Drug Discovery for Leishmaniasis

Luis Rivas, Carmen Gil, Luis Rivas, Carmen Gil

• Publisher: Royal Society of Chemistry
• Year: 2017
• ISBN:
  9781788012584
• Edition: 1
• Topic: Naturwissenschaften
• Subtopic: Klinische Chemie

For human health, leishmaniasis is among the most important protozoan diseases, superseded only by malaria. Globally, 10 to 12 million people are infected with 1.5 million new cases every year. The development of cheaper new drugs is urgently needed for this neglected disease that is developing resistance to current treatments. Chemotherapy remains the only treatment option for the bulk of patients. However, this is largely unaffordable for most. In the past three years numerous advances in drug discovery have been made for treating this disease by exploiting diverging metabolic pathways between the Leishmania enzymes and their hosts, using nanotechnology to target the immune cell phagolysosomes where Leishmania resides.

Drug Discovery for Leishmaniasis aims to provide a perspective of the current treatments and their challenges, blended with the emerging strategies and methodologies that will drive new target appraisals and drug developments, as well as addressing the molecular basis of resistance in Leishmania.

Recent studies have shown that leishmaniasis affects some of the poorest people in the world, with 95% of fatal cases occurring in only 6 countries. With the WHO goal of eliminating this public health problem in the South-east Asia Region by 2020, this book will be important for anyone who is interested in neglected tropical diseases.

III. The Quest for Achille’s Heel of Leishmania. Singular Targets as New Avenues for Drug Development

CHAPTER 12

Addressing the Molecular Biology of Leishmania for Drug Development

BRIANNA NORRIS-MULLINS AND MIGUEL A. MORALES^*

Eck Institute for Global Health, Department of Biological Sciences, University of Notre Dame, Notre Dame, IN 46556, USA

^*Email: [email protected]

12.1 Introduction

Kinetoplastid parasites of the genus Leishmania are the causative agents of leishmaniasis, a vector-borne disease that affects millions of people worldwide.^1–3 Globally, approximately 310 million people are at risk of acquiring the disease, which is endemic in over 90 countries spanning four continents.⁴ In its digenetic life cycle Leishmania parasites cycle between the sand fly vector, where they persist as promastigotes, and phagolysosomes of mammalian macrophages, where they survive as amastigotes. The ability of the parasites to survive these distinct environments is dependent on the developmental regulation of numerous genes, whose identification may result in therapeutic targets.⁵ Though there is currently no human safe vaccine for leishmaniasis, some treatments are available and are, typically, species-specific. Current existing therapies for leishmaniasis are inadequate due to resistance, safety and cost, underscoring the necessity for safer therapies with alternate modes of action. Leishmaniasis affects the poorest sectors of low-income countries, therefore, public health therapies to combat the disease have little prospect of major economic return for private industry. Consequently, there are few large-scale drug discovery programs to identify new chemical entities to combat leishmaniasis.^4,6,7 In implementing effective control strategies against leishmaniasis, it is vital to have a thorough understanding of the host–pathogen–vector relationship and, more importantly, of their interactions at a molecular level.^8,9 Recent advances in ‘omics’ technologies, including genomics, transcriptomics and proteomics, make available the tools necessary for advancing our current understanding of the molecular biology of Leishmania⁹ with the purpose of exploiting this information for the development of new alternatives for anti-leishmanial therapeutics.

12.2 The Leishmania Genome

In 2005, the first ever sequenced genomes of Leishmania major¹⁰ and two distantly related kinetoplastid protozoa, Trypanosoma brucei¹¹ and Trypanosoma cruzi,¹² revealed large-scale genetic preservation.¹³ Despite a conserved core of genes, more than 1000 Leishmania-specific genes were identified; many of which remain uncharacterized.¹⁴ In 2007, genomic studies were extended to two additional Leishmania species: Leishmania infantum and Leishmania braziliensis. Upon bioinformatics analysis, more than 99% gene conservation between the three Leishmania genomes (L. major, L. infantum and L. braziliensis) was identified, yielding roughly 200 differences at the gene or pseudogene level with only a fraction of these being species-specific.¹⁴ The genomes of Leishmania amazonensis, Leishmania mexicana, Leishmania donovani and Leishmania tarentolae have since been sequenced, and comparative analyses have shown similar results.^9,15 While findings indicate that minimal species-specific genes are important for pathogenesis, the underlying profound and unexpected conclusion is that the parasite genome plays a very small part in determining disease clinical presentation;¹⁴ this makes it nearly impossible to use species-specific genes for the development of therapeutic agents aimed at treating distinct clinical manifestations caused by Leishmania spp.

Leishmania parasites exhibit unique biological features ranging from gene organization, through transcription and mRNA processing, to post-translational modifications.^5,16 For example, at the genomic level, many Leishmania genes are arranged into long, strand-specific, polycistronic clusters (Figure 12.1).^10,17,18 This major difference between Leishmania spp. and higher eukaryotes has been extensively studied, has potential therapeutic implications for drug discovery and will be discussed in detail in the following section.

Figure 12.1 Gene Expression in Trypanosomatids vs. Higher Eukaryotes. In trypanosomatid parasites, large clusters of unrelated genes (coding regions) are organized as polycistronic transcription units (PTUs) under the control of a single promoter and termination codon (top). This differs from higher eukaryotes, where each individual gene has its own set of transcription signals (bottom).

12.3 The Leishmania Transcriptome

Although comparing the sequenced genomes of Leishmania species provides a solid basis for beginning to understand mechanisms of host–parasite interactions, it seems likely that a better understanding of disease pathogenesis could be gained by studying the regulation of gene expression during different parasite life cycle stages.⁹ The advent of high-throughput transcriptomic technologies, such as RNA sequencing, have facilitated such studies, however, the peculiar mechanisms of Leishmania gene expression partially dampen the enthusiasm for large-scale transcriptomic analyses. Indeed, most Leishmania genes have been shown to be constitutively expressed throughout the life cycle. It is important to note, though, that the noticeable variation in chromosome and gene copy numbers among L. infantum, L. braziliensis and L. major indicates that genome plasticity, rather than differential expression of single genes, could be the key to the different tissue tropism of Leishmania spp.⁹

12.3.1 Polycistronic Transcription and Trans-splicing Mechanisms

Similar to other eukaryotic organisms, Leishmania parasites use three different types of RNA polymerases (pol) to transcribe their genes. RNA pol I and RNA pol III are responsible for transcription of ribosomal RNA (rRNA) and transfer RNA (tRNA) genes, respectively, while RNA pol II transcribes protein-coding genes.¹⁹ It has been suggested, however, that in Leishmania, RNA pol II does not regulate transcription; this is due to the lack of transcription initiation sites for protein-coding genes within the organism.²⁰ Furthermore, common transcription facto...