Safety, immunogenicity, and efficacy of the ML29 reassortant vaccine for Lassa fever in small non-human primates☆
Introduction
Lassa virus (LASV), a human pathogen of the family Arenaviridae, is a rodent-borne virus that causes Lassa fever (LF) [1], [2]. LF is endemic in West African countries with the highest incidence in Nigeria, Guinea, Liberia, and Sierra Leone where up to 300,000 cases and 5000 deaths occur annually [3], [4]. LASV antibodies were detected in 8–52, 4–55, and 21% of the population in Sierra Leone, Guinea, and Nigeria, respectively, bringing the population at risk to 59 million with an estimated annual incidence of illness of 3 million [4], [5], [6]. Increasing international travel has resulted in importation of LF to non-endemic areas including European countries and the US [7]. The sizeable disease burden and the possibility that LASV can be used as an agent of biological warfare make a strong case for vaccine development [8], [9], [10].
LASV, like other members of the Arenaviridae, has a bi-segmented (L and S) ambisense RNA genome [11], [12]. The L RNA encodes a large protein (L, or RNA-dependent RNA polymerase) [13], and a small zinc-binding, Z protein [14]. The S RNA encodes the major structural proteins, nucleoprotein (NP), and glycoprotein precursor (GPC), cleaved into GP1, GP2, and signal peptide [15], [16], [17].
We previously described a live attenuated experimental vaccine for LF, clone ML29 [18], [19], [20]. The ML29 vaccine candidate encodes the NP and GPC of LASV and the Z protein and L protein of MOPV. Eighteen mutations distinguish the ML29 genome from the parental strains and likely contribute to the attenuated phenotype. The ML29 genotype was stable throughout 12 passages in tissue cultures.
Proof-of-concept studies in rodent models showed that the ML29 vaccine was attenuated and induced protective cell-mediated immune responses [19], [21]. However, LASV, a rodent-borne virus, is treated differently by the immune system of rodents and non-human primates [4]. Recently we developed a small non-human primate model of LF in the common marmoset, Callithrix jacchus[22]. We have used this model to evaluate safety, immunogenicity, and efficacy of the ML29 vaccine candidate.
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Viruses and cells
LASV (strain Josiah), MOPV (clone An 20410), MOP/LAS (clone ML29), and LCMV-WE viruses were previously described [18], [19], [20], [23]. The viruses were grown in Vero cells (ATCC, CRL-1586) and cultured in Dulbecco's modified minimum Eagle's medium (DMEM, GIBCO-BRL) with 2% fetal calf serum (FCS, GIBCO-BRL), 1% penicillin–streptomycin, and l-glutamine (2 mM) at 37 °C in 5% CO2 by using a multiplicity of infection (MOI) of 0.01. Supernatants were collected at 72 h post-infection, titrated on Vero
ML29 immunization of marmosets induces asymptomatic infection with low levels of transient viremia and viral load in tissues
None of the ML29-immunized marmosets had clinical manifestations during the observation period and hematological and chemical parameters were in normal ranges. With the high dose of ML29 we observed a minor elevation of alanine aminotransferase (ALT) in the plasma in comparison with animals immunized with the low dose. However, this elevation was still in the normal range (Fig. 1). In LASV-infected marmosets ALT levels in plasma were highly elevated indicating that liver was involved in LF
Discussion
There are several reasons to justify a replication-competent, “live”, vaccine as an attractive approach to control LF: (i) cell-mediated immunity plays the major role in LF patient recovery and in protection; (ii) a live vaccine provides the most effective natural pathway to process and present protective antigens to MHC molecules; (iii) epidemiological observations provide evidence that a single LASV exposure will induce long-term protection against disease [10]; and (iv) a vaccine candidate
Acknowledgements
This work was supported by grant RO1 AI052367 (to I.S.L.) from the National Institutes of Health and by the Regional Centers of Excellence in Biodefense and Emerging Infectious Diseases Researches, MARCE (U54 AI057168, subcontract to I.S.L.) and WRCE (U54 AI057156, subcontract to J.L.P.). Funding for microarray studies was from NIH grant AI053620 (to M.S.S.). We acknowledge the New England Primate Center (grant P51RR00168-45) for histological support.
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This work was presented in part at the Keystone Symposia “Challenge of Global Vaccine Development”, 8–13 October 2007, Cape Town, South Africa, and at the 13th International Congress on Infectious Diseases, 19–22 June 2008, Kuala Lumpur, Malaysia.
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Both authors have contributed equally to this paper.