An Improved Manufacturing Process for a Recombinant Polyprotein Vaccine

Combating the devastating global disease of leishmaniasis with a new therapeutic and prophylactic vaccine


A new recombinant protein vaccine against leishmaniasis has been produced by fusing portions of three leishmania-derived proteins. To simplify manufacturing, process improvements included removal of a 6-histidine sequence near the amino terminus, and the mutation of a proteolytic hot spot. The improved manufacturing method consists of fermenting an engineered version of the HMS-174 strain of E. coli following current good manufacturing practice regulations at the 30-L scale. A purification scheme yields purified protein product at greater than 100 mg/L. The purified protein then yields a stable cake that can safely be sent to clinical trials. The product was tested with the adjuvant monophosphoryl lipid A in stable emulsion (MPL-SE) in a murine potency assay. The Leish-110f recombinant protein product reported here is an improved version of the original leishmaniasis vaccine.

Leishmaniasis is a devastating disease that causes untold human suffering and numerous deaths worldwide.1 Leishmaniasis is a spectrum of diseases ranging from a fatal visceral form to a localized self-healing cutaneous lesion. The global prevalence of leishmaniasis, caused by the intracellular protozoan Leishmania species, is estimated to be approximately 12,000,000 cases total, with 500,000 new reports of visceral leishmaniasis and 1,500,000 million new reports of cutaneous leishmaniasis each year.2 Although much progress has been made in developing drug therapies for leishmaniasis, a number of clinical problems, such as length of treatment and prohibitive expense, are associated with these therapies. Therefore, new vaccines and improvements to existing clinical products are valuable additions to the fight against this terrible disease.

The first-line clinical therapy, antimonial drug, was developed almost a century ago. This clinical regimen requires multiple injections (four weeks of daily injections for the visceral form of leishmaniasis), is costly, often associated with unpleasant side effects, and is becoming ineffective in many endemic locales. With resistance to therapeutic antimonials increasing to 60% in certain parts of India, the mortality rate from visceral leishmaniasis has been very high.3 Recent progress has been made by an oral anti-leishmaniasis drug, miltefosine, with the current clinical recommendation to use this drug as frontline treatment for some forms of leishmaniasis, including cutaneous leishmaniasis. Miltefosine, however, is teratogenic and requires an extended therapeutic regimen. Another therapeutic, liposomal amphotericin B, also has considerable efficacy against clinical leishmaniasis, but the expense of the drug renders it impractical in all but severe cases of the disease. Both miltefosine and amphotericin B function independently of the immune system, therefore, when the drug is withdrawn before completion of the therapeutic regimen, parasite loads re-emerge.4 Control of the phlebotomine sand fly vectors is also not feasible given the prohibitive expense associated with insecticide use.

The development of a safe, effective, and economical vaccine product is possible, given that controlled vaccination with live parasites has been practiced for centuries in the Middle East, and individuals who recover from clinical leishmaniasis develop immunity against re-infection. Additionally, recent trials with killed parasites demonstrate good efficacy of such a vaccine in post-infection settings of cutaneous leishmaniasis,5–8 and numerous studies have demonstrated the effectiveness of such vaccines in experimental models of leishmaniasis.9–14

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