© 1999 Nature America Inc. • http://biotech.nature.com REVIEW Advances in vaccine adjuvants Manmohan Singh* and Derek O’Hagan Chiron Corporation, 5300 Chiron Way, Emeryville, CA 94608. *Corresponding author ([email protected]). Received 8 December 1998; accepted 20 September 1999 Currently, aluminum salts and MF59 are the only vaccine adjuvants approved for human use. With the development of new-generation vaccines (including recombinant subunit and mucosal vaccines) that are less immunogenic, the search for more potent vaccine adjuvants has intensified. Of the novel compounds recently evaluated in human trials, immunostimulatory molecules such as the lipopolysaccharide derived MPL and the saponin derivative QS21 appear most promising, although doubts have been raised as to their safety in humans. Preclinical work with particulate adjuvants, such as the MF59 microemulsion and lipid–particle immune-stimulating complexes (Iscoms), suggest that these molecules are also potent elic- itors of humoral and cellular immune responses. In addition, preclinical data on CpG oligonucleotides appear to be encouraging, particularly with respect to their ability to selectively manipulate immune responses. While all these adjuvants show promise, further work is needed to better define the mecha- nisms of adjuvant action. Ultimately, the development of more potent adjuvants may allow vaccines to be used as therapeutic, rather than prophylactic, agents. Keywords: adjuvant, recombinant vaccine delivery, mucosal delivery, vaccine targeting Vaccines have traditionally consisted of live attenuated pathogens, first introduced more than 50 years ago5. Many experimental adju- whole inactivated organisms, or inactivated toxins. Although these vants have advanced to clinical trials and some have demonstrated http://biotech.nature.com • have proved successful in the past, several drawbacks have limited high potency, but most have proved too toxic for routine clinical their use against more challenging diseases such as hepatitis C or use. For standard prophylactic immunization in healthy individu- AIDS. First, certain live-attenuated vaccines can cause disease in als, only adjuvants that induce minimal side effects will prove immunosuppressed individuals by reverting to a more virulent phe- acceptable. In contrast, for adjuvants that are designed to be used in notype. Second, whole inactivated vaccines (e.g., Bordetella pertus- life-threatening situations (e.g., cancer), the acceptable level of sis) contain reactogenic components that can cause undesirable side adverse events is likely to be higher. Additional issues that are effects. Third, some pathogens are difficult or even impossible to important for adjuvant development include biodegradability, sta- grow in culture (e.g., hepatitis B, hepatitis C, and human papillo- bility, ease of manufacture, cost, and applicability to a wide range of mavirus), making preparation of a vaccine problematic. vaccines. Ideally, for ease of administration and patient compliance, In the past decade, several new approaches to vaccine develop- an adjuvant should allow a vaccine to be given by a mucosal route. 1999 Nature America Inc. © ment have emerged that may have significant advantages over tradi- Examples of adjuvants that have been evaluated in clinical trials are tional approaches. These new approaches include recombinant pro- shown in Table 1. Although the mechanisms of action of adjuvants tein subunits, synthetic peptides, and plasmid DNA. Although they are often poorly understood2,5, they can be broadly classified on the offer advantages such as reduced toxicity, they are poorly immuno- basis of their principal mode of action. However, these classifica- genic when administered alone. This is particularly true for vaccines tions become more arbitrary as immunostimulatory adjuvants are based on recombinant proteins or peptides. Traditional vaccines are added to vaccine delivery systems and adjuvants are combined in heterogeneous and contain many epitopes, some of which can pro- single formulations. vide additional T-cell help or function as adjuvants (e.g., bacterial DNA in whole-cell vaccines). Therefore, a great need exists for The role of adjuvants in vaccine development immunological adjuvants that are potent, safe, and compatible with Adjuvants can be used to improve the immune response to vaccine new-generation vaccines, including DNA vaccines. antigens in many different ways: they can increase the immuno- Immunological adjuvants were originally described by Ramon1 genicity of weak antigens; they can be used to enhance the speed and as “substances used in combination with a specific antigen that duration of the immune response; they can modulate antibody avid- produce more immunity than the antigen alone.” This broad defin- ity, specificity, isotype, or subclass distribution; they can stimulate ition encompasses a very wide range of materials2. However, cell-mediated immunity; they can promote the induction of mucos- despite extensive evaluation over many years, the only adjuvants al immunity; they can enhance immune responses in immunologi- currently approved by the US Food & Drug Administration cally immature, or senescent individuals; they can decrease the dose (Rockville, MD) are aluminum-based mineral salts (generically of antigen and reduce vaccine costs; or they can help overcome anti- called alum). Alum has a good safety record, but comparative stud- gen competition in combination vaccines. ies show that it is a weak adjuvant for antibody induction to pro- The mechanisms of adjuvant action are currently only poorly tein subunits and a poor adjuvant for cell-mediated immunity3. understood, which has hampered rational design of new adjuvant Moreover, alum adjuvants can induce immunoglobulin E (IgE) compounds. Immunization often activates several complex cascades antibody responses and have been associated with some allergic of immune effectors, only some of which are relevant to the induc- reactions in human subjects3,4. tion of an antigen-specific response. Often, ascertaining the exact The most important issue in adjuvant development is safety, effectors enhanced by a particular adjuvant is difficult to clearly which has restricted the development of adjuvants since alum was define in vivo. NATURE BIOTECHNOLOGY VOL 17 NOVEMBER 1999 http://biotech.nature.com 1075 © 1999 Nature America Inc. • http://biotech.nature.com REVIEW Table 1. Selected examples of vaccine adjuvantsa guidance in adjuvant selection. However, adjuvant choice is complicated by commercial constraints and availability. In Mineral salts Aluminum hydroxide* addition, some adjuvants need to be manufactured using spe- Aluminum phosphate* Calcium phosphate* cialized techniques and equipment that are not generally avail- able or suitable for all antigens (e.g., see Iscoms below). Immunostimulatory adjuvants Cytokines (e.g., IL-2, IL-12, GM-CSF) Saponins (e.g., QS21) Immunostimulatory adjuvants MDP derivatives Immunostimulatory adjuvants are thought to exert their effects CpG oligos predominantly at the cytokine level through the activation of LPS MPL MHC molecules, through costimulatory signals, or through polyphosphazenes related intracellular signaling pathways. One class of immunos- Lipid particles Emulsions (e.g., Freund’s, SAF, MF59*) timulatory adjuvants is derived from the lipopolysaccharide Liposomes (LPS) of Gram-negative bacteria. The most extensively evaluat- Virosomes* ed member of this family, monophosphoryl Lipid A (MPL), is Iscoms obtained from Salmonella minnesota. Although the mechanism Cochleates Microparticulate adjuvants PLG microparticles of action of MPL has not been clearly defined, it is likely to be Poloxamer particles similar to LPS. Recent evidence suggests that LPS-mediated Virus-like particles immune activation occurs as a consequence of the interaction Mucosal adjuvants Heat-labile enterotoxin (LT) of LPS with toll-like receptors (TLR), following binding to sol- Cholera toxin (CT) uble serum factors, including CD1410–12. Human TLR2 is Mutant toxins (e.g., LTK63 and LTR72) involved in activation by LPS11,12 and triggers cells to produce Microparticles k Polymerized liposomes proinflammatory cytokines via NF- B, an important element of the response of the innate immune system to infection. aWith the exception of CpGs, cochleates, and polymerized liposomes, all of these adju- In several preclinical studies, MPL has been shown to vants have been evaluated in clinical trials. However, only those marked with an asterisk induce the synthesis and release of cytokines, particularly IFN- are currently included as adjuvants in approved vaccine products for human use. g, which promotes the generation of Th1 responses13,14. As a single adjuvant, MPL does not appear to be very potent for That said, adjuvants can mediate their effects by any of the fol- antibody induction, although it appears effective for the induction lowing mechanisms: increasing cellular infiltration, inflammation, of CD4+ T-cell mediated immunity. Clinically, MPL has often been http://biotech.nature.com • and trafficking to the injection site—particularly for antigen-pre- used in complex formulations, including liposomes and emulsions, senting cells (APCs); promoting the activation state of APCs by and has also been used in adjuvant combinations with alum and upregulating costimulatory signals or major histocompatibility QS2115. One problem with such combination studies is that it is dif- complex (MHC) expression; enhancing antigen presentation;