B. Weiner and Michele A. Kutzler
Pages 1607 - 1619
http://dx.doi.org/10.4161/hv.22574
Abstract | Full Text | PDF

A significant hurdle in vaccine development for many infectious pathogens is the ability to generate appropriate immune responses at the portal of entry, namely mucosal sites. The development of vaccine approaches resulting in secretory IgA and mucosal cellular immune responses against target pathogens is of great interest and in general, requires live viral infection at mucosal sites. Using HIV-1 and influenza A antigens as models, we report here that a novel systemically administered DNA vaccination strategy utilizing co-delivery of the specific chemokine molecular adjuvant CCL25 (TECK) can produce antigen-specific immune responses at distal sites including the lung and mesenteric lymph nodes in mice. The targeted vaccines induced infiltration of cognate chemokine receptor, CCR9+/CD11c+ immune cells to the site of immunization. Furthermore, data shows enhanced IFN-λ secretion by antigen-specific CD3+/CD8+ and CD3+/CD4+ T cells, as well as elevated HIV-1-specific IgG and IgA responses in secondary lymphoid organs, peripheral blood, and importantly, at mucosal sites. These studies have significance for the development of vaccines and therapeutic strategies requiring mucosal immune responses and represent the first report of the use of plasmid co-delivery of CCL25 as part of the DNA vaccine strategy to boost systemic and mucosal immune responses following intramuscular injection.

IL-12 DNA as molecular vaccine adjuvant increases the cytotoxic T cell responses and breadth of humoral immune responses in SIV DNA vaccinated macaques

Rashmi Jalah, Vainav Patel, Viraj Kulkarni, Margherita Rosati, Candido Alicea, Brunda Ganneru, Agneta von Gegerfelt, Wensheng Huang, Yongjun Guan, Kate E. Broderick, Niranjan Y. Sardesai, Celia LaBranche, David C. Montefiori, George N. Pavlakis and Barbara K. Felber
Pages 1620 - 1629
http://dx.doi.org/10.4161/hv.21407
Abstract | Full Text | PDF

Intramuscular injection of macaques with an IL-12 expression plasmid (0.1 or 0.4 mg DNA/animal) optimized for high level of expression and delivered using in vivo electroporation, resulted in the detection of systemic IL-12 cytokine in the plasma. Peak levels obtained by day 4–5 post injection were paralleled by a rapid increase of IFN-γ, indicating bioactivity of the IL-12 cytokine. Both plasma IL-12 and IFN-γ levels were reduced to basal levels by day 14, indicating a short presence of elevated levels of the bioactive IL-12. The effect of IL-12 as adjuvant together with an SIVmac239 DNA vaccine was further examined comparing two groups of rhesus macaques vaccinated in the presence or absence of IL-12 DNA. The IL-12 DNA-adjuvanted group developed significantly higher SIV-specific cellular immune responses, including IFN-γ⁺ Granzyme B⁺ T cells, demonstrating increased levels of vaccine-induced T cells with cytotoxic potential, and this difference persisted for 6 mo after the last vaccination. Coinjection of IL-12 DNA led to increases in Gag-specific CD4⁺ and CD4⁺CD8⁺ double-positive memory T cell subsets, whereas the Env-specific increases were mainly mediated by the CD8⁺ and CD4⁺CD8⁺ double-positive memory T cell subsets. The IL-12 DNA-adjuvanted vaccine group developed higher binding antibody titers to Gag and mac251 Env, and showed higher and more durable neutralizing antibodies to heterologous SIVsmE660. Therefore, co-delivery of IL-12 DNA with the SIV DNA vaccine enhanced the magnitude and breadth of immune responses in immunized rhesus macaques, and supports the inclusion of IL-12 DNA as vaccine adjuvant.

DNA prime-protein boost using subtype consensus Env was effective in eliciting neutralizing antibody responses against subtype BC HIV-1 viruses circulating in China

Mingshun Zhang, Lu Zhang, Chunhua Zhang, Kunxue Hong, Yiming Shao, Zuhu Huang, Shixia Wang and Shan Lu
Pages 1630 - 1637
http://dx.doi.org/10.4161/hv.21648
Abstract | Full Text | PDF

Previously, we have shown that DNA prime-protein boost is effective in eliciting neutralizing antibodies (NAb) against randomly selected HIV-1 isolates. Given the genetic diversity of HIV-1 viruses and the unique predominant subtypes in different geographic regions, it is critical to test the DNA prime-protein boost approach against circulating viral isolates in key HIV endemic areas. In the current study, the same DNA prime-protein boost vaccine was used as in previous studies to investigate the induction of NAb responses against HIV-1 clade BC, a major subtype circulating in China. A codon optimized gp120-BC DNA vaccine, based on the consensus envelope (Env) antigen sequence of clade BC, was constructed and a stable CHO cell line expressing the same consensus BC gp120 protein was produced. The immunogenicity of this consensus gp120-BC was examined in New Zealand White rabbits by either DNA prime-protein boost or protein alone vaccination approaches. High levels of Env-specific antibody responses were elicited by both approaches. However, DNA prime-protein boost but not the protein alone immune sera contained significant levels of NAb against pseudotyped viruses expressing HIV-1 BC Env antigens. Furthermore, high frequencies of CD4 binding site-targeted antibodies were found in the DNA prime- protein boost rabbit sera indicating that the positive NAb may be the result of antibodies against conformationally sensitive epitopes on HIV-1 Env. The findings support that DNA prime-protein boost was effective in eliciting NAb against a key HIV-1 virus subtype in China. This result may lead to the development of regional HIV vaccines through this approach.

Pilot study on the immunogenicity of paired Env immunogens from mother-to-child transmitted HIV-1 isolates

Shixia Wang, Michael Kishko, Shengqin Wan, Yan Wang, Frank Brewster, Glenda E. Gray, Avye Violari, John L. Sullivan, Mohan Somasundaran, Katherine Luzuriaga and Shan Lu
Pages 1638 - 1647
http://dx.doi.org/10.4161/hv.22414
Abstract | PDF

Recent studies have reported that founder viruses play unique roles in establishing HIV-1 infection. Understanding the biological and immunological features of envelope glycoproteins (Env) from such viruses may facilitate the development of effective vaccines against HIV-1. In this report, we evaluated the immunogenicity of gp120 immunogens from two pairs of clade B and two pairs of clade C mother-to-child transmitted (MTCT) HIV-1 variants that had various levels of sensitivity to broadly neutralizing monoclonal antibodies. Individual gp120 DNA and protein vaccines were produced from each of the eight MTCT Env antigens included in the current study. Rabbits were immunized with these gp120 immunogens by the DNA prime-protein boost approach. High level Env-specific antibody responses were elicited by all MTCT gp120 immunogens. However, their abilities to elicit neutralizing antibody (NAb) responses differed and those from relatively neutralization-resistant variants tended to be more effective in eliciting broader NAb. Results of this pilot study indicated that not all MTCT Env proteins have the same potential to elicit NAb. Understanding the mechanism(s) behind such variation may provide useful information in formulating the next generation of HIV vaccines.

DNA-based HIV vaccines do not induce generalized activation in mucosal tissue T cells

Morgan A. Reuter, Sally Yuan, Preston A. Marx, Michele A. Kutzler, David B. Weiner and Michael R. Betts
Pages 1648 - 1653
http://dx.doi.org/10.4161/hv.22247
Abstract | Full Text | PDF

HIV preferentially infects activated T cells, and activated mucosal CD4⁺ T cells are the primary sites of viral replication. One potential explanation for increased HIV acquisition rates in the STEP study is that vaccination with adenoviral (Ad) vectors increased CD4⁺ T cell activation levels at the site of infection, a concept that others and we continue to explore.^1,2 Whether vaccination with HIV vaccine platforms increases the activation state of CD4⁺ T cells within peripheral tissues, such as the gastro-intestinal (GI) mucosa, is exceptionally important to determine as a vaccine safety measure, given the susceptibility of activated CD4⁺ T cells to HIV infection.

 

In this study we examined whether vaccination with DNA plasmids and chemokine adjuvants alter the activation state of T cells within the GI mucosa, inguinal LN, and peripheral blood. T cell activation state was measured by expression of CD25, CD69, and HLA-DR over the course of the prime/boost study. DNA plasmid vaccination did not increase expression of any of these markers in the 3 tissues studied. Addition of the gut-homing chemokine TECK during DNA plasmid vaccination did not alter activation levels of CD4⁺ T cells at any of these sites. These findings indicate that DNA vaccines do not elicit generalized mucosal T cell activation. Thus, DNA platforms may be especially suitable for HIV vaccine development, where bystander activation could promote increased HIV transmission.

REVIEW

Open Access Article

Co-expression of HIV-1 virus-like particles and granulocyte-macrophage colony stimulating factor by GEO-D03 DNA vaccine

Michael Hellerstein, Yongxian Xu, Tracie Marino, Shan Lu, Hong Yi, Elizabeth R. Wright and Harriet L. Robinson
Pages 1654 - 1658
http://dx.doi.org/10.4161/hv.21978
Abstract | Full Text | PDF

Here, we report on GEO-D03, a DNA vaccine that co-expresses non-infectious HIV-1 virus-like particles (VLPs) and the human cytokine, granulocyte-macrophage colony-stimulating factor (GM-CSF). The virus-like particles display the native gp160 form of the HIV-1 Envelope glycoprotein (Env) and are designed to elicit antibody against the natural form of Env on virus and virus-infected cells. The DNA-expressed HIV Gag, Pol and Env proteins also have the potential to elicit virus-specific CD4 and CD8 T cells. The purpose of the co-expressed GM-CSF is to target a cytokine that recruits, expands and differentiates macrophages and dendritic cells to the site of VLP expression. The GEO-D03 DNA vaccine is currently entered into human trials as a prime for a recombinant modified vaccinia Ankara (MVA) boost. In preclinical studies in macaques using an SIV prototype vaccine, this vaccination regimen elicited both anti-viral T cells and antibody, and provided 70% protection against acquisition during 12 weekly rectal exposures with a heterologous SIV. Higher avidity of the Env-specific Ab for the native form of the Env in the challenge virus correlated with lower likelihood of SIV infection.

RESEARCH PAPERS

Sequential administration of cytokine genes to enhance cellular immune responses and CD4⁺ T memory cells during DNA vaccination

Baowei Su, Junpeng Wang, Gan Zhao, Xiao Wang, Jinyao Li and Bin Wang
Pages 1659 - 1667
http://dx.doi.org/10.4161/hv.22105
Abstract | PDF

Antigen specific memory T cells (Tm) have shown to be an important factor in protecting hosts against subsequent infection by previously encountered pathogens. During T-cell activation, several cytokines including IL-6, IL-7 and IL-15, play crucial roles in the development of T cells into memory T cells. With the aim of generating specific Tm, we examined a strategy of sequential administration of molecular adjuvants. In this strategy a DNA vaccine encoding the VP1 capsid protein of foot and mouth disease virus (designated pcD-VP1) was co-delivered to mice along with an IL-6 expressing plasmid (pVAX-IL-6) as an initial molecular adjuvant and boosted with either an IL-7 or IL-15 expressing plasmid, (pVAX-IL-7 or proVAX-IL-15) as the secondary adjuvant. During the pcD-VP1 immunization, we demonstrated that the groups primed with IL-6 and boosted with either IL-7 or IL-15 resulted in the enhancement of cellular and humoral immune responses, maturation of dendritic cells (DCs) and macrophages, and a higher frequency of CD4⁺ Tm (characterized by expressing CD44^highCD62L^low markers, compared with the other groups). Thus, we took advantage of the different effects of cytokines on T cell development, not only to induce a higher level of immune responses after vaccination, but also to generate a higher ratio of CD4⁺ Tm in this sequential cytokine prime-boost study. This would then lead to the mounting of an effective long-term antigen specific immune response.

Vaccination with synthetic constructs expressing cytomegalovirus immunogens is highly T cell immunogenic in mice

Devon J. Shedlock, Kendra T. Talbott, Stephan J. Wu, Christine M. Wilson, Karuppiah Muthumani, Jean D. Boyer, Niranjan Y. Sardesai, Sita Awasthi and David B. Weiner
Pages 1668 - 1681
http://dx.doi.org/10.4161/hv.22447
Abstract | Full Text | PDF

There is no licensed vaccine or cure for human cytomegalovirus (CMV), a ubiquitous β-herpesvirus infecting 60–95% of adults worldwide. Infection can cause congenital abnormalities, result in severe disease in immunocompromised patients, and is a major impediment during successful organ transplantation. In addition, it has been associated with numerous inflammatory diseases and cancers, as well as being implicated in the development of essential hypertension, a major risk factor for heart disease. To date, limited data regarding the identification of immunogenic viral targets has frustrated CMV vaccine development. Based upon promising clinical data suggesting an important role for T cells in protecting against disease in the transplantation setting, we designed a novel panel of highly-optimized synthetic vaccines encoding major CMV proteins and evaluated their immune potential in murine studies. Vaccination induced robust CD8+ and CD4+ T cells of great epitopic breadth as extensively analyzed using a novel modified T cell assay described herein. Together with improved levels of CMV-specific T cells as driven by a vaccine, further immune evaluation of each target is warranted. The present model provides an important tool for guiding future immunization strategies against CMV.

REVIEWS

Harnessing DNA-induced immune responses for improving cancer vaccines

Andrés A. Herrada, Nicole Rojas-Colonelli, Paula González-Figueroa, Jonathan Roco, César Oyarce, Maarten A. Ligtenberg and Alvaro Lladser
Pages 1682 - 1693
http://dx.doi.org/10.4161/hv.22345
Abstract | Full Text | PDF

DNA vaccines have emerged as an attractive strategy to promote protective cellular and humoral immunity against the encoded antigen. DNA vaccines are easy to generate, inexpensive to produce and purify at large-scale, highly stable and safe. In addition, plasmids used for DNA vaccines act as powerful “danger signals” by stimulating several DNA-sensing innate immune receptors that promote the induction of protective adaptive immunity. The induction of tumor-specific immune responses represents a major challenge for DNA vaccines because most of tumor-associated antigens are normal non-mutated self-antigens. As a consequence, induction of potentially self-reactive T cell responses against such poorly immunogenic antigens is controlled by mechanisms of central and peripheral tolerance as well as tumor-induced immunosuppression. Although several DNA vaccines against cancer have reached clinical testing, disappointing results have been observed. Therefore, the development of new adjuvants that strongly stimulate the induction of antitumor T cell immunity and counteract immune-suppressive regulation is an attractive approach to enhance the potency of DNA vaccines and overcome tumor-associated tolerance. Understanding the DNA-sensing signaling pathways of innate immunity that mediate the induction of T cell responses elicited by DNA vaccines represents a unique opportunity to develop novel adjuvants that enhance vaccine potency. The advance of DNA adjuvants needs to be complemented with the development of potent delivery systems, in order to step toward successful clinical application. Here, we briefly discuss recent evidence showing how to harness DNA-induced immune response to improve the potency of cancer vaccines and counteract tumor-associated tolerance.

What you always needed to know about electroporation based DNA vaccines

Anita Gothelf and Julie Gehl
Pages 1694 - 1702
http://dx.doi.org/10.4161/hv.22062
Abstract | Full Text | PDF

Vaccinations are increasingly used to fight infectious disease, and DNA vaccines offer considerable advantages, including broader possibilities for vaccination and lack of need for cold storage. It has been amply demonstrated, that electroporation augments uptake of DNA in both skin and muscle, and it is foreseen that future DNA vaccination may to a large extent be coupled with and dependent upon electroporation based delivery. Understanding the basic science of electroporation and exploiting knowledge obtained on optimization of DNA electrotransfer to muscle and skin, may greatly augment efforts on vaccine development. The purpose of this review is to give a succinct but comprehensive overview of electroporation as a delivery modality including electrotransfer to skin and muscle. As well, this review will speculate and discuss future uses for this powerful electrotransfer technology.

SHORT REPORT

A multiagent filovirus DNA vaccine delivered by intramuscular electroporation completely protects mice from ebola and Marburg virus challenge

Rebecca J. Grant-Klein, Nicole M. Van Deusen, Catherine V. Badger, Drew Hannaman, Lesley C. Dupuy and Connie S. Schmaljohn
Pages 1703 - 1706
http://dx.doi.org/10.4161/hv.21873
Abstract | Full Text | PDF

We evaluated the immunogenicity and protective efficacy of DNA vaccines expressing the codon-optimized envelope glycoprotein genes of Zaire ebolavirus, Sudan ebolavirus, and Marburg marburgvirus (Musoke and Ravn). Intramuscular or intradermal delivery of the vaccines in BALB/c mice was performed using the TriGrid™ electroporation device. Mice that received DNA vaccines against the individual viruses developed robust glycoprotein-specific antibody titers as determined by ELISA and survived lethal viral challenge with no display of clinical signs of infection. Survival curve analysis revealed there was a statistically significant increase in survival compared to the control groups for both the Ebola and Ravn virus challenges. These data suggest that further analysis of the immune responses generated in the mice and additional protection studies in nonhuman primates are warranted.

RESEARCH PAPERS

Contactless magneto-permeabilization for intracellular plasmid DNA delivery in-vivo

Thomas J. Kardos and Dietmar P. Rabussay
Pages 1707 - 1713
http://dx.doi.org/10.4161/hv.21576
Abstract | Full Text | PDF

Electroporation, an attractive process for delivering DNA and other molecules into target cells in vivo and in vitro is limited by the necessity of electrodes that need to be in contact with the subject or object to be electroporated. We have used magnetic fields, which do not require material contact with the subject, to temporarily permeabilize cells in guinea pig skin in vivo to enhance uptake and expression of GFP plasmid DNA. The results show for the first time that magnetic fields can trigger a process likely similar to electroporation. In designing the magnetic pulses, our most important criterion was a high rate of change of the magnetic field, based on the principle described by Michael Faraday which is expressed by the formula: E = -dB/dt, (E, electric field, B, magnetic field, t, time). Magnetic fields were generated by a flat electromagnet in a hand-held applicator positioned above the target tissue. The magnetic pulses had a peak magnetic flux density of 4 tesla; 50 pulses were applied in 5 sec. Biphasic magnetic pulses were twice as effective as monophasic pulses and about equally effective as traditional electroporation pulses . Advantages of magnetopermeabilization over electoporation include: No contact between applicator and subject (“contact-less”); no need for invasive, disposable, sterile electrodes (“needle-less”); no pain from needles and reduced overall pain; no known side effects; easier and faster to administer than electroporation; less expensive due to absence of disposables; and, importantly, greater tissue penetration of the magnetic field allowing treatment of anatomical areas inaccessible by electroporation.

Electroporation driven delivery of both an IL-12 expressing plasmid and cisplatin synergizes to inhibit B16 melanoma tumor growth through an NK cell mediated tumor killing mechanism

Ha Kim and Jeong-Im Sin
Pages 1714 - 1721
http://dx.doi.org/10.4161/hv.22346
Abstract | Full Text | PDF

Combined therapy using chemotherapeutic drugs and immunotherapeutics offers some promise for treating patients with cancer. In this study, we evaluated whether cisplatin delivered by intratumoral (IT)-electroporation (EP) might enhance antitumor activity against established B16 melanoma and whether further addition of intramuscular (IM)-EP of IL-12 cDNA to IT-EP of cisplatin might augment antitumor therapeutic activity, with a focus on the underlining antitumor mechanism(s). When tumor (7 mm)-bearing animals were treated locally with cisplatin by IT-EP, they showed tumor growth inhibition significantly more than those without IT-EP. Moreover, IL-12 cDNA delivered by IM-EP was also able to inhibit tumor growth significantly more than control vector delivery. This tumor growth inhibition was mediated by NK cells, but not CD4+ T or CD8+ T cells, as determined by immune cell subset depletion and IFN-γ induction. Moreover, concurrent therapy using IT-EP of cisplatin plus IM-EP of IL-12 cDNA displayed antitumor therapeutic synergy. This therapeutic synergy appeared to be mediated by increased sensitivity of cisplatin-treated tumors to NK cell-mediated tumor killing. Taken together, these data support that cisplatin delivery by IT-EP plus IL-12 gene delivery by IM-EP are more effective at inducing antitumor therapeutic responses through increased sensitivity of cisplatin-treated tumors to NK cell-mediated tumor killing. This combined approach might have some implication for treating melanoma in patients.

Electrogenetherapy of B16.F10 murine melanoma tumors with an interleukin-28 expressing DNA plasmid

Kevin Shah, Richard J. Connolly, Taryn Chapman, Mark J. Jaroszeski and Kenneth E. Ugen
Pages 1722 - 1728
http://dx.doi.org/10.4161/hv.22560
Abstract | Full Text | PDF

Augmented delivery of cytokine-expressing DNA plasmids to subcutaneous tumors has been demonstrated to result in a level of enhanced anti-tumor activity. One delivery enhancement method which has been evaluated is in vivo electroporation (EP), a contact-dependent delivery technique where electric pulses are hypothesized to augment the transfer of DNA into cells and tissues through the induction of temporary cell membrane pores. Previous work by members of our group, as well as others, has demonstrated the anti-tumor effects of DNA plasmids expressing the cytokines IL-12 and IL-15. In this report the potential anti-tumor activity of a relatively newly-described cytokine, IL-28, was measured when administered intratumorally as a DNA expression plasmid (designated pIL28) to established murine (B16.F10) melanoma tumors. The administration of the IL-28 expressing plasmid was performed through enhanced delivery methods. One method was EP and the other a non-contact dependent technique using a helium plasma stream. IL-28 is a member of the type III interferon family of cytokines that has been characterized as possessing potent anti-viral activity. This cytokine has been demonstrated to function as an adjuvant in small animal model vaccination protocols and stimulates CD8+ CTL responses. In addition, stimulation of anti-tumor activity has been demonstrated in several studies using IL-28. Based on these activities, it was hypothesized that this cytokine could, when delivered through a DNA expression plasmid, mediate anti-tumor activity. The results of this study indicated that enhanced delivery of pIL-28 resulted in attenuation of tumor growth, compared with non-enhanced delivery. Of note, this is the first proof-of-concept experiment, of our knowledge, documenting the ability of a non-contact dependent helium plasma-based delivery method to mediate the enhancement of an anti-tumor effect by a cytokine-expressing DNA plasmid. This suggests the use of the helium plasma delivery method as an alternative or adjunctive method to EP for the effective delivery of agents that possess potential anti-tumor activity.

Non-contact helium-based plasma for delivery of DNA vaccines: Enhancement of humoral and cellular immune responses

Richard J. Connolly, Taryn Chapman, Andrew M. Hoff, Michele A. Kutzler, Mark J. Jaroszeski and Kenneth E. Ugen
Pages 1729 - 1733
http://dx.doi.org/10.4161/hv.21624
Abstract | Full Text | PDF

Non-viral in vivo administration of plasmid DNA for vaccines and immunotherapeutics has been hampered by inefficient delivery. Methods to enhance delivery such as in vivo electroporation (EP) have demonstrated effectiveness in circumventing this difficulty. However, the contact-dependent nature of EP has resulting side effects in animals and humans. Noncontact delivery methods should, in principle, overcome some of these obstacles. This report describes a helium plasma–based delivery system that enhanced humoral and cellular antigen-specific immune responses in mice against an intradermally administered HIV gp120-expressing plasmid vaccine (pJRFLgp120). The most efficient plasma delivery parameters investigated resulted in the generation of geometric mean antibody-binding titers that were 19-fold higher than plasmid delivery alone. Plasma mediated delivery of pJRFLgp120 also resulted in a 17-fold increase in the number of interferon-gamma spot-forming cells, a measure of CD8+ cytotoxic T cells, compared with non-facilitated plasmid delivery. This is the first report demonstrating the ability of this contact-independent delivery method to enhance antigen-specific immune responses against a protein generated by a DNA vaccine.

REVIEW

Plasmid IL-12 electroporation in melanoma

Edward Cha and Adil Daud
Pages 1734 - 1738
http://dx.doi.org/10.4161/hv.22573
Abstract | Full Text | PDF

Intratumoral gene electroporation uses electric charges to facilitate entry of plasmid DNA into cells in a reproducible and highly efficient manner, especially to accessible sites such as cutaneous and subcutaneous melanomas. Effective for locally treated disease, electroporation of plasmid DNA encoding interleukin-12 can also induce responses in untreated distant disease, suggesting that adaptive immune responses are being elicited that can target melanoma-associated antigens. In vivo electroporation with immunomodulatory cytokine DNA is a promising approach that can trigger systemic anti-tumor immune responses without the systemic toxicity associated with intravenous cytokine delivery and potentially offer complete long-term tumor regression.