Distinct immunological activation profiles of dSLIM® and ProMune® depend on their different structural context.

Introduction: DNA-based TLR9 agonists are potent activators of the immune system. ProMune® and dSLIM® belong to different families of TLR9 agonists and both have been established as cancer immunotherapeutics in clinical proof-of-concept studies. Unfortunately, ProMune® failed in pivotal oncological trials.

Design and characterization of the tumor vaccine MGN1601, allogeneic fourfold gene-modified vaccine cells combined with a TLR-9 agonist.

The tumor vaccine MGN1601 was designed and developed for treatment of metastatic renal cell carcinoma (mRCC). MGN1601 consists of a combination of fourfold gene-modified cells with the toll-like receptor 9 agonist dSLIM, a powerful connector of innate and adaptive immunity. Vaccine cells originate from a renal cell carcinoma cell line (grown from renal cell carcinoma tissue), express a variety of known tumor-associated antigens (TAA), and are gene modified to transiently express two co-stimulatory molecules, CD80 and CD154, and two cytokines, GM-CSF and IL-7, aimed to support immune response.

Design and Structural Requirements of the Potent and Safe TLR-9 Agonistic Immunomodulator MGN1703.

Single-stranded oligodeoxynucleotides (ODN), containing nonmethylated cytosine-guanine motifs (CpG ODN), are recognized by the innate immune system as "danger signals." CpG ODN are efficacious immunomodulators but require phosphorothioate (PT) or other backbone modifications for metabolic stability, which cause toxicities in mice and primates. We therefore designed a covalently closed DNA molecule (dSLIM(®)) where two single-stranded loops containing CG motifs are connected through a double-stranded stem in the absence of any nonnatural DNA component.

Phase I clinical study of the toll-like receptor 9 agonist MGN1703 in patients with metastatic solid tumours.

Purpose: This study was initiated to evaluate safety, toxicity, pharmacokinetics, and pharmacodynamics of treatment with MGN1703, a novel synthetic DNA-based toll-like receptor 9 (TLR9)-immunomodulator.

Methods: The study consisted of an escalating single dose regimen followed by a multiple dose part. Dose levels of 0.

Cationic lipid-formulated DNA vaccine against hepatitis B virus: immunogenicity of MIDGE-Th1 vectors encoding small and large surface antigen in comparison to a licensed protein vaccine.

Currently marketed vaccines against hepatitis B virus (HBV) based on the small (S) hepatitis B surface antigen (HBsAg) fail to induce a protective immune response in about 10% of vaccinees. DNA vaccination and the inclusion of PreS1 and PreS2 domains of HBsAg have been reported to represent feasible strategies to improve the efficacy of HBV vaccines. Here, we evaluated the immunogenicity of SAINT-18-formulated MIDGE-Th1 vectors encoding the S or the large (L) protein of HBsAg in mice and pigs.

Genuine Immunomodulation With dSLIM.

Toll-like receptors are sensing modulators of the innate immune system. One member of this protein family, Toll-like receptor (TLR)-9, is increasingly being investigated as therapeutic target for infectious diseases and cancer. Double-Stem Loop ImmunoModulator (dSLIM) is a new TLR-9 agonist in clinical development for patients with metastatic colorectal carcinoma.

Modular multiantigen T cell epitope-enriched DNA vaccine against human leishmaniasis.

The leishmaniases are protozoal diseases that severely affect large populations in tropical and subtropical regions. There are only limited treatment options and preventative measures. Vaccines will be important for prevention, control and elimination of leishmaniasis, and could reduce the transmission and burden of disease in endemic populations.

Combination of MIDGE-Th1 DNA vaccines with the cationic lipid SAINT-18: studies on formulation, biodistribution and vector clearance.

We have previously shown that the combination of MIDGE-Th1 DNA vectors with the cationic lipid SAINT-18 increases the immune response to the encoded antigen in mice. Here, we report on experiments to further optimize and characterize this approach. We evaluated different formulations of MIDGE-Th1 vectors with SAINT-18 by assessing their influence on the transfection efficiency in cell culture and on the immune response in mice.

Immune response induced by a linear DNA vector: influence of dose, formulation and route of injection.

Previously, minimalistic, immunogenetically defined gene expression (MIDGE) vectors were developed as effective and sophisticated carriers for DNA vaccination. Here we evaluate the influence of dose, formulation and delivery route on the immune response after vaccination with MIDGE-Th1 vectors encoding hepatitis B virus surface antigen (HBsAg). An HBsAg-specific IgG1 and IgG2a antibody response was induced in a dose-dependent manner, whereas the IgG2a/IgG1 ratio was independent of the injected DNA dose.

Peripheral non-viral MIDGE vector-driven delivery of beta-endorphin in inflammatory pain.

Background: Leukocytes infiltrating inflamed tissue produce and release opioid peptides such as beta-endorphin, which activate opioid receptors on peripheral terminals of sensory nerves resulting in analgesia. Gene therapy is an attractive strategy to enhance continuous production of endogenous opioids. However, classical viral and plasmid vectors for gene delivery are hampered by immunogenicity, recombination, oncogene activation, anti-bacterial antibody production or changes in physiological gene expression.

Minimal size MIDGE vectors improve transgene expression in vivo.

Viral and plasmid vectors may cause immunological side-effects resulting from the expression of therapeutically unwanted genes and from CpG motifs contained in their sequence. A new vector type for minimalistic, immunological-defined gene expression (MIDGE) may overcome these problems. MIDGE is a minimal size gene transfer unit consisting of the expression cassette, including promotor, gene and RNA-stabilizing sequences, flanked by two short hairpin oligonucleotide sequences.

Brief communication: stability and catalytic activity of novel circular DNAzymes.

Unlabelled: DNAzymes represent a new generation of catalytic nucleic acids for specific RNA targeting in order to inhibit protein translation from the specifically cleaved mRNA. The 10-23 DNAzyme was found to hydrolyze RNA in a sequence-specific manner both in vitro and in vivo. Although single-stranded DNAzymes may represent the most effective nucleic acid drug to date, they are nevertheless sensitive to nuclease degradation and require modifications for in vivo application.

Ballistic CTLA4 and IL-4 gene transfer into the lower lid prolongs orthotopic corneal graft survival in mice.

Purpose: To explore outflow from the eye and to determine and modulate the influence of lymphatic drainage on corneal graft survival in mice.

Methods: Tracer experiments were conducted in BALB/c mice using the (99m)Tc colloidal albumin Nanocoll. Count rates were determined in the eyes, submandibular lymph nodes, spleen, liver and blood 24 h after subconjunctival, intracorneal, intracameral (anterior chamber), intravenous and subcutaneous lower-lid or upper-lid injections ( n=6 each).

Influence of ballistic gene transfer on antigen-presenting cells in murine corneas.

Purpose: The outcome of corneal transplantation may depend on passenger cells in corneal epithelium and stroma. Their presence in normal corneas is controversial. This study aimed at examining this question and elucidating the still unknown influence of ballistic gene transfer.