Mosquito-borne disease which is Malaria can infected people to become sick with high fever, chills, and flu-likeillness. The most dangerous it can alsocause death. According to National Institute of Allergy , and InfectiousDiseases (NIAID) ,substantial progress hasbeen made globally to control and eliminate malaria, but it continues to be asignificant public health problem with roughly 3.2 billion people worldwide atrisk for the disease. Malaria is disease that are responsibleglobally for 500 million cases of clinical disease and presents a public healthproblem for 2.4 billion people in over 90 countries representing 40% of the world’s population.
( MalariaFoundation International). Thereare four species of protozoan parasites in Malaria human which the genus is Plasmodium:P. falciparum, P.
vivax, P. ovale, and P. malariae.Although P. vivax is responsible for most malaria infectionsin the world, the most severe form of malaria is caused by P.falciparum.
The severity of malarial illness depends largely on theimmunological status of the person who is infected. Partial immunity developsover time through repeated infection, and without recurrent infection, immunityis relatively short-lived. (Instituteof Medicine (US) Committee for the Study on Malaria Prevention and Control ,1991 ).
Thehuman stage of the life cycle beginswith the exoerythrocytic phase. When an infected mosquito bites a human, sporozoites in the mosquito’s saliva enterthe bloodstream . The sporozoites travel to the liver, where they invadehepatocytes over a period of up to 4 weeks, the infected hepatocytes matureinto schizonts. In Plasmodium vivax and P.ovale infections only, some schizonts may remaindormant as hypnozoites for weeks to years before causing clinical relapses.With schizont rupture, merozoites are released into the bloodstream .
In theerythrocytic phase, merozoites invade erythrocytes and either undergo anasexual cycle of reproduction or develop into non multiplying sexual formsgametocytes. These gametocytes are crucial for perpetuating the life cycle, asthey are ingested by a feeding mosquito and undergo sexual reproduction withinthe mosquito midgut; thousands of infective sporozoites are produced, whichthen migrate to the salivary glands, ready to initiate another life cycle ( Suhet al., 2004 ).1.2 Drug resistant problemIn general, malaria is acurable disease, and if everyone has access to early treatment, nobody shoulddie from it.
For the past 50 years, there have been two main classes ofantimalarial agents in use, the antifolates and the cinchona alkaloids or thequinoline-containing drugs. In most cases, these agents are targeted at theasexual erythrocytic stage of the parasite ( Philips , 2001 ).Antimalarialdrug resistance has confront with one of the greatest challenges facing malariacontrol nowadays. According to Peter B. Bloland, (2001), the definition of resistance is ability of a parasite strain to surviveand/or multiply despite the administration and absorption of a drug given indoses equal to or higher than those usually recommended but within tolerance ofthe subject then to specify that the drug must gain access to the parasite orthe infected red blood cell for the duration of the time necessary for itsnormal action .
Drug resistance has been implicated in thespread of malaria to new areas and re-emergence of malaria in areas where thedisease had been eradicated. Drug resistance has also played a significant rolein the occurrence and severity of epidemics in some parts of the world.Population movement has introduced resistant parasites to areas previously freeof drug resistance.
The economics of developing new pharmaceuticals fortropical diseases, including malaria, are such that there is a great disparitybetween the public health importance of the disease and the amount of resourcesinvested in developing new cures . This disparity comes at a time when malariaparasites have demonstrated some level of resistance to almost everyantimalarial drug currently available, significantly increasing the cost andcomplexity of achieving parasitological cure.Thegoal of antimalarial drug resistance monitoring has been to detect cases oftreatment failure caused by drug-resistant parasites before they becomewidespread in the population and lead to increased morbidity and mortality.This is deceptively simple, because recurrent parasitemia in endemic countriescan also result from a host of other factors including noncompliance, poorquality drug, inadequate drug levels, new infection and even varying levels ofimmunity to malaria.
Artemisinin therapies (ACT), in which more than one drug is mixed together toprevent the parasite from developing resistance to any one treatment, is a farmore effective therapy than pure chloroquine. However, ACTs cost 10 times morethan mono-drug therapy. Since pharmaceutical companies stand to gain littlefinancially from ACT sales in the developing world, their use remains low as noincentive structure exists to encourage their wider distribution (Arrow et.al.,2005). There are many factors that have been associated with antimalarial drug resistanceinclude such disparate issues as human behaviour ,vector and parasite biology,pharmacokinetics, and economics. Futhermore, the factor that decrease the effectiveness of the immunesystem in clearing parasite residuum after treatment also appear to increasesurvivorship of parasites and facilitate development and intensification of resistance.
This mechanism has been suggested as a significant contributor to resistance inSouth-East Asia, where parasites are repeatedly cycled through populations ofnon-immune individuals.1.3 Why kinase is the targetfor new malaria drug.In eukaryotic cells, proteinkinases are known to play key roles in cell cycle regula? tion and signalingpathways. Protein kinases are fascinating biological catalysts with a rapidlyexpanding knowledge base, a growing appreciation in cell regulatory control,and an ascendant role in successful therapeutic intervention protein kinases make up almost 2% of the humangenome and control most important biological processes.The activity ofcAMP?dependent protein kinase (PKA) depends on A?kinase anchoring proteins(AKAPs) through protein interactions. While several components of the cAMPdependent pathway includ?ing the PKA catalytic and regulatory subunits—havebeen characterized in P.
falciparum, whether AKAPs are involved in this pathwayremains unclear. Here, PfAKAL, an open reading frame of a potential AKAP?likeprotein in the P. falciparum genome was identified, and its protein partnersand putative cellular functions characterized.Identification and validation of the target asthe true direct driver is the most important step in drug discovery, or inassociation with other proteins may cause an altered pathological phenotype.Many failures during the drug discovery process are ascribed mainly to a wrongtarget, or that the animal model does not mimic the exact phenotype in humans.Other factors such as physicochemical properties, poor absorption,distribution, metabolism and excretion characteristics and in vivotoxicological outcomes also impact the attrition rate of drug candidates (Waring et al. , 2015).
Most kinases are promiscuousand will catalyze the phosphotransfer from ATP to alternative substrates withdiffering degrees of catalytic efficiency. Protein kinases (PKs) catalyze the phosphorylation ofproteins at serine, threonine and tyrosine residues. This post-translationmodification is important to most physiological processes; there are hundredsof different protein kinases that catalyze specific phosphorylation reactions.Kinases are important targets for drug discovery with now over 30 medicinesapproved that inhibit protein kinases .(Haubrich and Swinney , 2016). CHAPTER2 MOLECULAR CHARACTERIZATION OF MALARIA KINASE2.
1 Calcium Dependent Protein Kinase 1 (CDPK1)According to Garcia (1999), the presence ofcalcium in the medium used in the culture of P. falciparum is requiredfor parasite invasion of erythrocytes and that chelating calcium in infectederythrocytes results in developmental arrest and impairs parasite invasion inculture.In Plasmodium falciparum,calcium-dependent protein kinase 1 (PfCDPK1) is expressed during schizogony inthe erythrocytic stage as well as in the sporozoite stage.
It is coexpressedwith genes that encode the parasite motor complex, a cellular componentrequired for parasite invasion of host cells, parasite motility and potentiallycytokinesis. It does cause the suddenarrest of developmental progression in late schizogony, resulting in areduction in the number of ring-stage parasites in culture. From Kato et al.,(2008) findings, along withthe results obtained by the ontology-based pattern identification analysis,strongly suggest that PfCDPK1 is involved in regulating parasitemotor-dependent processes that occur in the late schizont stage.
These mayinclude cytokinesis, egress and invasion, all of which are closely linked inmalaria parasites. Recently, Plasmodium falciparum calcium-dependent proteinkinase 1 (PfCDPK1) has been found in the membrane and organelle fraction of theparasite. The kinase contains three motifs for membrane binding at itsN-terminus, a consensus sequence for myristoylation, a putative palmitoylationsite and a basic motif.Knockdownof CDPK1 in sexual stages resulted in developmentally arrested parasites andprevented mosquito transmission, and these effects were independent of thepreviously proposed function for CDPK1 in regulating parasite motility.In-depth translational and transcriptional profiling of arrested parasitesrevealed that CDPK1 translationally activates mRNA species in the developingzygote that in macrogametes remain repressed via their 30 and 50 UTRs. Thesefindings indicate that CDPK1 is a multifunctional protein that translationallyregulates mRNAs to ensure timely and stage-specific protein expression.
( Sarah et al., 2012)2.2. Calcium Dependant Protein Kinase 4 (CDPK4) According to Oliver (2004), a member of afamily of plant-like calcium dependent, CDPK4, is identified as the molecular switch that translates the xanthurenicacid (XA)which is small mosquito molecule induced calcium signal into a cellular response by regulating cellcycle pro- during gametogenesis in Plasmodiumberghei, a malaria parasite gression in the male gametocyte. CDPK4 is shownbe essential for the sexual reproduction and mosquito specialized of P.berghei. The study from an unexpected function for a plant-like signalingpathway in cell cycle regulation and life cycleprogression of malaria parasite.Their developmental regulation makes thesekinases prime candidates for the molecular switches that translate ubiquitousCa2+ signals into appropriate cellular responses at specificstages of the parasite’s life cycle.
CDPK4 triggers cell cycle progression byinitiating the raoid threefold replication of the genome that is completed withabout 8 minute of activation (Billker et al.,20014). It is also essential toinduce to the three rounds of mitosis that take place in parallel with DNAreplication and to initiate the polymerization of axonmes that form the motilebackbone of the microgamets ( Tewari et al .
, 2005).According to Sarah (2012),CDPK4 is the Ca2+ kinase critical for initiation od DNAreplication during the first minute afer microgametocytes activation but thiskinase was not required for the emergence of gametocytes from their host cellsleading to the other calcium effectors that invoved. 2.3Mitogen – associated Protein Kinase 2 (MAPK2) The purified recombinant enzyme displayed functionalcharacteristics of MAPKs such as (i) ability to undergo autophosphorylation,playimportant roles in signal transduction pathways regulating adaptative responseto a wide range of stimuli. Several pathways involving different MAP kinasescoexist in the cell (ii) ability to phosphorylate myelin basic protein, aclassical MAPK substrate, (iii) regulation of kinase activity by aMAPK-specific phosphatase, and (iv) ability to be activated by component(s)present in cell extracts. Mutational analysis of the recombinant protein allowedthe identification of residues that are important for enzymatic activity.
CHAPTER 3 SUBSTRATE IDENTIFICATION OF MALARIA KINASE3.1 Casein Kinase I (CKI)Casein kinase I (CKI) was one of the firstserine/threonine protein kinase activities to be isolated and characterised thatphosphorylates a large number of protein substrates .The casein kinase I (CKI)isoforms from yeast have been genetically linked to vesicular trafficking, DNArepair, cell cycle progression and cytokinesis.CKI was found to be highly conserved in allorganisms which perform various functions in both the cytoplasm and nucleus,such as DNA repair, cellcycle, cytokinesis, vesicular trafficking, morphogenesis andcircadian rhythm.
CKI uses ATP as a phosphate donor to phosphorylate serine andthreonine residues of target proteins. Many proteins, such as glycogensynthase, tumor suppressor p53, the cAMP-responsive element modulator (CREM),and the type I protein phosphatase inhibitor-2 are substrates of CK1.CKI contains a highly conserved kinase domainresponsible for catalytic activity at the N-terminus and a highly diverseregulatory domain responsible for both regulation of kinase activity andsubcellular targeting at the C-terminus. 3.2 Casein Kinase ?(CK?) Mammalianprotein kinase CK2 is a pleiotropic serine/ threonine protein kinase know toact on hundreds of cellular substrates involved in crucial cellular processessuch as differentiation, proliferation, apoptosis, stress response, DNA damageand circadian rhythm. CK2 was constitutivelyactive in unfertilized and fertilized oocytes. The enzymatic activityoscillated through meiosis showing three major peaks: soon after fertilization(metaphase I exit), before metaphase II, and at the exit from metaphase I(Holland et al.
,2008). CK2 catalyzes thephosphorylation of a great number of substrates presenting multiple acidicresidues surrounding the phosphoacceptor amino acid and the potential role of PfCK2 in the nucleushas identified a number of interacting partners and substrate nuclear proteinsinvolved in chromatin assembly and dynamics( Graciotti et al.,2014). CHAPTER 4DISCUSSION4.
1 Potential as a newmalarial drug target Newopportunities to discover medicines for neglected diseases can be leveraged bythe extensive kinase tools and knowledge created in targeting human kinases.Theprotein kinase is in its relevant state of activation with the relevantsubstrates present; this only selects cell penetrant compounds and compoundsthat have a lethal effect; it would detect compounds that were acting onmultiple proteins. The disadvantages of this approach include the unknownidentity of the target(s), which could make optimization of hits problematic,particularly if there were pharmacokinetic or toxicological issues.( Irene ,2017)4.
2 Controvercy ofantimalarial drug resisstance Anti-malarialresistance, especially to artemisinin-based combination therapy (ACT), is amajor threat to malaria control efforts. Resistance generally results frominappropriate, incomplete or inadequate courses of treatment. This mostcommonly occurs in the context of poverty, counterfeit medications, and weakhealthcare infrastructure.
( World Malaria Report 2012). CHAPTER 5CONCLUSION A molecular understanding of the life cycle of P.falciparum will facilitate the rational design of new therapies.
Efficientegress of P. falciparum out of an infected human red blood cell is afundamental step in the parasite life cycle.
