ISCOMATRIX™ ADJUVANT LINKS INNATE AND ADAPTIVE IMMUNE RESPONSES

The ISCOMATRIX™ adjuvant has antigen delivery and presentation properties as well as immunomodulatory capabilities which combine to provide enhanced and accelerated immune responses. The responses are broad, including a range of sub classes of antibodies as well as both CD4+ and CD8+ T cells. A range of ISCOMATRIX™ vaccines (ISCOMATRIX™ adjuvant combined with antigen) have now been tested in clinical trials and have been shown to be generally safe and well tolerated as well as immunogenic, generating both antibody and T cell responses. The mechanisms by which ISCOMATRIX™ adjuvant facilitates its immune effects is the scope of significant study and indicates that ISCOMATRIX™ adjuvant (i) rapidly traffics antigen into the cytosol of multiple dendritic cell subsets, (ii) induces the induction of an array of cytokines and chemokines and (iii) links the innate and adaptive immune responses in vivo in a TLR-independent but MyD88-dependent manner. These data highlight the clinical utility of ISCOMATRIX™ adjuvant in the development of novel prophylactic and therapeutic vaccines

CpG ODN and ISCOMATRIX Adjuvant: A Synergistic Adjuvant Combination Inducing Strong T-Cell IFNResponses

For the induction of robust humoral and cellular immune responses, a strong rationale exists to use vaccine-adjuvant combinations possessing both immune modulatory and enhanced delivery capabilities. Herein, we evaluated the combination of 2 different adjuvants, a TLR9 agonist, composed of synthetic oligodeoxynucleotides (ODN) containing immunostimulatory CpG motifs (CpG), and ISCOMATRIX adjuvant (ISCOMATRIX), composed of saponin, phospholipid, and cholesterol, which possesses both immunostimulatory and delivery properties. While both individual adjuvants have been shown effective in numerous preclinical and clinical studies, it is likely that for optimal adjuvant activity a combined adjuvant approach will be necessary. Herein, using three different antigens, namely, hepatitis B surface antigen (HBsAg), ovalbumin (OVA), and influenza A haemagglutinin antigen (HA), we show in mice that some adjuvant effects of CpG and ISCOMATRIX are further enhanced if they are used in combination. In particular, with all three antigens, IFN-levels were greatly increased with the CpG/ISCOMATRIX combination. The ability of the CpG/ISCOMATRIX combination to induce antitumor responses when administered with OVA following administration to mice of a highly metastatic OVA-secreting tumor cell line (B16-OVA melanoma) was also demonstrated. Thus the CpG/ISCOMATRIX combination may prove to be a valuable tool in the development of novel or improved vaccines

ISCOMATRIX vaccines mediate CD8+ T-cell cross-priming by a MyD88-dependent signaling pathway

Generating a cytotoxic CD8+ T-cell response that can eradicate malignant cells is the primary objective of cancer vaccine strategies. In this study we have characterized the innate and adaptive immune response to the ISCOMATRIX adjuvant, and the ability of vaccine antigens formulated with this adjuvant to promote antitumor immunity. ISCOMATRIX adjuvant led to a rapid innate immune cell response at the injection site, followed by the activation of natural killer and dendritic cells (DC) in regional draining lymph nodes. Strikingly, major histocompatibility complex (MHC) class I cross-presentation by CD8α+ and CD8α− DCs was enhanced by up to 100-fold when antigen was formulated with ISCOMATRIX adjuvant. These coordinated features enabled efficient CD8+ T-cell cross-priming, which exhibited prophylactic and therapeutic tumoricidal activity. The therapeutic efficacy of an ISCOMATRIX vaccine was further improved when co-administered with an anti-CD40 agonist antibody, suggesting that ISCOMATRIX-based vaccines may combine favorably with other immune modifiers in clinical development to treat cancer. Finally, we identified a requirement for the myeloid differentiation primary response gene 88 (MyD88) adapter protein for both innate and adaptive immune responses to ISCOMATRIX vaccines in vivo. Taken together, our findings support the utility of the ISCOMATRIX adjuvant for use in the development of novel vaccines, particularly those requiring strong CD8+ T-cell immune responses, such as therapeutic cancer vaccines

Long-term storage of DNA-free RNA for use in vaccine studies

RNA replicons represent potential vaccine delivery vehicles, but are considered too unstable for such use. This study examined the recovery, integrity and function of in vitro transcribed replicon RNA encoding hepatitis C virus (HCV) proteins. To remove residual template DNA, the RNA was digested with TURBO DNase followed by RNeasy DNase set and purified through an RNeasy column. The RNA was freeze-dried in distilled water or trehalose, stored under nitrogen gas for up to 10 months and analyzed at different time points. The recovery of RNA stored at ≤4°C that was freeze-dried in distilled water varied between 66% to zero of that recovered from RNA freeze-dried in 10% trehalose, a figure that depended on the duration of storage. In contrast, the recovery of the RNA stored in trehalose was consistently high for all time points. After recovery, both RNAs were translationally competent and expressed high levels of proteins after transfection, although the level of expression from the trehalose-stored RNA was consistently higher. Thus the addition of trehalose permitted stable storage of functional RNA at 4°C for up to 10 months and this permits the development of RNA vaccines, even in developing countries where only minimum storage conditions (e.g., 4°C) can be achieved.Kathryn L. Jones, Debbie Drane and Eric J. Gowan

Development of prophylactic and therapeutic vaccines using the ISCOMATRIX adjuvant

Adjuvants are components that when added to subunit antigen (Ag) vaccines boost their immunogenicity and thus immune efficacy. However, there are few adjuvants that are approved for clinical use resulting in a critical need for the development of safe and effective adjuvants for use in both prophylactic and therapeutic vaccines. The paucity of appropriate adjuvants is more chronic for the development of therapeutic vaccines for cancer and chronic infectious disease, which need to induce cytotoxic T-cell responses via cross-presentation of the vaccine Ag by dendritic cells. The ISCOMATRIX adjuvant represents a unique adjuvant system that facilitates Ag delivery and presentation as well as immunomodulation to provide enhanced and accelerated immune responses. The immune responses generated are of broad specificity to the vaccine Ag, and include robust antibody responses of multiple subclasses as well as both CD4+ and CD8+ T-cell responses. Here we discuss our understanding of the mechanisms of action by which ISCOMATRIX adjuvant may facilitate these integrated immune responses and touch on insights gained through its clinical experience

CpG ODN and ISCOMATRIX Adjuvant: A Synergistic Adjuvant Combination Inducing Strong T-Cell IFN-γ Responses

For the induction of robust humoral and cellular immune responses, a strong rationale exists to use vaccine-adjuvant combinations possessing both immune modulatory and enhanced delivery capabilities. Herein, we evaluated the combination of 2 different adjuvants, a TLR9 agonist, composed of synthetic oligodeoxynucleotides (ODN) containing immunostimulatory CpG motifs (CpG), and ISCOMATRIX adjuvant (ISCOMATRIX), composed of saponin, phospholipid, and cholesterol, which possesses both immunostimulatory and delivery properties. While both individual adjuvants have been shown effective in numerous preclinical and clinical studies, it is likely that for optimal adjuvant activity a combined adjuvant approach will be necessary. Herein, using three different antigens, namely, hepatitis B surface antigen (HBsAg), ovalbumin (OVA), and influenza A haemagglutinin antigen (HA), we show in mice that some adjuvant effects of CpG and ISCOMATRIX are further enhanced if they are used in combination. In particular, with all three antigens, IFN-γ levels were greatly increased with the CpG/ISCOMATRIX combination. The ability of the CpG/ISCOMATRIX combination to induce antitumor responses when administered with OVA following administration to mice of a highly metastatic OVA-secreting tumor cell line (B16-OVA melanoma) was also demonstrated. Thus the CpG/ISCOMATRIX combination may prove to be a valuable tool in the development of novel or improved vaccines

Association of antigens to ISCOMATRIX™ adjuvant using metal chelation leads to improved CTL responses

The association of antigen with ISCOMATRIX™ adjuvant has been shown to be important for the optimal induction of cytotoxic T lymphocyte (CTL) responses. Here, we describe a simple broadly applicable method for associating recombinant proteins with hexa-histidine tags to ISCOMATRIX™ adjuvant utilising metal-affinity chelating interactions. The metal chelation binding step can be performed in a wide range of buffers, including commonly used denaturants such as urea, which makes it an ideal strategy for formulating proteins which are otherwise insoluble. Following association of protein with the chelating ISCOMATRIX™ adjuvant, the denaturant can be removed. Further, we show enhanced CTL responses with a protein-associated chelating ISCOMATRIX™ vaccine compared to a non-associated ISCOMATRIX™ vaccine

Association of antigens to ISCOMATRIX™ adjuvant using metal chelation leads to improved CTL responses

The association of antigen with ISCOMATRIX™ adjuvant has been shown to be important for the optimal induction of cytotoxic T lymphocyte (CTL) responses. Here, we describe a simple broadly applicable method for associating recombinant proteins with hexa-histidine tags to ISCOMATRIX™ adjuvant utilising metal-affinity chelating interactions. The metal chelation binding step can be performed in a wide range of buffers, including commonly used denaturants such as urea, which makes it an ideal strategy for formulating proteins which are otherwise insoluble. Following association of protein with the chelating ISCOMATRIX™ adjuvant, the denaturant can be removed. Further, we show enhanced CTL responses with a protein-associated chelating ISCOMATRIX™ vaccine compared to a non-associated ISCOMATRIX™ vaccine