drug discov today moghimi sm, symonds p, murray jc, hunter ac, debska g and szewczyk a a twostage polyethyleniminemediated cytotoxicity implications for gene transfertherapy mol ther gbadamosi jk, hunter ac and moghimi sm pegylation of microspheres generates a heterogeneous population of particles with differential surface characteristics and biological performance febs lett genetic vaccines a role for liposomes gregory gregoriadis, andrew bacon, brenda mccormack and peter laing introduction prevention of microbial infections by the use of vaccines is a preferred alternative to treatment vaccines have been applied successfully, for example, in pravachol gastric pain the eradication of smallpox as well as against tetanus, diphtheria, whooping cough, polio and measles, thus preventing millions of deaths each year however, vaccines made of attenuated organisms, which mimick pravachol gastric pain natural infections usually without the disease, can be potentially unsafe for instance, there is a pravachol gastric pain risk of reversion during replication of live viruses or even mutation to a more pathogenic state pravachol gastric pain furthermore, with immunocompromised individuals, some of the attenuated viruses may still provoke disease on the other hand, with killed virus vaccines, their extracellular localization and subsequent phagocytosis by professional antigen pravachol gastric pain presenting cells apc or antigenspecific � cells, lead to mhcii class restricted presentation and to t helper cell and humoural immunity however, they do not elicit significant cytotoxic t cell pravachol gastric pain ctl responses moreover, subunit vaccines produced from biological fluids may not be entirely free of infectious pravachol gastric pain agents even with subunit and peptide vaccines produced recombinantly or synthetically and thus considered safe, pravachol gastric pain immune responses are weak and often not of the appropriate kind the great variety of pravachol gastric pain immunological adjuvants that are now available go a long way in rendering subunit and peptide vaccines stronger and more efficient however, more than seventy years after the introduction of aluminium salts as an adjuvant, only two other adjuvants, liposomes and mf, have been approved for use in humans thus, inspite of considerable progress, the road to the ideal vaccine appears as pravachol gastric pain elusive as ever, until recently recent developments have led to a novel and exciting concept, pravachol gastric pain namely de novo production of the required vaccine antigen by the hosts cells in vivo, pravachol gastric pain which promises to revolutionize vaccination especially where vaccines are either ineffective or unavailable the concept pravachol gastric pain entails the direct injection of antigenencoding plasmid dna which, on uptake by cells, localizes to some pravachol gastric pain extent into the nucleus where it transfects the cells episomally the produced antigen is recognized pravachol gastric pain as foreign by the host and is thus subjected to pathways similar to those observed for antigens of internalized viruses but without their disadvantages, leading to protective humoural and cell mediated immunity a series of publications since first established the ability of plasmid dna to pravachol gastric pain induce an immune antibody response to the encoded foreign protein in experiments with dna encoding influenza nucleoprotein, immunity was both humoural and cellmediated, and also protective in mice challenged with the pravachol gastric pain virus, this was the first demonstration of an experimental dna vaccine another observation was the pravachol gastric pain induction of humoural and cellmediated immunity against hiv using plasmids encoding the hiv rev and env proteins similar results were obtained with a gene for the hepatitis � surface antigen hbsag dna immunization was also found pravachol gastric pain to apply in cancer treatment for instance, injection of plasmids pravachol gastric pain encoding tumor antigens promoted immune responses, which were protective in an animal model the concept pravachol gastric pain of dna immunization has now been adopted by vaccinologists worldwide using an ever increasing number pravachol gastric pain of plasmids encoding immunogens from bacterial, viral and parasitic pathogens, and a variety of tumors, pravachol gastric pain in many of these studies, genetic immunization has led to the protection of animals from pravachol gastric pain infection a number of clinical trials for the therapy of, or prophylaxis against, a variety of pravachol gastric pain infections are in progress the dna vaccine a plasmid dna vaccine is usually supercoiled and consists of the gene encoding the vaccine antigen the section of the target pathogen which pravachol gastric pain elicits protective immunity, a promoter sequence which is often derived from cytomegalovirus cmv or rous sarcoma virus rsv, an mrna stability polyadenylation region at the end of the insert, and pravachol gastric pain the plasminogen activator gene which controls the secretion of the recombinant product in addition, there are pravachol gastric pain an origin of replication for the amplification of the plasmid in bacteria, and a gene for antibiotic resistance to select the transformed bacteria immunization procedures with dna vaccines are carried pravachol gastric pain out by the intramuscular and, to a lesser extent, the intraepidermal route other routes include the oral, nasal, vaginal, intravenous, intraperitoneal and subcutaneous routes, intramuscular injection of dna vaccines leads pravachol gastric pain to such types of immunity as ctl, this was unexpected because antigen presentation requires the function pravachol gastric pain of professional apc however, myocytes which were shown to take up the plasmid only to a small extent and with only a fraction of cells participating in the uptake, are pravachol gastric pain not professional apcs although myocytes carry mhc class i molecules and can present endogenously produced pravachol gastric pain viral peptides to the cd cells to induce ctls, they do so inefficiently as they lack vital costimulatory molecules eg the b molecule it is thus difficult to accept that antigen presentation, leading to a ctl response, occurs via myocytes instead, it was reported that ctl responses occur as a result of the transfer of antigenic material between the myocytes and professional apc to some extent in parallel, it could also be that plasmid secreted by pravachol gastric pain the myocytes or as such, is taken up directly by apc infiltrating the injected site pravachol gastric pain such apc would include dendritic cells which will express and present peptides to cd cells following transport to the lymph nodes or spleen on the other hand, cd cells may propecia results months pictures be activated by apcs via mhc class ii presentation of antigen secreted by the myocytes or pravachol gastric pain released from them after their destruction via a tc response and captured by the cells such events would lead to both cellular th and humoural th immunity indeed, it has pravachol gastric pain been shown that dendritic cells are the essential apc involved in immune responses elicited by intramuscularly given dna vaccines dna vaccination via liposomes vaccination with naked dna by the intramuscular route is dependent on the ability of myocytes to take up the plasmid however, some of pravachol gastric pain the dna may also be engulfed by apc infiltrating the site of injection, or in the lymph nodes following migration of the dna to the lymphatics the extent of dna degradation by extracellular deoxyribonucleases is unknown, but depending on the time of its residence interstitially, degradation could be considerable therefore, approaches that protect dna from the extracellular nucleases and promote dna uptake by cells more efficiently, or target it to apc, should contribute to the optimal design of dna vaccines it has been suggested that as apc are a preferred alternative pravachol gastric pain to muscle cells for dna vaccine uptake and expression, liposomes known to be taken up avidly by apc infiltrating the site of injection or in the lymphatics, an event that has pravachol gastric pain been implicated in their immunoadjuvant activity would be a suitable means of delivery of entrapped pravachol gastric pain dna to such cells liposomes would also protect their dna content from deoxyrubonuclease attack moreover, the structural versatility of the system would ensure that its tranfection efficiency is further improved pravachol gastric pain by the judicial choice of its structural characteristics or by the coentrapment of cytokine genes, pravachol gastric pain other adjuvants eg immunostimulatory sequences, or indeed protein antigens see later together with the plasmid vaccine pravachol gastric pain as a number of injectable liposomebased drug formulations, including vaccines against hepatitis a and influenza, pravachol gastric pain have been already licensed for clinical use, acceptance of the system clinically would be less problematic than with other systems that are still at an experimental stage procedure for the entrapment of plasmid dna into liposomes ready to use mlv, =nm dna, antigen or both pravachol gastric pain entrapped over entrapment yield a variety, of plasmid dnas have been quantitatively entrapped into liposomes by pravachol gastric pain a mild dehydrationrehydration procedure the procedure fig consists of mixing preformed small unilamellar vesicles suv pravachol gastric pain with a solution of the dna destined for entrapment, freezedrying of the mixture, followed by pravachol gastric pain controlled rehydration of the formed powder, and centrifugation to remove non entrapped material formed liposomes pravachol gastric pain are multilamellar however, when an appropriate amount of sucrose is added to the suv and pravachol gastric pain dna mixture prior to dehydration, the resulting liposomes are much smaller about nm in diameter as expectcd, dna incorporation values, were higher up to of the amount used when a cationic lipid was present in the bilayers no apparent relationship was observed between amount of dna used g and the values of incorporation for the compositions and lipid mass used, the pravachol gastric pain possibility that dna was not entrapped within the bilayers of cationic liposomes, but was rather complexed with their surface as suggested by the high fig entrapment of dna andor protein into pravachol gastric pain cationic liposomes the procedure entails mixing up empty suv with the solutes destined for entrapment and subsequent dehydration on rehydration, most of the solutes is recovered entrapped within the generated pravachol gastric pain multilamellar liposomes liposomes suv = rim dna, antigen or both naked d laaxea dna q pravachol gastric pain m complexed d � naked d complexed d fig gel electrophoresis of a mixture of pravachol gastric pain cationic suv and prccmv hbs before com plexed dna and after entrapped dna dehydrationrehydration of the mixture !?