Nové imunologické adjuvanty v protichřipkových vakcínách - přínosy a rizika

V rámci pandemických plánů WHO se počítá s aktivní imunizací světové veřejnosti proti prasečí chřipce (kmen H1N1), přičemž vzhledem k časové náročnosti přípravy dostatečného množství dávek vakcín doporučuje WHO nově použít adjuvanty na bázi skvalenu.¹ Jedná se o emulzi minerálního oleje ve vodě (skvalenového nebo jiného) , který zvyšuje reakci imunitního systému na vakcínu (co do výše i šířky imunitní odpovědi) a dovoluje použít řádově menší množství antigenu k vyvolání imunitní odpovědi.^{2, 3, 4, 5} Vyhlášení šestého pandemického stupně přitom umožňuje zkrácení schvalovací procedury dosud neregistrovaných vakcín.

Nejstarším olejovým adjuvantem je Freundovo adjuvans, které bylo objeveno v roce 1930 a vykazuje výraznou imunogenitu⁵, ovšem z důvodu toxicity se používá pouze v experimentech na zvířatech, nekompletní Freundovo adjuvans je rovněž patentován pro použití ve veterinární medicíně. V humánní medicíně byl zatím omezeně používán (viz dále) AS03 (resp. MF-59) - jedná se o směs skvalenu a tokoferolu ve vodě s podporou detergentu Tween 80 ve formě submikronové nanoemulze (velikost částic je podobná velikosti pórů membrán). Mechanismus účinku není zcela objasněn.^5,6

Nyní je plánováno použití MF-59 v protipandemických vakcínách (výrobci Baxter a Novartis) navzdory tomu, že vakcíny obsahující skvalenové adjuvanty jsou podezřelé z vyvolání Syndromu války v Zálivu (Gulf War Syndrome)⁷, onemocnění, kterým trpí velká část veteránů z války v Perském zálivu v roce 1991, kteří byli imunizováni nelicencovanou vakcínou proti antraxu a pertusi obsahující skvalenové adjuvanty.⁸ GWS trpí rovněž očkovaní vojáci, kteří nebyli nasazeni v Perském zálivu. Přes 20 000 vojáků muselo být hospitalizováno po vakcinaci mezi roky 1998-2000.⁹ Možným spojením mezi GWS a vakcinací se zabývá pouze několik málo vědeckých článků.^{10,11,12,13,14,15,16,17}

Rovněž mechanismus případných toxických účinků není objasněn. Předpokládá se například posun imunitní odpovědi k Th-2 profilu jako následek vytvoření dlouhodobě perzistentní zánětlivé léze v mízních uzlinách v blízkosti podání vakcíny, vedoucí ke stálé stimulaci imunitního systému. Dá se předpokládat, že GWS je autoimunním onemocněním. Je prokázáno, že pouhé i.p. resp. i.v. podání určitých minerálních olejů laboratorním zvířatům vede k rozvoji onemocnění na autoimunním podkladě (je popsána tzv. adjuvant disease, onemocnění z adjuvantů).^20,21,22,23

Existují studie, které mají dokládat vysokou účinnost a bezpečnost těchto adjuvantů vakcín^24,25, tyto studie ovšem mají řadu nedostatků (příliš krátká doba sledování pacientů, problematické dokazování vztahu příčina-důsledek u nežádoucích účinků atd.). Skutečnost, že látky podávané jako adjuvanty jsou podány v příliš malém množství, než aby měli přímé toxické účinky, může být nepodstatná, připustíme-li, že se zde uplatňují mechanismy imunologické. Již samotná schopnost adjuvantu být nosným médiem pro aktivní komponentu vakcíny, a schopnost submikronové emulze přecházet přes řadu tělních bariér (hematoencefalická bariéra, cerebrospinální bariéra, bariéra krev-varle, placenta).

Poznámka. Autor článku není lékařem, k sepsání článku jej vedla snaha otevřít diskusi o bezpečnosti protipandemických vakcín, která by byla alespoň částečně založena na odborných základech.

Odkazy

• 1- http://www.who.int/csr/disease/swineflu/notes/h1n1_vaccine_20090713/en/index.html
• 2- http://www.gdk-cds.ch/fileadmin/vks/Bilder/H1N1_vaccine_strategy_KA_16.07.2009_D.Koch_e.pdf

• 3 - Vaccine adjuvants: the dream becomes real.

  Tagliabue A, Rappuoli R.

  ALTA R&D in Biotechnology, Siena, Italy. tagliabue@altaweb.eu

  Abstract

  After about 70 years two new adjuvants have been approved for human vaccines. The first is MF59 developed by the ex-Chiron now Novartis Vaccines and it consists in an oil-in-water emulsion, comprising a low content of biodegradable squalene oil (4.3%) as the dispersed phase, which is stabilized by two non-ionic surfactants (Tween 80 and Span 85), and a low ionic strength citrate buffer as the continuous phase. The second one, defined as AS04, has been developed by GSK Biologics and consists in 3-0-desacyl-4'-monophosphoryl lipid A (MPL) that comes from the cell wall LPS of Gram-negative Salmonella minnesota R595 and is detoxified by mild hydrolytic treatment and purification. It is absorbed on aluminum hydroxide or aluminum phosphate. Thus, new molecules are available to improve the immune response to vaccines also in humans: this is the beginning of a new era in vaccinology.

  Hum Vaccin. 2008 Sep-Oct;4(5):347-9. Epub 2008 Sep 16. Links

• 4 - Enhancement of potent antibody and T-cell responses by a single-dose, novel nanoemulsion-formulated pandemic influenza vaccine.

  Huang MH, Huang CY, Lin SC, Chen JH, Ku CC, Chou AH, Liu SJ, Chen HW, Chong P, Leng CH.

  Vaccine Research and Development Center, National Health Research Institutes, Taiwan, ROC.

  Abstract

  Vaccine shortages are a major obstacle to influenza pandemic preparedness. Increasing vaccine efficiency provides a potentially effective way to overcome this problem. Specifically, using single-dose immunization to induce protective immunity is an attractive approach to emergency/massive vaccination. In this report, we propose a novel nanoemulsion comprised of the bioresorbable polymer, Span 85, and squalene forming a ready-to-use adjuvant, called PELC. After formulation with PELC, inactivated H5N1 virus was intramuscularly administered to mice via a single injection. The data demonstrate that inactivated virus containing 0.5microg hemagglutinin (HA) and formulated with PELC induced more potent antigen-specific antibodies, hemagglutination inhibition, and virus neutralization than non-adjuvanted inactivated virus containing 5microg HA. In addition, T-cell proliferative responses, as well as interferon-gamma (IFN-gamma) and interleukin-4 (IL-4) secretion were significantly enhanced after immunization with PELC-adjuvanted inactivated virus. These results indicate that PELC can be used for effective single-dose immunization and could thus play an important role in influenza pandemic preparedness.

  Microbes Infect. 2009 May-Jun;11(6-7):654-60. Epub 2009 Apr 1. Links

• 5- O adjuvantech obecně

  http://en.wikipedia.org/wiki/Immunologic_adjuvant

  O Freundovu adjuvantu

  http://en.wikipedia.org/wiki/Freund_adjuvant

• 6 - Molecular and cellular signatures of human vaccine adjuvants.

  Mosca F, Tritto E, Muzzi A, Monaci E, Bagnoli F, Iavarone C, O'Hagan D, Rappuoli R, De Gregorio E.

  Novartis Vaccines and Diagnostics, 53100 Siena, Italy.

  Abstract

  Oil-in-water emulsions are potent human adjuvants used for effective pandemic influenza vaccines; however, their mechanism of action is still unknown. By combining microarray and immunofluorescence analysis, we monitored the effects of the adjuvants MF59 oil-in-water emulsion, CpG, and alum in the mouse muscle. MF59 induced a time-dependent change in the expression of 891 genes, whereas CpG and alum regulated 387 and 312 genes, respectively. All adjuvants modulated a common set of 168 genes and promoted antigen-presenting cell recruitment. MF59 was the stronger inducer of cytokines, cytokine receptors, adhesion molecules involved in leukocyte migration, and antigen-presentation genes. In addition, MF59 triggered a more rapid influx of CD11b+ blood cells compared with other adjuvants. The early biomarkers selected by microarray, JunB and Ptx3, were used to identify skeletal muscle as a direct target of MF59. We propose that oil-in-water emulsions are the most efficient human vaccine adjuvants, because they induce an early and strong immunocompetent environment at the injection site by targeting muscle cells.

  Proc Natl Acad Sci U S A. 2008 Jul 29;105(30):10501-6. Epub 2008 Jul 23. Links

• 7 - http://en.wikipedia.org/wiki/Gulf_War_syndrome
• 8- http://orig.clarionledger.com/news/0105/16/m11.html
• 9 - http://www.vermontguardian.com/dailies/122005/1220.shtml

• 10 - Gulf war syndrome: could it be triggered by biological warfare-vaccines using pertussis as an adjuvant?

  Tournier JN, Jouan A, Mathieu J, Drouet E.

  Département de biologie des agents transmissibles, CRSSA, La Tronche, France. j.tournier@eudoramail.com

  Abstract

  Several recent epidemiological studies have shown that vaccinations against biological warfare using pertussis as an adjuvant were associated with the Gulf war syndrome. If such epidemiological findings are confirmed, we propose that the use of pertussis as an adjuvant could trigger neurodegeneration through induction of interleukin-1beta secretion in the brain. In turn, neuronal lesions may be sustained by stress or neurotoxic chemical combinations. Particular susceptibility for IL-1beta secretion and potential distant neuronal damage could provide an explanation for the diversity of the symptoms observed on veterans. Copyright 2002 Elsevier Science Ltd. All rights reserved.

  Med Hypotheses. 2002 Apr;58(4):291-2. Links

• 11 - Aluminum adjuvant linked to Gulf War illness induces motor neuron death in mice.

  Petrik MS, Wong MC, Tabata RC, Garry RF, Shaw CA.

  Department of Ophthalmology and Program in Neuroscience, University of British Columbia, Vancouver, British Columbia, Canada. mspetrik@interchange.ubc.ca

  Abstract

  Gulf War illness (GWI) affects a significant percentage of veterans of the 1991 conflict, but its origin remains unknown. Associated with some cases of GWI are increased incidences of amyotrophic lateral sclerosis and other neurological disorders. Whereas many environmental factors have been linked to GWI, the role of the anthrax vaccine has come under increasing scrutiny. Among the vaccine's potentially toxic components are the adjuvants aluminum hydroxide and squalene. To examine whether these compounds might contribute to neuronal deficits associated with GWI, an animal model for examining the potential neurological impact of aluminum hydroxide, squalene, or aluminum hydroxide combined with squalene was developed. Young, male colony CD-1 mice were injected with the adjuvants at doses equivalent to those given to US military service personnel. All mice were subjected to a battery of motor and cognitive-behavioral tests over a 6-mo period postinjections. Following sacrifice, central nervous system tissues were examined using immunohistochemistry for evidence of inflammation and cell death. Behavioral testing showed motor deficits in the aluminum treatment group that expressed as a progressive decrease in strength measured by the wire-mesh hang test (final deficit at 24 wk; about 50%). Significant cognitive deficits in water-maze learning were observed in the combined aluminum and squalene group (4.3 errors per trial) compared with the controls (0.2 errors per trial) after 20 wk. Apoptotic neurons were identified in aluminum-injected animals that showed significantly increased activated caspase-3 labeling in lumbar spinal cord (255%) and primary motor cortex (192%) compared with the controls. Aluminum-treated groups also showed significant motor neuron loss (35%) and increased numbers of astrocytes (350%) in the lumbar spinal cord. The findings suggest a possible role for the aluminum adjuvant in some neurological features associated with GWI and possibly an additional role for the combination of adjuvants.

  Neuromolecular Med. 2007;9(1):83-100.Links

• 12 - Lessons from macrophagic myofasciitis: towards definition of a vaccine adjuvant-related syndrome

  Gherardi RK.

  Groupe Nerf-Muscle, Département de Pathologie, Hôpital Henri Mondor, Créteil. romain.gherardi@hmn.ap-hop-paris.fr

  Abstract

  Macrophagic myofasciitis is a condition first reported in 1998, which cause remained obscure until 2001. Over 200 definite cases have been identified in France, and isolated cases have been recorded in other countries. The condition manifests by diffuse myalgias and chronic fatigue, forming a syndrome that meets both Center for Disease Control and Oxford criteria for the so-called chronic fatigue syndrome in about half of patients. One third of patients develop an autoimmune disease, such as multiple sclerosis. Even in the absence of overt autoimmune disease they commonly show subtle signs of chronic immune stimulation, and most of them are of the HLADRB1*01 group, a phenotype at risk to develop polymyalgia rheumatica and rheumatoid arthritis. Macrophagic myofasciitis is characterized by a stereotyped and immunologically active lesion at deltoid muscle biopsy. Electron microscopy, microanalytical studies, experimental procedures, and an epidemiological study recently demonstrated that the lesion is due to persistence for years at site of injection of an aluminum adjuvant used in vaccines against hepatitis B virus, hepatitis A virus, and tetanus toxoid. Aluminum hydroxide is known to potently stimulate the immune system and to shift immune responses towards a Th-2 profile. It is plausible that persistent systemic immune activation that fails to switch off represents the pathophysiologic basis of chronic fatigue syndrome associated with macrophagic myofasciitis, similarly to what happens in patients with post-infectious chronic fatigue and possibly idiopathic chronic fatigue syndrome. Therefore, the WHO recommended an epidemiological survey, currently conducted by the French agency AFSSAPS, aimed at substantiating the possible link between the focal macrophagic myofasciitis lesion (or previous immunization with aluminium-containing vaccines) and systemic symptoms. Interestingly, special emphasis has been put on Th-2 biased immune responses as a possible explanation of chronic fatigue and associated manifestations known as the Gulf war syndrome. Results concerning macrophagic myofasciitis may well open new avenues for etiologic investigation of this syndrome. Indeed, both type and structure of symptoms are strikingly similar in Gulf war veterans and patients with macrophagic myofasciitis. Multiple vaccinations performed over a short period of time in the Persian gulf area have been recognized as the main risk factor for Gulf War syndrome. Moreover, the war vaccine against anthrax, which is administered in a 6-shot regimen and seems to be crucially involved, is adjuvanted by aluminium hydroxide and, possibly, squalene, another Th-2 adjuvant. If safety concerns about long-term effects of aluminium hydroxide are confirmed it will become mandatory to propose novel and alternative vaccine adjuvants to rescue vaccine-based strategies and the enormous benefit for public health they provide worldwide.

  Rev Neurol (Paris). 2003 Feb;159(2):162-4. Links

• 13 - Antibodies to squalene in recipients of anthrax vaccine.

  Asa PB, Wilson RB, Garry RF.

  Department of Microbiology, Tulane University Medical School, New Orleans, Louisiana 70112, USA.

  Abstract

  We previously reported that antibodies to squalene, an experimental vaccine adjuvant, are present in persons with symptoms consistent with Gulf War Syndrome (GWS) (P. B. Asa et al., Exp. Mol. Pathol 68, 196-197, 2000). The United States Department of Defense initiated the Anthrax Vaccine Immunization Program (AVIP) in 1997 to immunize 2.4 million military personnel. Because adverse reactions in vaccinated personnel were similar to symptoms of GWS, we tested AVIP participants for anti-squalene antibodies (ASA). In a pilot study, 6 of 6 vaccine recipients with GWS-like symptoms were positive for ASA. In a larger blinded study, only 32% (8/25) of AVIP personnel compared to 15.7% (3/19) of controls were positive (P > 0.05). Further analysis revealed that ASA were associated with specific lots of vaccine. The incidence of ASA in personnel in the blinded study receiving these lots was 47% (8/17) compared to an incidence of 0% (0/8; P < 0.025) of the AVIP participants receiving other lots of vaccine. Analysis of additional personnel revealed that in all but one case (19/20; 95%), ASA were restricted to personnel immunized with lots of vaccine known to contain squalene. Except for one symptomatic individual, positive clinical findings in 17 ASA-negative personnel were restricted to 4 individuals receiving vaccine from lots containing squalene. ASA were not present prior to vaccination in preimmunization sera available from 4 AVIP personnel. Three of these individuals became ASA positive after vaccination. These results suggest that the production of ASA in GWS patients is linked to the presence of squalene in certain lots of anthrax vaccine.

  Exp Mol Pathol. 2002 Aug;73(1):19-27. Links

• 13 - Vaccines with the MF59 adjuvant do not stimulate antibody responses against squalene.

  Del Giudice G, Fragapane E, Bugarini R, Hora M, Henriksson T, Palla E, O'hagan D, Donnelly J, Rappuoli R, Podda A.

  Research Center, Novartis Vaccines, Via Fiorentina 1, 53100 Siena, Italy. giuseppe_del_giudice@chiron.com

  Abstract

  Squalene is a naturally occurring oil which has been used in the development of vaccine adjuvants, such as the oil-in-water emulsion MF59. In past years, by use of noncontrolled and nonvalidated assays, a claim was made that antisqualene antibodies were detectable in the sera of individuals with the so-called Gulf War syndrome. Using a validated enzyme-linked immunosorbent assay for the quantitation of immunoglobulin G (IgG) and IgM antibodies against squalene, we demonstrated that antisqualene antibodies are frequently detectable at very low titers in the sera of subjects who were never immunized with vaccines containing squalene. More importantly, vaccination with a subunit influenza vaccine with the MF59 adjuvant neither induced antisqualene antibodies nor enhanced preexisting antisqualene antibody titers. In conclusion, antisqualene antibodies are not increased by immunization with vaccines with the MF59 adjuvant. These data extend the safety profile of the MF59 emulsion adjuvant.

  Clin Vaccine Immunol. 2006 Sep;13(9):1010-3. Links

• 14 - Gulf War syndrome: is it due to a systemic shift in cytokine balance towards a Th2 profile?

  Rook GA, Zumla A.

  Department of Bacteriology, University College London Medical School, UK.

  Abstract

  The symptoms of Gulf War syndrome are compatible with the hypothesis that the immune system of affected individuals is biased towards a Th2-cytokine pattern. Factors that could lead to a Th2 shift among Gulf War veterans include exposure to multiple Th2-inducing vaccinations under stressful circumstances and the way in which such vaccinations were administered, which would be expected to maximise Th2 immunogenicity. These factors may have led to a long-term systemic shift towards a Th2-cytokine balance and to mood changes related to the immunoendocrine state. Other vaccines that lead to similar long-term, non-specific shifts in cytokine balance are well-established. If our hypothesis is proven, treatment may be possible with regimens that induce a systemic Th1 bias.

  Lancet. 1997 Jun 21;349(9068):1831-3. Links

• 15 - Beef allergy and the Persian Gulf syndrome.

  Hollander DH.

  Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA.

  Abstract

  It is suggested that the Persian Gulf Syndrome (PGS) is caused by beef allergy. In the first symptomless phase, as a result of an energetic US Army immunizing program, using sera with adjuvants to produce detectable antibody levels, the subjects not only developed immunity to the targeted substances, but also became sensitized to one or more of the other substances in the immunizing sera, and specifically to beef protein. The subjects remained healthy while in the war zone on a restricted diet essentially free from beef, but developed PGS after they came home, and were again able to obtain steaks and hamburgers.

  Med Hypotheses. 1995 Sep;45(3):221-2. Links

• 16 - Infection and vaccination in chronic fatigue syndrome: myth or reality?

  Appel S, Chapman J, Shoenfeld Y.

  Department of Neurology, Sheba Medical Center, Tel Hashomer, Israel.

  Abstract

  Chronic fatigue syndrome (CFS) is characterized by severe disabling fatigue lasting for more than 6 months associated with physical and mental disturbances such as headache, arthralgia, myalgia, memory impairment, sore throat and tender lymph nodes. The exact pathogenesis is still unknown. Several models were proposed to explain its etiology including chronic infection, endocrine dysfunction, autonomic imbalance, depression, decreased immunity states and an aberrant reaction to infection. No convincing evidence was found to support any of the suggested pathogenic mechanisms. The current concept is that CFS pathogenesis is a multi factorial condition in which an infective agent cause an aberrant immune response characterized by a shift to Th-2 dominant response. When the response fails to be switched-off, a chronic immune activation occurs and clinically expressed as the symptomatology of CFS. Vaccinations are used in order to stimulate the immune system to induce a persistent immunity against the favorable antigens. Several syndromes that contain chronic fatigue as one of their symptoms, such as "Gulf war syndrome" and macrophagic myofasciitis were related to vaccinations. Can vaccinations induce the aberrant immune response of CFS? Little is known about this issue. There are some reports on CFS occurring after vaccination, but few prospective and retrospective studies failed to find such an association. A working group of the Canadian Laboratory Center for Disease Control (LCDC) that was founded in order to examine the suspected association between CFS and vaccinations concluded that there is no evidence that relates CFS to vaccination. Further studies are requested to examine this issue since it is very conceivable that if infection can lead to CFS, vaccination may also lead to it in the same immune-mediated pathogenesis.

  Autoimmunity. 2007 Feb;40(1):48-53. Links

• 17 - Autoimmunity induced by adjuvant hydrocarbon oil components of vaccine.

  Kuroda Y, Nacionales DC, Akaogi J, Reeves WH, Satoh M.

  Division of Rheumatology and Clinical Immunology, Department of Medicine, University of Florida, ARB-R2-156, 1600 SW Archer Road, P.O. Box 100221 Gainesville, FL 32610-0221, USA.

  Abstract

  Adjuvant oils such as Bayol F (Incomplete Freund's adjuvant: IFA) and squalene (MF59) have been used in human and veterinary vaccines despite poor understanding of their mechanisms of action. Several reports suggest an association of vaccination and various autoimmune diseases, however, few were confirmed epidemiologically and the risk of vaccination for autoimmune diseases has been considered minimal. Microbial components, not the adjuvant components, are considered to be of primary importance for adverse effects of vaccines. We have reported that a single intraperitoneal injection of the adjuvant oils pristane, IFA or squalene induces lupus-related autoantibodies to nRNP/Sm and -Su in non-autoimmune BALB/c mice. Induction of these autoantibodies appeared to be associated with the hydrocarbon's ability to induce IL-12, IL-6, and TNF-alpha, suggesting a relationship with hydrocarbon's adjuvanticity. Whether this is relevant in human vaccination is a difficult issue due to the complex effects of vaccines and the fact that immunotoxicological effects vary depending on species, route, dose, and duration of administration. Nevertheless, the potential of adjuvant hydrocarbon oils to induce autoimmunity has implications in the use of oil adjuvants in human and veterinary vaccines as well as basic research. Copyright 2004 Elsevier SAS Biomed Pharmacother. 2004 Jun;58(5):325-37. Links

• 18 - Human adjuvant disease: presentation as a multiple sclerosis-like syndrome.

  Shoaib BO, Patten BM.

  CompreCare Clinics, Houston, TX 77030, USA.

  Abstract

  Twenty-six women had a systemic disease with central nervous system (CNS) involvement at a mean age of 39.2 years (range, 23 to 64 years) after receiving silicone breast implants (n = 25) or silicone fluid injections into breasts (n = 1). The median latency period between breast surgery and onset of symptoms was 5.71 years (range, 3 months to 15 years). All patients had evidence of disseminated CNS lesions; 20 patients also had evidence of peripheral neuropathy. Additional problems included myalgia (n = 24), joint stiffness (n = 23), arthralgia (n = 22), sicca complex (dry eyes and dry mouth) (n = 19), headache (n = 16), skin rash (n = 15), joint swelling (n = 14), Raynaud's phenomena (n = 14), fever (n = 13), hair loss (n = 12), allergies (n = 11), sensitivity to sunlight (n = 10), and lymphadenopathy (n = 9). Magnetic resonance imaging brain scans were abnormal in 22 of 26 patients (21, white matter lesions; 1, ischemic lesions; 4, cerebral atrophy). Spinal tap revealed oligoclonal bands in 18 of 23 patients. Visual evoked responses were delayed in 14 of 23 patients, and autodirected antibodies were detected in 16 of 26. Sural nerve biopsy results showed loss of myelinated fibers in 15 of 15. Seventeen of 24 patients (71%) who had implant removal were found to have grossly ruptured implants. We believe our patients had a new syndrome triggered by the foreign material in their body. This syndrome appears as a systemic inflammatory autoimmune disease with central nervous system involvement resembling multiple sclerosis.

• 19 - http://www.autoimmune.com/GWSGen.html

• 20 - Immunoregulatory role of CD1d in the hydrocarbon oil-induced model of lupus nephritis.

  Yang JQ, Singh AK, Wilson MT, Satoh M, Stanic AK, Park JJ, Hong S, Gadola SD, Mizutani A, Kakumanu SR, Reeves WH, Cerundolo V, Joyce S, Van Kaer L, Singh RR.

  Department of Internal Medicine, Autoimmunity and Tolerance Laboratory, University of Cincinnati College of Medicine, Cincinnati, OH 45267, USA.

  Abstract

  Systemic lupus erythematosus (SLE) is a systemic autoimmune disease that is accompanied by the emergence of autoreactive T cells and a reduction in regulatory T cells. Humans and mice with SLE have reduced numbers of CD1d-restricted NK T cells, suggesting a role for these cells in the regulation of SLE. In this study, we show that CD1d deficiency exacerbates lupus nephritis induced by the hydrocarbon oil pristane. This exacerbation in disease is associated with: 1) reduced TNF-alpha and IL-4 production by T cells, especially during the disease induction phase; and 2) expansion of marginal zone B cells. Strikingly, inoculation of pristane in wild-type mice resulted in reduced numbers and/or functions of NK T cells and CD1d-expressing dendritic cells. These findings suggest that CD1d may play an immunoregulatory role in the development of lupus in the pristane-induced model.

  J Immunol. 2003 Aug 15;171(4):2142-53. Links

• 21 - Distinctive patterns of autoimmune response induced by different types of mineral oil.

  Kuroda Y, Akaogi J, Nacionales DC, Wasdo SC, Szabo NJ, Reeves WH, Satoh M.

  Division of Rheumatology and Clinical Immunology, Department of Medicine, University of Florida, Gainesville, Florida 32610-0221, USA.

  Abstract

  Although mineral oils are generally considered nontoxic and have a long history of use in humans, the mineral oil Bayol F (incomplete Freund's adjuvant, IFA) and certain mineral oil components (squalene and n-hexadecane) induce lupus-related anti-nRNP/Sm or -Su autoantibodies in nonautoimmune mice. In the present study, we investigated whether medicinal mineral oils can induce other types of autoantibodies and whether structural features of hydrocarbons influence autoantibody specificity. Female 3-month-old BALB/c (16-45/group) mice each received an i.p. injection of pristane (C19), squalene (C30), IFA, three medicinal mineral oils (MO-F, MO-HT, MO-S), or PBS. Sera were tested for autoantibodies and immunoglobulin levels. Hydrocarbons were analyzed by gas chromatography/mass spectrometry. IFA contained mainly C15-C25 hydrocarbons, whereas MO-HT and MO-S contained C20-C40, and MO-F contained C15-C40. Pristane and n-hexadecane were found in IFA (0.17% and 0.10% w/v, respectively) and MOs (0.0026-0.027%). At 3 months, pristane and IFA induced mainly IgG2a, squalene IgG1, and MOs IgG3 and IgM in sera. Anti-cytoplasmic antibodies were common in mice treated with MO-F, as well as those treated with pristane, squalene, and IFA. Anti-ssDNA and -chromatin antibodies were higher in MO-F and MO-S than in untreated/PBS, squalene-, or IFA-treated mice, suggesting that there is variability in the induction of anti-nRNP/Sm versus -chromatin/DNA antibodies. The preferential induction of anti-chromatin/ssDNA antibodies without anti-nRNP/Sm/Su by MO-S and MO-F is consistent with the idea that different types of autoantibodies are regulated differently. Induction of autoantibodies by mineral oils considered nontoxic also may have pathogenetic implications in human autoimmune diseases.

  Toxicol Sci. 2004 Apr;78(2):222-8. Epub 2004 Jan 12. Links

• 22 - Induction of lupus-related specific autoantibodies by non-specific inflammation caused by an intraperitoneal injection of n-hexadecane in BALB/c mice.

  Kuroda Y, Ono N, Akaogi J, Nacionales DC, Yamasaki Y, Barker TT, Reeves WH, Satoh M.

  Division of Rheumatology and Clinical Immunology, Department of Medicine, ARB-R2-156, 1600 SW Archer Road, Box 100221, Gainesville, FL 32610-0221, USA.

  Abstract

  A single intraperitoneal (i.p.) injection of pristane, incomplete Freund's adjuvant (IFA), or the adjuvant oil squalene, but not high molecular weight medicinal mineral oils, induces lupus-related autoantibodies to nRNP/Sm and -Su in non-autoimmune strains of mice. This ability appears to be associated with the low molecular weight and adjuvanticity of hydrocarbon. n-Hexadecane (C(16)H(34)), which is present in petroleum, has adjuvant activity and induces arthritis in rodents like other lupus-inducing oils. In addition to dietary exposure to n-hexadecane in mineral oils, exposure also occurs via inhalation of oil mist, jet fuel, or diesel exhaust or by absorption through the skin. Since n-hexadecane is a low molecular weight adjuvant hydrocarbon oil similar to other lupus-inducing hydrocarbons, the present study examined whether it can also induce lupus-related autoantibodies in mice. Female BALB/cJ mice received a single i.p. injection of 0.5 ml of n-hexadecane, pristane, or saline (control). Pathology and serology (immunoglobulin levels, autoantibodies by immunofluorescence, immunoprecipitation, and ELISA) were examined 3 months later. Unexpectedly, all n-hexadecane-treated mice, but none in the other groups, developed inflammatory ascites within 2.5 months. n-Hexadecane induced hypergammaglobulinemia (IgG1, IgG2a), antinuclear (titer>1:160, 67%) and -cytoplasmic antibodies (58%) and autoantibodies to nRNP/Sm (25%), Su (33%), ssDNA (83%), and chromatin (100%). Therefore, non-specific inflammation caused by n-hexadecane resulted in the production of a limited set of specific autoantibodies. These previously unrecognized immunological effects of n-hexadecane may have implications in monitoring human exposure to hydrocarbons and in the pathogenesis of autoimmune diseases.

  Toxicology. 2006 Feb 1;218(2-3):186-96. Epub 2005 Nov 23. Links

• 23 - Rats made congenic for Oia3 on chromosome 10 become susceptible to squalene-induced arthritis.

  Holm BC, Xu HW, Jacobsson L, Larsson A, Luthman H, Lorentzen JC.

  Center for Molecular Medicine, Department of Medicine, Unit of Rheumatology, Karolinska Institutet, S-17176 Stockholm, Sweden. barbro.holm@cmm.ki.se

  Abstract

  Several quantitative trait loci (QTLs) regulating the risk of experimental arthritis have been identified by genome-wide linkage analyses, but only the MHC has thus far been reported to transfer arthritis susceptibility in congenic animals. We have produced a congenic strain for Oia3, a genetic factor originally identified as an oil-induced arthritis (OIA) QTL in arthritis-prone DA rats. A 46 cM telomeric region of chromosome 10 encompassing Oia3 was transferred from DA rats to MHC-identical but minutely arthritis-susceptible LEW.1AV1 rats by selective breeding. Arthritis development was provoked in Oia3-congenic rats by intradermal injection of different adjuvant oils. One successful arthritis trigger was squalene, which is approved for vaccinations in humans and has been implicated in Gulf War syndrome. The endogenous cholesterol precursor squalene induced T cell infiltration into joints and macroscopic arthritis in Oia3-congenic rats and DA rats, whereas LEW.1AV1 rats were almost resistant. Arthritis onset, approximately 14 days post-injection, coincided with arrested body-weight gain and increased plasma levels of the inflammation markers fibrinogen and alpha 1-acid glycoprotein. Congenic rats displayed intermediate phenotypes compared with the two parental strains, and similar to rheumatoid arthritis in humans, female preponderance was observed in Oia3-congenic rats. Finally, recombinant rat strains were constructed and were used to map a susceptibility gene(s) in females to a telomeric 4--19 cM Oia3 subregion. The experimental system described allows transformation of multifactorial arthritis susceptibility into dichotomous phenotypes.

  Hum Mol Genet. 2001 Mar 15;10(6):565-72. Links

• 24 - Safety of MF59 adjuvant.

  Schultze V, D'Agosto V, Wack A, Novicki D, Zorn J, Hennig R.

  Novartis Vaccines and Diagnostics, Marburg, Germany. viola.schultze@novartis.com

  Abstract

  The need to enhance the immunogenicity of purified subunit antigens has prompted the development of new adjuvants. The adjuvant emulsion MF59 has been tested in animals in combination with different antigens and finally evaluated in humans. It was licensed after the successful outcome of preclinical and clinical testing. This paper summarizes the main characteristics of the MF59 adjuvant, including animal testing, clinical experience with various vaccines, and information from current postmarketing surveillance data. This review supports the hypothesis that MF59 is a safe adjuvant for human use.

  Vaccine. 2008 Jun 19;26(26):3209-22. Epub 2008 Apr 21. Links

• 25 - MF59-adjuvanted vaccines for seasonal and pandemic influenza prophylaxis.

  Banzhoff A, Pellegrini M, Del Giudice G, Fragapane E, Groth N, Podda A.

  Novartis Vaccines, Marburg, Germany. angelika.banzhoff@novartis.com

  Abstract

  Influenza is a major cause of worldwide morbidity and mortality through frequent seasonal epidemics and infrequent pandemics. Morbidity and mortality rates from seasonal influenza are highest in the most frail, such as the elderly, those with underlying chronic conditions and very young children. Antigenic mismatch between strains recommended for vaccine formulation and circulating viruses can further reduce vaccine efficacy in these populations. Seasonal influenza vaccines with enhanced, cross-reactive immunogenicity are needed to address these problems and can confer a better immune protection, particularly in seasons were antigenic mismatch occurs. A related issue for vaccine development is the growing threat of pandemic influenza caused by H5N1 avian strains. Vaccines against strains with pandemic potential offer the best approach for reducing the potential impact of a pandemic. However, current non-adjuvanted pre-pandemic vaccines offer suboptimal immunogenicity against H5N1. For both seasonal and pre-pandemic vaccines, the addition of adjuvants may be the best approach for providing enhanced cross-reactive immunogenicity. MF59, the first oil-in-water emulsion licensed as an adjuvant for human use, can enhance vaccine immune responses through multiple mechanisms. A trivalent MF59-adjuvanted seasonal influenza vaccine (Fluad has shown to induce significantly higher immune responses to influenza vaccination in the elderly, compared with non-adjuvanted vaccines, and to provide cross-reactive immunity against divergent influenza strains. Similar results have been generated with a MF59-adjuvanted H5N1 pre-pandemic vaccine, which showed higher and broader immunogenicity compared with non-adjuvanted pre-pandemic vaccines.

  Influenza Other Respi Viruses. 2008 Nov;2(6):243-9.Links