miR-21 and miR-155 are associated with mitotic activity and lesion depth of borderline melanocytic lesions.

BACKGROUND: Expression of microRNAs (miRs) has been shown to be altered in many solid tumours and is being explored in melanoma. The malignant potential of some melanocytic lesions is difficult to predict. We hypothesised that characterisation of miR expression in borderline melanocytic proliferations would lead to the identification of a molecular profile that could be used with known prognostic factors to differentiate lesions with high malignant potential. METHODS: The miR expression profile of melanocytic lesions (benign naevi, malignant melanoma and borderline melanocytic tumours) was evaluated by real-time PCR. RESULTS: PCR analysis revealed primary cutaneous melanomas had an 8.6-fold overexpression of miR-21 and a 7.5-fold overexpression of miR-155 compared with benign naevi (P<0.0001). In situ hybridisation confirmed these results. miR-21 and miR-155 were significantly overexpressed within borderline lesions (P=0.0011 and P=0.0048, respectively). When borderline lesions were categorised by mitotic activity and Breslow thickness, miR-21 was associated with mitotic activity and miR-155 was associated with thickness (P<0.025). Among 14 patients with borderline lesions who underwent sentinel lymph node biopsy (SLNB), positive SLNB was associated with increased miR-21 and miR-155 in the primary lesion compared with lesions with a negative SLNB. CONCLUSION: MicroRNA expression profiles can be used to characterise atypical melanocytic lesions.

TGF-beta inhibition of CTL re-stimulation requires accessory cells and induces peroxisome-proliferator-activated receptor-gamma (PPAR-gamma).

Effective cellular immunity to Epstein-Barr virus (EBV), necessary to prevent or cure many post-transplant lymphoproliferative disorders (PTLD), can be inhibited by transforming growth factor-beta (TGF-beta). In vitro, TGF-beta inhibits memory CTL re-stimulation from whole PBMC. We show that the effect of TGF-beta on CTL re-stimulation is not directly on the T cell, but requires an accessory cell (AC) population. Further, pre-treatment of AC with TGF-beta significantly reduces memory CTL re-stimulation and suppresses delayed type hypersensitivity (DTH) responses. Addition of exogenous interferon-gamma to the AC overcomes the effects of TGF-beta. TGF-beta pre-treatment also up-regulates expression of peroxisome-proliferator-activated receptor-gamma (PPAR-gamma) in CD14(+) AC. Importantly, pre-treatment of AC with the PPAR-gamma ligand, ciglitazone, results in significantly reduced memory CTL re-stimulation. Thus, the effects of TGF-beta in this system may be mediated in part via PPAR-gamma, and PPAR-gamma activation could have significant inhibitory effects on memory T-cell responses by affecting AC function. These data have important implications in understanding how memory CTL are re-stimulated and function to prevent disease, especially PTLD.

Novel vaccine strategies to T-independent antigens.

T cell independent antigens do not require T cell help to induce an immune response, and are characterized by a lack of immunologic memory. These antigens can be divided into two classes, TI-1 or TI-2. TI-1 antigens, such as bacterial lipopolysaccharide, are potent B-cell mitogens, capable of non-specific, polyclonal activation of B cells. In contrast, TI-2 antigens can only activate mature B cells and consist of highly repetitive structures, such as capsular polysaccharides (CPS) from bacteria. Many vaccines currently in use consist of purified capsular polysaccharides from pathogenic bacteria such as Streptococcus pneumoniae and Neisseria meningitidis. These vaccines are efficacious in immune-competent adults, however, due to their TI-2 nature, are not effective in children <2 years of age. Converting polysaccharides into T cell dependent (TD) antigens, allows children, <2, to produce an effective immune response. This review focuses on various strategies used to convert the immune response to polysaccharide antigens from TI-2 to a TD response. Conjugate vaccines, anti-idiotypic antibodies, phage display library technology and DNA vaccines are discussed.

A DNA vaccine encoding a peptide mimic of Streptococcus pneumoniae serotype 4 capsular polysaccharide induces specific anti-carbohydrate antibodies in Balb/c mice.

Streptococcus pneumoniae is a leading cause of morbidity and mortality in both the developing and developed world. The T-independent nature of the current polysaccharide vaccine renders it ineffective in elderly adults and children <2 years of age. Although the recently licensed conjugate vaccines are capable of producing T-cell-dependent immunity, they also have their limitations, namely a lack of response in certain populations. Our laboratory has focused on a different approach, DNA vaccination. We have defined a peptide sequence (pep4) that mimics the S. pneumoniae serotype 4 capsular polysaccharide (PPS4) using a monoclonal antibody to PPS4 (mAb4) and phage display library. Pep4 was synthesized, complexed to proteosomes and used to immunize mice. We have shown by ELISA that this peptide mimic is capable of eliciting an anti-PPS4 immune response significantly higher than in negative control mice (P<0.05). A PPS4-DNA vaccine was made by cloning an oligodeoxynucleotide encoding pep4 into the HBcAg vector. Following epidermal immunization with the PPS4-DNA vaccine, mice produced an anti-PPS4 antibody response significantly higher than in mice immunized with an impertinent DNA vector (P<0.05). Our results demonstrate the feasibility of peptide mimicry in DNA vaccine development.

Vaccines against polysaccharide antigens.

Encapsulated bacteria such as Streptococcus pneumoniae, Neisseria meningitidis, and Haemophilus influenzae serogroup B (Hib) are a major cause of disease worldwide. Vaccine development against these organisms has targeted their capsular polysaccharides (CPS), as anti-capsular antibodies often protect against disease. The capsular polysaccharide vaccines that have been available against these organisms are neither immunogenic nor protective in young children and certain immunocompromised individuals. In general, polysaccharide (PS) antigens elicit a T-independent immune response, characterized by lack of memory, and poor immunogenicity at the extremes of life. Efforts to overcome the poor immunogenicity of CPS vaccines have led to development of conjugate vaccines. By conjugating CPS to carrier proteins it is possible to induce a T-dependent immune response against these antigens. Although conjugate vaccines have been successful against Hib disease, their applicability to multi-serotype/serogroup pathogens like the pneumococcus or the meningococcus is questioned. As a result, alternative vaccines including (1) surface proteins conserved across serotypes/serogroups, (2) peptides that mimic PS antigens and (3) DNA vaccines are presently under investigation. This review will highlight the potential and limitations of both CPS and CPS-conjugate vaccines against encapsulated bacteria as well as alternative strategies against PS antigens.