Biological Adjuvants Research Articles

Augmentation of HIV-1 Subtype C Vaccine Constructs Induced Immune Response in Mice by CpG Motif 1826-ODN

The greatest biomedical challenge of this century is to develop a preventive vaccine against Human Immunodeficiency Virus (HIV-1). For an HIV vaccine to be effective, it appears logical to develop new strategies that enhance the level of the immune response as well as steer it towards the desirable cellular type. In view of this, there is a need for rational inclusion of biological adjuvants into the HIV-1 vaccination strategies that could potentiate the immune responses both qualitatively and quantitatively. The adjuvant may include the immunostimulatory oligonucleotides containing CpG motifs, whose immunomodulatory characters are well established and represent the basis for an effective vaccine adjuvant. In our study, we investigated the use of an immunostimulatory oligonucleotide (or CpG motif), 1826-ODN to augment the immune response elicited by plasmid DNA vaccine constructs containing Indian subtype C HIV-1 envelope gp120 and gag-protease genes in Balb/c mouse model system. A dose of 2-microg CpG motifs/mouse was found to be optimum when co-administered with the DNA vaccine constructs with the peak level of humoral and cell mediated immune responses at 6 weeks post immunization. Murine IFN-gamma ELISpot assay demonstrated that the use of 1826-ODN led to a broad based and long term recognition of the subtype C envelope and gag peptides. The use of CpG motifs has been effective in augmenting the immune responses generated by the DNA constructs. Taken together, these results are an important advancement towards the design of future preclinical and clinical trials of these vaccine constructs.

DNA Vaccines

Compared with conventional approaches, DNA vaccine technology is a relatively new methodology for producing effective vaccines and has the major advantage of being simple and not requiring any special techniques for the purification and characterisation of recombinant proteins in the correct structure and conformation. The beauty of this approach lies in the ability of DNA vaccines to induce both cellular and humoral responses, thereby having special applications for making improved prophylactic vaccines against diseases for which traditional approaches have failed. Furthermore, the technology also provides an opportunity for developing therapeutic vaccines to treat chronic diseases such as HIV infection and viral hepatitis. These types of vaccines utilise advances in the fields of immunology and molecular biology to tailor more specifically the types of immune responses needed (cellular and/or humoral) against selected targets. In addition, DNA vaccines are potentially safer because, by delivering only the gene(s) encoding the particular immunogen(s) against which a protective or therapeutic immune response is desired, one can avoid the limitations and risks of certain other approaches such as live attenuated pathogens. So far, the efficacy of DNA vaccines has been low to modest in clinical trials, therefore significant efforts have been made to increase the potency of DNA vaccines by increasing the expression levels of the gene of interest, developing novel formulations and delivery technologies, and utilising biological adjuvants. Second-generation DNA vaccines are under development. This review will focus on the status of DNA vaccines against disease and the progress made in increasing the potency and efficacy of DNA vaccines.

Whole cell preparations of MCF-7 breast carcinoma cells are more active than lysed cell preparations in lymphocyte blastogenesis assays (LBAs) of breast cancer patients

2596 Background: We have previously reported that whole cell preparations of breast carcinoma cells from either an autologous or allogeneic source (MCF-7 cells) could produce immune responses, increased lymphocyte proliferation in a LBA, in breast cancer patients vaccinated with tumor cells, tumor antigens and two biological adjuvants (IL-2 and GM-CSF). We also showed that after vaccination with the tumor antigens (CEA, CA 15–3 and CA125), there were significant increases in the proliferation of the breast cancer patients' lymphocytes in response to the antigens in the LBA. The present study was undertaken to determine if tumor cell lysates would be more antigenic than the whole cell preparations. Methods: MCF-7 cells were lysed by two cycles of freeze thawing and a sample was tested in cell culture for viability. The lysate was then tested in LBAs and the results compared to those from whole cell preparations of MCF-7 cells (detached from cell culture plate, allowed to regenerate surface antigens while being agitated, and treated with mitomycin C to prevent further proliferation). Results: We found that there was significantly less activity in the lysate (38% to 81% less) than the whole cell preparation in 6 of the LBAs from 9 breast cancer patients. However, the lysate was much more active in 2 of the patients' LBAs (280% and 350% increases). One patient showed no difference between the two preparations. Conclusions: Whole cell preparations of allogeneic MCF-7 cells have immunologic activity in more breast cancer patients than cell lysates, as determined by LBAs. However, a small but significant proportion (20 to 25%) of patients have a significantly greater immunologic response to the cell lysate. Further investigation of the pre-vaccine LBA activity of the whole cell and cell lysate as they relate to immunologic response to our breast cancer vaccine is warranted to determine if the cell lysate should be added to the vaccine. No significant financial relationships to disclose.

Multiple antigens in a breast cancer vaccine result in a higher immune response rate in breast cancer patients

2607 Background: We have previously reported that whole cell preparations of breast carcinoma cells from either an autologous or allogeneic source (MCF-7 cells) and tumor antigens (CA 15–3, CEA and CA125) could produce immune responses, increased lymphocyte proliferation in a Lymphocyte Blastogenesis Assay (LBA), in breast cancer patients vaccinated with tumor cells, tumor antigens and two biological adjuvants (IL-2 and GM-CSF). The present study was undertaken to determine if the use of multiple antigens, cells and tumor antigens, in a breast cancer vaccine could increase the proportion of patients demonstrating an immune response to vaccination. Methods: Twenty-four patients were vaccinated 6 times with allogeneic cells, CA 15–3, CEA, and CA 125 antigens. LBAs were done before and after vaccination and the ratio of antigen to the media control was determined for each antigen before and after vaccination (a ratio of 1 means no stimulation by the antigen). The post-vaccine ratios for each patient's antigens were compared to the pre-vaccine ratio and an increase of 2 times was considered a significant immune response. Results: The following immune response rates were found: 25% (6 of 24 patients) for allogeneic MCF-7 cells, 21% (5 of 24 patients) for CA 15–3, 25% (6 of 24 patients) for CEA and 22% (5 of 23 patients) for CA 125. When an immune response to either the whole cell or any one of the tumor antigens was considered an immune response for that patient, then 58% (14 of 24) of the patients had immune responses. Conclusions: Whole cell preparations of allogeneic MCF-7 cells and tumor antigens (CA 15–3, CEA and CA 125) can induce immune responses in breast cancer patients. Each antigen produces an immune response, as measured by a LBA, in approximately 25% of the patients but by using multiple antigens over half of the patients will have an immune response to one or more of the antigens. This suggests that the use of multiple antigens in cancer vaccines may increase the clinical response rate, but a larger study with long-term follow-up will be needed. No significant financial relationships to disclose.

Multiple antigens in a breast cancer vaccine result in a higher immune response rate in breast cancer patients

2607 Background: We have previously reported that whole cell preparations of breast carcinoma cells from either an autologous or allogeneic source (MCF-7 cells) and tumor antigens (CA 15–3, CEA and CA125) could produce immune responses, increased lymphocyte proliferation in a Lymphocyte Blastogenesis Assay (LBA), in breast cancer patients vaccinated with tumor cells, tumor antigens and two biological adjuvants (IL-2 and GM-CSF). The present study was undertaken to determine if the use of multiple antigens, cells and tumor antigens, in a breast cancer vaccine could increase the proportion of patients demonstrating an immune response to vaccination. Methods: Twenty-four patients were vaccinated 6 times with allogeneic cells, CA 15–3, CEA, and CA 125 antigens. LBAs were done before and after vaccination and the ratio of antigen to the media control was determined for each antigen before and after vaccination (a ratio of 1 means no stimulation by the antigen). The post-vaccine ratios for each patient's antigens were compared to the pre-vaccine ratio and an increase of 2 times was considered a significant immune response. Results: The following immune response rates were found: 25% (6 of 24 patients) for allogeneic MCF-7 cells, 21% (5 of 24 patients) for CA 15–3, 25% (6 of 24 patients) for CEA and 22% (5 of 23 patients) for CA 125. When an immune response to either the whole cell or any one of the tumor antigens was considered an immune response for that patient, then 58% (14 of 24) of the patients had immune responses. Conclusions: Whole cell preparations of allogeneic MCF-7 cells and tumor antigens (CA 15–3, CEA and CA 125) can induce immune responses in breast cancer patients. Each antigen produces an immune response, as measured by a LBA, in approximately 25% of the patients but by using multiple antigens over half of the patients will have an immune response to one or more of the antigens. This suggests that the use of multiple antigens in cancer vaccines may increase the clinical response rate, but a larger study with long-term follow-up will be needed. No significant financial relationships to disclose.

Internal limiting membrane: ultrastructural relationships, with clinical implications for macular hole healing

Background: Recent data on macular hole formation have changed our understanding of the mechanisms involved. However, the mechanisms of macular hole closure, despite the introduction of internal limiting membrane (ILM) removal, are not fully understood. We reviewed the charts of patients who underwent surgery for macular hole with and without intentional removal of ILM and reviewed the basic science literature on the components and relationships of the ILM-glial cell interface to help interpret our surgical data.Methods: We reviewed the records of 205 eyes with a clinical diagnosis of macular hole at a university centre in Montreal between 1998 and 2002. Of these, 117 eyes of 108 patients were included. Vitrectomy with intentional ILM removal was done under indocyanine green (ICG) staining. Macular hole stage and rates of hole closure, failure and reopening were recorded, comparing the eyes with intentional ILM peeling versus no ILM peeling. Anatomic success was defined as a flat/closed configuration. A basic science literature review was done through the PubMed search engine with the key words “internal limiting membrane”, “internal limiting lamina”, “macular hole”, “retinal glial cells” and “retinal Muller cells”.Results: Eighty-four eyes had ICG-guided ILM removal (26 stage 2 macular holes, 45 stage 3 holes and 13 stage 4 holes), and 33 eyes had no ILM removal (10 stage 2 holes and 23 stage 3 holes). Of the 117 eyes 110 (94.0%) were successfully closed. Four (3.6%) of the closed holes reopened (none had ILM peeling), and 7 (6.0%) of the 117 eyes had primary failure. Ten of the 11 eyes that had primary failure or hole reopening were reoperated, and 8 were successfully closed after the second procedure. One patient refused reoperation. Including second procedures, the final anatomic closure rate was 97.4% (114/117).Interpretation: Intentional ILM peeling is not mandatory for all macular holes to close but does play a role in preventing reopening. Removal of ILM increases the rate of flat/closed hole configuration on reoperation without the need for biologic adjuvants. Review of the basic science evidence helped clarify the possible role of ILM removal.

Cytokine, chemokine, and costimulatory molecule modulation to enhance efficacy of HIV vaccines.

Understanding key intervention points in developing immune responses may allow the rational inclusion of biological adjuvants into vaccines that could potentiate the immune response both quantitatively and qualitatively and enhance effective memory responses. Cytokine and chemokine combinations can potentially help target antigen to the appropriate antigen presenting cell and initiate maturation of these presenting cells, attract cells expressing different chemokine receptors, steer cellular immune responses toward Th1 and CD8 CTL, and enhance systemic and mucosal IgG and secretory IgA antibodies and determine their isotype balance. Animal protection studies suggest that synergistic combinations of cytokines and immunomodulating molecules may be required to protect from a viral challenge. For example, GM-CSF has been shown to be synergistic with IL-12 or CD40 ligand for induction of CTL and for antiviral protection, and the triple combination of GM-CSF, IL-12, and TNF alpha appears to induce the most effective protection in some mouse models. Chemokine-antigen fusions have also been shown to enhance immunogenicity of the antigen. Combinations of costimulatory molecules have been found to be synergistic when incorporated in a vaccine. Combined use of newer more potent vaccine constructs, containing codon optimized epitopes, relevant CpG motifs, cytokines, costimulatory molecules and chemokines, used in heterologous prime-boost strategies with viral vector vaccines or recombinant proteins, might afford the most potent vaccine approaches yet developed. In this review we will discuss the application and delivery of cytokines, costimulatory molecules, and chemokines toward improving current vaccine strategies.

Differentiation of immunostimulatory stem-cell- and monocyte-derived dendritic cells involves maturation of intracellular compartments responsible for antigen presentation and secretion.

Dendritic cells (DCs) are important for the induction of primary T-cell responses and may serve as "biologic adjuvants" in therapeutic protocols. However, given the "plasticity" of this antigen-presenting cell, it remains unclear which DC type (source, subtype, and stage of differentiation) should be applied clinically. To provide additional insight in this selection process, we have, for the first time, analyzed the in vitro differentiation of CD34(+) precursor-derived and monocyte-derived DCs for ultrastructure, phenotype, and function. The ultrastructural intracytoplasmic differentiation of DCs correlated with increasing T-cell stimulatory activity of these cells. "Early-stage"-DCs proliferate, exhibit high levels of soluble antigen uptake, and moderate T-cell stimulatory capacity, and are characterized by centrally located nuclei and numerous enlarged mitochondria. "Intermediate-stage"-DCs are enlarged cells with enhanced T-cell stimulatory activity and pronounced cytoplasmic protein synthesis machinery. "Late-stage" (LS)-DCs exhibit a mature secretory cell phenotype and low proliferative index. They express high levels of the HLA-DR, CD40L, B7-1, and B7-2 molecules and CD83, a specific marker of mature DCs, and appear maximally stimulatory to T cells. Ultrastructurally, LS-DCs feature an accentric nucleus, an enlarged cytoplasm, containing numerous secretory storage vesicles, along with a fully developed Golgi complex. LS-DCs exhibited numerous multivesicular and multilaminar structures containing major histocompatibility complex class II molecules, consistent with the MIIC (peptide-loading) compartment. In extended studies, cultured CD14(+) monocyte-derived DCs displayed a similar, but accelerated, temporal differentiation staging pattern.

Hydroxyapatite: Catalyst or conjuror?

A prospective, randomized, intermediate-term study of 174 patients (199 hips) comparing proximally hydroxyapatite (HA)-coated and non-HA-coated surfaces of identical stem design is presented. Clinical and radiographic data from 152 patients (174 hips) confirmed comparable outcomes in both groups according to the Hospital for Special Surgery modification of the Postel–d'Aubigne Hip Rating System. The non-HA group showed significantly higher (P=.03) activity-related thigh pain, however. There were no significant radiographic differences, with all stems showing bone ongrowth. The use of HA-coated stems resulted in early recovery of function and reduced activity-related trochanteric and thigh pain. A possible explanation for the difference is most likely early superior osseointegration. HA as a biologic adjuvant should gain widespread acceptance because of rapid recovery of function and lesser trochanteric and thigh pain. Copyright 2002, Elsevier Science (USA). All rights reserved.

Toll-like receptors: molecular mechanisms of the mammalian immune response.

The unrelenting bombardment by microbial pathogens from the environment and their increasing resistance to medical treatments are a constant threat to the survival of all organisms on earth. Microbes are covered by molecular patterns that are common among a broad range of pathogens. These include the lipopolysaccharides (LPS) of Gram-negative bacteria, lipoteichoic acids of Gram-positive bacteria, lipoproteins of bacteria and parasites, glycolipids of mycobacteria, mannans of yeast and double-stranded RNAs of viruses.1,2 Recognition of and responses to these molecules are controlled by a wide variety of cellular receptors. The best characterized receptors are the T-cell receptor and B-cell antibody receptor of the adaptive immune response, the specificity of which is randomly generated and clonally selected during the development of T and B lymphocytes.3 Unlike the receptors of the adaptive immune system, the pattern recognition receptors of the innate immune system have predetermined specificity generated early on in evolution and play an essential role in the determination of self versus non-self during the initial rapid responses to infection.1,2 As described in this review, a family of cell surface receptors, termed Toll-like receptors, are emerging as key regulators of host responses to infection.

Macular hole surgery in 2000.

Patient selection and surgical experience have increased the closure rates for idiopathic macular holes to more than 90%. The best surgical results are obtained in patients with acute idiopathic macular holes. Biologic adjuvants are not as useful as they were once thought to be. Internal limiting membrane peeling and endolaser are useful for failed macular holes, reopened macular holes, and chronic macular holes. Internal limiting membrane peeling may not be necessary for acute idiopathic macular holes, provided a complete posterior vitreous detachment is created. Silicone oil tamponade should be considered in patients who have positioning problems or air travel plans. Humidifying the infusion air may reduce postoperative visual field defects. Every patient with a macular hole should be given the opportunity to improve his or her vision with macular hole surgery.

Alloantigenic stimulation bypasses CD28-B7 costimulatory blockade by an interleukin-2-dependent mechanism.

Allogeneic leukocytes have been used as biological adjuvants for T cell-specific responses to tumor and recall antigens, but the mechanisms underlying this effect have not been fully understood. The present study investigates whether alloantigen stimulation of human T cells would bypass an in vitro T cell costimulatory dysfunction induced by CTLA4Ig blockage of CD28-B7 interaction. Here, we demonstrate that costimulation with intact allogeneic leukocytes plus viral antigen circumvented the inhibition of this costimulatory pathway via interleukin-2 (IL-2) production, resulting in the generation of influenza-specific cytotoxic T lymphocytes (CTL). The alloantigen-induced help for influenza-specific CTL generation did not require cell-to-cell contact between responding and allogeneic stimulator cells. These results suggest that alloantigens can be used to bypass defects in the CD28-B7 costimulatory pathway and, therefore, may contribute to understanding the mechanisms of alloantigen-induced restoration of T cell-mediated immunity.

REDUCTION IN SERUM IL-6 AFTER VACINATION OF BREAST CANCER PATIENTS WITH TUMOUR-ASSOCIATED ANTIGENS IS RELATED TO ESTROGEN RECEPTOR STATUS

Elevated serum IL-6 concentrations have been associated with poor prognosis in a variety of cancers, and decreases in serum IL-6 concentrations have been reported after chemotherapy. We have demonstrated that serum IL-6 concentrations are elevated in breast cancer patients [normal women 0.7±2.5pg/ml (n=36), breast cancer patients 38.3±138.7pg/ml (n=111)]. After vaccination of breast cancer patients with a combination of tumour-associated antigens and biological adjuvants (IL-2 and GM-CSF), the concentration of IL-6 decreased significantly (P<0.05) to 8.1±14.6pg/ml (n=85). Other studies have shown that oestrogen suppresses IL-6 production in oestrogen receptor positive breast cancer cells. We have demonstrated that the decrease in IL-6 associated with vaccination is related to the oestrogen receptor status of the tumours from breast cancer patients, as a decrease in IL-6 from 124.0±267.5pg/ml (n=26) to 6.2±11.0pg/ml (n=34) only occurs in patients with oestrogen receptor negative tumours. The IL-6 concentration in breast cancer patients with oestrogen receptor positive tumours remained unchanged (9.5pg/ml before vaccination, and 9.3pg/ml after vaccination). These results suggest that postmenopausal women with oestrogen receptor negative breast cancers, who do not respond well to either hormonal therapy with tamoxifen or adjuvant chemotherapy, may have a significant response to vaccination with autologous tumour-associated antigens.

Long-term incidence of reopening of macular holes

Objective To report the incidence of macular hole reopening after initial successful surgery. Design A noncomparative retrospective case series. Participants From a series of 137 consecutive cases of idiopathic macular holes operated on between August 1993 and May 1996, the authors analyzed the charts of 116 cases of successful surgery. Setting A single university-based ophthalmology department. Intervention Pars plana vitrectomy, posterior hyaloid peeling, fluid-air and air-gas exchange with a nonexpansile 17% C 2F 6 mixture followed by face-down positioning. Eighty-eight patients (64%) received autologous platelets as a biologic adjuvant. The anatomic success rate at the first postoperative month was 116 of 137 (85%). One hundred and six patients (91%) were followed up for more than 2 years. Main outcome measure Macular anatomic status. Results Mean follow-up was 27 months. Eleven eyes of 11 patients (9.5%) had reopening of the macular hole. The mean period between hole surgery and reopening was 15.4 months (range, 8–29). In 8 of these 11 cases, reopening occurred after cataract extraction. In two cases, an epiretinal membrane was noted. In another case cystoid macular edema was present. The final anatomic success rate with one surgical procedure was 105 of 137 (77%). Eight reopened holes were reoperated on and all were anatomic successes at 1 month. However, four patients experienced a second reopening. Conclusions Macular hole reopening occurred in 9.5% of cases (11 of 116). The cause of reopening might have been any anatomic stress such as epiretinal membrane formation or macular edema. However, in most of our reopened cases, no definite cause was evident. Four patients experienced recurrent reopening.

In vivo IL-12 administration induces profound but transient commitment to T helper cell type 1-associated patterns of cytokine and antibody production.

There is much interest in the utility of exogenous IL-12 as a biologic adjuvant in immediate hypersensitivity and infectious or parasitic diseases where the induction of Th1 responses is strongly associated with protective immunity. Using an immediate hypersensitivity model in which C57Bl/6 mice immunized with OVA (alum) normally generate Th2-dominated responses, we examined the ability of rIL-12 to direct and maintain OVA-specific cytokine and Ab responses in a Th1 direction. Exogenous IL-12 administered coincident with OVA immunization stimulated elevated serum IFN-gamma levels, enhanced IFN-gamma synthesis, and inhibited IL-4 synthesis in bulk culture; 80 to 99% inhibited primary Ag-specific serum IgE and IgG1 responses; and 15- to 20-fold enhanced IgG2a synthesis. However, each of these effects was highly transient, as exogenous IL-12, given at levels up to those associated with serious toxicity, failed to have a lasting impact on the OVA-specific T or B cell response. This transience was evident in primary bulk culture cytokine synthesis; in limiting dilution analysis of the frequency of OVA-specific IFN-gamma-, IL-4-, or IL-10-producing CD4 T cells; and, most importantly, in vivo effector responses such as IgE production to secondary and tertiary OVA immunization. The finding that the intense Th1-like phenomena seen following in vivo administration of rIL-12 with this exogenous Ag are highly transient and are not associated with alterations in the allergen-specific CD4 T cell repertoire to Th1-like patterns suggests a need for caution in the enthusiasm for the use of this cytokine as a biologic adjuvant.

Carcinoembryonic antigen as a target for cancer vaccines.

The vaccine approach to cancer therapy is still in the early stages. Since any immune response has a limited capacity, immunotherapeutic manipulations to destroy tumor will probably be most effective against small tumor masses or micrometastases. Consequently, immunotherapy with these CEA cancer vaccines may prove most effective in the adjuvant setting, where disease has been controlled or stabilized with conventional therapies. The development of CEA cancer vaccines involves many parameters, including the appropriate form of the vaccine, i.e., recombinant protein, peptides, vectors etc., the use of classical adjuvants and/or biological adjuvants such as cytokines, and the use of T cell costimulatory molecules. The integration of these parameters into the development of a cancer vaccine will be the challenge for the next decade [33].

170 The regulation of CO-stimulatory molecules on Langerhans cells by various biological response modifiers, e.g., adjuvants and superantigens

170 The regulation of CO-stimulatory molecules on Langerhans cells by various biological response modifiers, e.g., adjuvants and superantigens

Induction of Cytokines and Cytotoxicity against Tumor Cells by Newcastle Disease Virus

The use of NDV as biological adjuvant in vaccines against human cancer is still actual in several clinical treatment protocols. In this study, we have investigated in vitro-effects of Newcastle disease virus (NDV) strain 73-T on isolated mononuclear blood cells and cultured tumor cells. Cellular cytotoxicity of PBMC freshly isolated from healthy donors against tumor cells was enhanced significantly (p < 0.01) after coincubation of NDV with effector cells. NDV failed to enhance cytotoxicity of effector cells when PBMC were stimulated three days with 500 IU recombinant interleukin-2 (rIL-2) per ml prior to coincubation with the virus. No significant enhancement of cellular lysis was seen when only target cells were coincubated with NDV. As shown by depletion of various lymphocyte subsets, NK cells were the predominant mediator of lysis. Enhancement of cytotoxicity correlated with the induction of interferon-alpha (IFN-alpha) in PBMC by NDV. NDV also induced high amounts of tumor necrosis factor-alpha (TNF-alpha) in PBMC. Induction of interferon-gamma (IFN-gamma) was weak. A direct cytopathic effect (CPE) of NDV on different target cells was detected by colorimetric measurement of metabolic cell activity. The human tumor cell lines A-498, A-704, Caki-1, Caki-2, and K-562 and the fibroblast line MRC-5 showed progressive cellular destruction 48 h after infection with NDV, whereas PBMC and Daudi cells remained unaffected during the observation period. The nontransformed monkey kidney cell line CV-1 and the transformed monkey kidney cell line COS-1 were both lysed by NDV with marginal difference in time course of CPE. Our results indicate a reasonable potential of pleiotropic modifications of the immune response against tumors by NDV.

Two-Dimensional Protein Crystals (S-Layers): Fundamentals and Application Potential

ABSTRACTCrystalline cell surface layers (S-layers) represent the outermost cell envelope component in many bacteria. The oblique, square or hexagonal lattices are formed of assemblies of a single protein or glycoprotein species. Isolated S-layer subunits are endowed with the ability to assemble into monomolecular arrays in suspension, on solid surfaces (e.g. metals, polymers, glass, carbon, silicon), at the air/water interface, or on lipid films generated by the Langmuir Blodgett technique. S-layer lattices are isoporous structures with functional groups located on the surface and in the pores in an identical position and orientation. These characteristic features have led to applications of S-layers as (i) ultrafiltration membranes with pores of identical size and morphology and a broad chemical modification potential, (ii) matrices for the controlled and reproducible immobilization of functional macromolecules, as required for affinity and enzyme membranes, affinity microcarriers and biosensors, (iii) carriers for Langmuir-Blodgett films and reconstituted biological membranes, (iv) immobilization matrices and adjuvants for weakly immunogenic antigens and haptens and (v) patterning elements in molecular nanotechnology.

Skin grafting across the H-Y barrier and the effect of biological adjuvants

The results of clinical cancer trials which use biological adjuvants as therapeutic agents have been encouraging but variable, since little is known of the mechanism of action of adjuvants and their method of use has not been rationalized. Skin grafting across the murine H-Y barrier was investigated for its suitability as a laboratory system to analyse the immunological effects of these adjuvants. Background work revealed that male-female skin grafts in strain C57 Bl mice provide consistent control data and are susceptible to immunological manipulation, including the arousal of a second set rejection response, sensitization by prior injection of male spleen cells, adoption of immunity by spleen cell transfer and the demonstration of enhancement occurring with passive transfer of anti-H-Y antisera. It was also demonstrated that the Y antigen is present on organs other than skin. The effect of biological adjuvants in this system varied with the circumstance under which the adjuvant was administered. For instance, B.C.G. led to shortened skin graft survival if given 10 days before grafting, but if given after grafting, it actually had an enhancing effect. The B.C.G. effect was also more marked depending on the dose used. Corynebacterium parvum also led to prolonged graft survival if given 10 days after grafting, but had an immunopotentiating effect if given before grafting or within 4 days after grafting. C. parvum showed no variation of effect in the dosage range investigated. Thus, skin grafting across the H-Y barrier appears to be useful for investigating adjuvants. These adjuvants appear to be immunopotentiating under certain circumstances, but may also have an enhancing effect, depending on treatment conditions. Further experimental study of the effects of these agents is mandatory before clinical studies are undertaken on non-terminal cancer patients.