Protect Host Cells Research Articles

Lack of association between complement factor H polymorphisms and coronary artery disease or myocardial infarction

Complement factor H (CFH) plays a critical role in the protection of host cells and tissues from damage by complement activation and has been suggested to protect against the progression of atherosclerosis. A polymorphism in the CFH gene, Y402H, known to be strongly associated with age-related macular degeneration, has been analyzed in relation to coronary artery disease (CAD) in several studies with conflicting results. We investigated the association of polymorphisms of the CFH gene in two large-scale studies on CAD and myocardial infarction (MI). The AtheroGene Study included a cohort of cases with CAD (n = 1,303) prospectively followed for a median period of 6.2 years, among whom198 experienced a cardiovascular event, and a group of 483 control subjects. The AtheroGene Study population was genotyped for the Y402H, I62V, and E936D polymorphisms. There was no significant difference in genotypic or allelic frequencies between CAD cases and controls. Among cases, no significant association was found with prospective cardiovascular outcome. Many inflammatory proteins, including the C-reactive protein, were measured, and none of the polymorphisms showed an association with these markers. The Etude Cas-Témoin de l'Infarctus du Myocarde (ECTIM) Study compared 1,034 patients with MI and 1,039 controls from France and United Kingdom. The ECTIM Study population was genotyped for the Y402H polymorphism. Genotype and allele frequencies were similar in cases and controls. These results do not support an involvement of common nonsynonymous polymorphisms of the CFH gene in predisposition to CAD and its complications.

Legionella pneumophila inhibits macrophage apoptosis by targeting pro-death members of the Bcl2 protein family

To establish a vacuole that supports bacterial replication, Legionella pneumophila translocates a large number of bacterial proteins into host cells via the Dot/Icm type IV secretion system. Functions of most of these translocated proteins are unknown, but recent investigations suggest their roles in modulating diverse host processes such as vesicle trafficking, autophagy, ubiquitination, and apoptosis. Cells infected by L. pneumophila exhibited resistance to apoptotic stimuli, but the bacterial protein directly involved in this process remained elusive. We show here that SidF, one substrate of the Dot/Icm transporter, is involved in the inhibition of infected cells from undergoing apoptosis to allow maximal bacterial multiplication. Permissive macrophages harboring a replicating sidF mutant are more apoptotic and more sensitive to staurosporine-induced cell death. Furthermore, cells expressing SidF are resistant to apoptosis stimuli. SidF contributes to apoptosis resistance in L. pneumophila-infected cells by specifically interacting with and neutralizing the effects of BNIP3 and Bcl-rambo, two proapoptotic members of Bcl2 protein family. Thus, inhibiting the functions of host pro-death proteins by translocated effectors constitutes a mechanism for L. pneumophila to protect host cells from apoptosis.

The C-terminus of complement factor H is essential for host cell protection

Complement is a powerful self-amplifying system of innate immune defense with the capacity to eliminate microbes directly. Factor H is a central regulator in plasma which protects host tissue from complement mediated damage. Here we characterize the relevance of surface attached factor H, and study the regulatory activity of factor H on endothelial cells. Although these cells expressed membrane bound regulators, cell bound factor H contributed substantially to complement regulatory activity at the cell surface. Blockade of the C-terminus of factor H with monoclonal antibodies inhibited cell binding of this soluble regulator and resulted in enhanced complement activation on the cells. In the absence of factor H, increased deposition and slower inactivation of C3b resulted in higher amount of membrane attack complexes on the cell surface. When the membrane regulators CD55 and CD59 were removed by enzymatic treatment, complement mediated cell lysis was enhanced in the absence of factor H. Importantly, inhibition of the C-terminus did not compromise the regulatory function of factor H in fluid phase. Altogether these data point to a highly relevant, yet so far underestimated role of factor H for complement control at cellular surfaces, and reveal a decisive role of the factor H C-terminus in host cell recognition and protection.

Cooperation between Human DAF and CD59 in Protecting Cells from Human Complement-mediated Lysis

The complement (C) regulatory proteins decay accelerating factor (DAF, CD55) and CD59 could protect host cells using different mechanisms from C-mediated damage at two distinct levels within the C pathway. Co-expression of DAF and CD59 would be an effective strategy to help overcome host C-induced xenograft hyperacute rejection. In this study, we made a construct of recombinant expression vector containing DAF and CD59 cDNA and the stable cell lines were obtained by G418 selection. Extraneous genes integration and co-expression were identified by PCR, RT-PCR and Western blot analysis. Human c-mediated cytolysis assays showed that NIH/3T3 cells transfected stably with pcDNA3-CD59, pcDNA3-DAF, and pcDNA3-CD59DAF-DP were protected from Cmediated damage and that synchronously expressed human CD59 and DAF provided the most excellent protection for host cells as compared with either human CD59 or DAF expressed alone. Therefore, the construct represents an effective and efficacy strategy to overcome C-mediated damage in cells and, ultimately, in animals.

Critical Role of the C-Terminal Domains of Factor H in Regulating Complement Activation at Cell Surfaces

The plasma protein factor H primarily controls the activation of the alternative pathway of complement. The C-terminal of factor H is known to be involved in protection of host cells from complement attack. In the present study, we show that domains 19-20 alone are capable of discriminating between host-like and complement-activating cells. Furthermore, although factor H possesses three binding sites for C3b, binding to cell-bound C3b can be almost completely inhibited by the single site located in domains 19-20. All of the regulatory activities of factor H are expressed by the N-terminal four domains, but these activities toward cell-bound C3b are inhibited by isolated recombinant domains 19-20 (rH 19-20). Direct competition with the N-terminal site is unlikely to explain this because regulation of fluid phase C3b is unaffected by domains 19-20. Finally, we show that addition of isolated rH 19-20 to normal human serum leads to aggressive complement-mediated lysis of normally nonactivating sheep erythrocytes and moderate lysis of human erythrocytes, which possess membrane-bound regulators of complement. Taken together, the results highlight the importance of the cell surface protective functions exhibited by factor H compared with other complement regulatory proteins. The results may also explain why atypical hemolytic uremic syndrome patients with mutations affecting domains 19-20 can maintain complement homeostasis in plasma while their complement system attacks erythrocytes, platelets, endothelial cells, and kidney tissue.

Transplantation of primed human fetal neural stem cells improves cognitive function in rats after traumatic brain injury

Traumatic brain injury (TBI) often produces cognitive impairments by primary or secondary neuronal loss. Stem cells are a potential tool to treat TBI. However, most previous studies using rodent stem or progenitor cells failed to correlate cell grafting and cognitive improvement. Furthermore, the efficacy of fetal human neural stem cells (hNSCs) for ameliorating TBI cognitive dysfunction is undetermined. This study therefore characterized phenotypic differentiation, neurotrophic factor expression and release and functional outcome of grafting hNSCs into TBI rat brains. Adult Sprague–Dawley rats underwent a moderate parasagittal fluid percussion TBI followed by ipsilateral hippocampal transplantation of hNSCs or vehicle 1 day post-injury. Prior to grafting, hNSCs were treated in vitro for 7 days with our previously developed priming procedure. Significant spatial learning and memory improvements were detected by the Morris water maze (MWM) test in rats 10 days after receiving hNSC grafts. Morphological analyses revealed that hNSCs survived and differentiated mainly into neurons in the injured hippocampus at 2 weeks after grafting. Furthermore, hNSCs expressed and released glial-cell-line-derived neurotrophic factor (GDNF) in vitro and when grafted in vivo, as detected by RT-PCR, immunostaining, microdialysis and ELISA. This is the first direct demonstration of the release of a neurotrophic factor in conjunction with stem cell grafting. In conclusion, human fetal neural stem cell grafts improved cognitive function of rats with acute TBI. Grafted cells survived and differentiated into neurons and expressed and released GNDF in vivo, which may help protect host cells from secondary damage and aid host regeneration.

Biosynthesis of ascorbic acid in legume root nodules.

Ascorbic acid (vitamin C) is a major antioxidant and redox buffer, but is also involved in other critical processes of plants. Recently, the hypothesis has been proposed that legume nodules are unable to synthesize ascorbate and have to import it from the shoot or root, thus providing a means by which the plant regulates nodule senescence. The last step of ascorbate biosynthesis in plants is catalyzed by L-galactono-1,4-lactone dehydrogenase (GalLDH). The mRNAs encoding GalLDH and three other enzymes involved in ascorbate biosynthesis are clearly detectable in nodules. Furthermore, an active membrane-bound GalLDH enzyme is present in nodule mitochondria. Biochemical assays on dissected nodules reveal that GalLDH activity and ascorbate are correlated in nodule tissues and predominantly localized in the infected zone, with lower levels of both parameters (relative to the infected tissues) in the apex (87%) and senescent region (43%) of indeterminate nodules and in the peripheral tissues (65%) of determinate nodules. In situ RNA hybridization showed that the GalLDH mRNA is particularly abundant in the infected zone of indeterminate and determinate nodules. Thus, our results refute the hypothesis that ascorbate is not synthesized in nodules and lend support to a previous conclusion that ascorbate in the infected zone is primarily involved in the protection of host cells against peroxide damage. Likewise, the high ascorbate and GalLDH activity levels found in the apex of indeterminate nodules strongly suggest a participation of ascorbate in additional functions during symbiosis, possibly related to cell growth and division and to molecular signaling.

Phenotypic expression of factor H mutations in patients with atypical hemolytic uremic syndrome

We investigated the phenotypic expression of factor H mutations in two patients with atypical hemolytic uremic syndrome (HUS). Factor H in serum was assayed by rocket immunoelectrophoresis, immunoblotting, and double immunodiffusion and in tissue by immunohistochemistry. Functional activity was analyzed by hemolysis of sheep erythrocytes and binding to endothelial cells. A homozygous mutation in complement control protein (CCP) domain 10 of factor H was identified in an adult man who first developed membranoproliferative glomerulonephritis and later HUS. C3 levels were very low. The patient had undetectable factor H levels in serum and a weak factor H 150 kDa band. Double immunodiffusion showed partial antigenic identity with factor H in normal serum owing to the presence of factor H-like protein 1. Strong specific labeling for factor H was detected in glomerular endothelium, mesangium and in glomerular and tubular epithelium as well as in bone marrow cells. A heterozygous mutation in CCP 20 of factor H was found in a girl with HUS. C3 levels were moderately decreased at onset. Factor H levels were normal and a normal 150 kDa band was present. Double immunodiffusion showed antigenic identity with normal factor H. Factor H labeling was minimal in the renal cortex. Factor H dysfunction was demonstrated by increased sheep erythrocyte hemolysis and decreased binding to endothelial cells. In summary, two different factor H mutations associated with HUS were examined: in one, factor H accumulated in cells, and in the other, membrane binding was reduced.

Group V and X secretory phospholipase A2 prevents adenoviral infection in mammalian cells

sPLA2 (secretory phospholipase A2) enzymes have been implicated in various biological events, yet their precise physiological functions remain largely unresolved. In the present study we show that group V and X sPLA2s, which are two potent plasma membrane-acting sPLA2s, are capable of preventing host cells from being infected with an adenovirus. Bronchial epithelial cells and lung fibroblasts pre-expressing group V and X sPLA2s showed marked resistance to adenovirus-mediated gene delivery in a manner dependent on their catalytic activity. Although adenovirus particles were insensitive to recombinant group V and X sPLA2s, direct addition of these enzymes to 293A cells suppressed both number and size of adenovirus plaque formation. Group V and X sPLA2s retarded the entry of adenovirus into endosomes. Moreover, adenoviral infection was suppressed by LPC (lysophosphatidylcholine), a membrane-hydrolytic product of these sPLA2s. Thus hydrolysis of the plasma membrane by these sPLA2s may eventually lead to the protection of host cells from adenovirus entry. Given that group V and X sPLA2s are expressed in human airway epithelium and macrophages and that the expression of endogenous group V sPLA2 is upregulated by virus-related stimuli in these cells, our present results raise the possibility that group V and X sPLA2s may play a role in innate immunity against adenoviral infection in the respiratory tract.

Analysis of membrane-bound complement regulatory proteins in prostate cancer

ObjectivesMembrane-bound complement regulatory proteins (mCRPs) are ubiquitously expressed throughout the cells of the hematopoietic system and protect host cells from complement-mediated lysis. The expression and functionality of mCRPs have been demonstrated in several types of cancers; however, no clear correlation has been appreciated between tumor stage and mCRP expression. MethodsWe investigated the expression pattern of CD46, CD55, CD59, and CD97 (a receptor for CD55) in prostate cancer by tissue microarray analysis using high-density tissue microarrays spotted with tissue cores from biopsy and autopsy specimens. Real-time polymerase chain reaction was used to confirm the tissue microarray data by quantifying mRNA expression in clinically identified tissue specimens. ResultsCD55 and CD97 demonstrated increased expression in prostatic intraepithelial neoplasia, localized prostate cancer, and metastatic prostate cancer specimens compared with normal tissue specimens. No appreciable difference was seen in CD46 or CD59 expression. These data were confirmed by real-time polymerase chain reaction of mRNA expression levels in grossly identified specimens. ConclusionsThese data suggest that mCRPs are differentially expressed in prostate cancer, with CD55 the predominant mCRP upregulated in malignant prostate epithelial cells. Additionally, CD97 expression correlated with the malignant phenotype and may contribute to the tumorigenic and metastatic potential of prostate cancer.

The expression of CD97EGF and its ligand CD55 on marginal epithelium is related to higher stage and depth of tumor invasion of gastric carcinomas

CD97EGF is a member of the EGF-TM7 family of class II seven-transmembrane (7TM), and its cellular ligand CD55 (also known as decay accelerating factor; DAF) protects host cells from complement attack. To determine whether the expression levels of these two molecules are correlated with the clinicopathological features of gastric carcinomas, a total of 35 gastric carcinomas and their corresponding margins and normal specimens were investigated by RT-PCR, Western blot analysis and immunohistochemistry. Transcript levels of CD97EGFand CD55 were higher in tumors than those in the margin and normal epithelial mucous tissues (P<0.05). However, the expression levels of CD97EGF and CD55 mRNA had no correlation with the clinicopathological features of gastric carcinoma patients. All three groups of specimens were immunoreactive for CD97EGF and the CD55 protein. Strong and specific immunoreactivities of CD97EGF were located in the mucosal epithelia of the marginal basal membrane. Expression of CD97(EGF) in the margins showed a marked difference between the depth of tumor invasion T1 and T2, 3 and 4, and stages I and II/III/IV of gastric carcinomas (P<0.05). The expression of CD55 protein was highly correlated with CD97EGF (R=0.6483, P<0.001). Western blot analysis confirmed the expression and distribution patterns of CD97EGF and CD55. Our findings suggest that CD97EGF may play a role in the development and invasion of gastric carcinomas by binding its cellular ligand CD55. Detection of the CD97EGF expression in the tumor margin could be referred to as the molecular edge of gastric carcinomas.

A mosquito 2-Cys peroxiredoxin protects against nitrosative and oxidative stresses associated with malaria parasite infection

Malaria parasite infection in anopheline mosquitoes induces nitrosative and oxidative stresses that limit parasite development, but also damage mosquito tissues in proximity to the response. Based on these observations, we proposed that cellular defenses in the mosquito may be induced to minimize self-damage. Specifically, we hypothesized that peroxiredoxins (Prxs), enzymes known to detoxify reactive oxygen species (ROS) and reactive nitrogen oxide species (RNOS), protect mosquito cells. We identified an Anopheles stephensi 2-Cys Prx ortholog of Drosophila melanogaster Prx-4783, which protects fly cells against oxidative stresses. To assess function, AsPrx-4783 was overexpressed in D. melanogaster (S2) and in A. stephensi (MSQ43) cells and silenced in MSQ43 cells with RNA interference before treatment with various ROS and RNOS. Our data revealed that AsPrx-4783 and DmPrx-4783 differ in host cell protection and that AsPrx-4783 protects A. stephensi cells against stresses that are relevant to malaria parasite infection in vivo, namely nitric oxide (NO), hydrogen peroxide, nitroxyl, and peroxynitrite. Further, AsPrx-4783 expression is induced in the mosquito midgut by parasite infection at times associated with peak nitrosative and oxidative stresses. Hence, whereas the NO-mediated defense response is toxic to both host and parasite, AsPrx-4783 may shift the balance in favor of the mosquito.

Regulator of Complement Activation (RCA) Locus in Chicken: Identification of Chicken RCA Gene Cluster and Functional RCA Proteins

A 150-kb DNA fragment, which contains the gene of the chicken complement regulatory protein CREM (formerly named Cremp), was isolated from a microchromosome by screening bacterial artificial chromosome library. Within 100 kb of the cloned region, three complete genes encoding short consensus repeats (SCRs, motifs with tandemly arranged 60 aa) were identified by exon-trap method and 3'- or 5'-RACE. A chicken orthologue of the human gene 6-phosphofructo-2-kinase/fructose-2,6-biphosphatase 2, which exists in close proximity to the regulator of complement activation genes in humans and mice, was located near this chicken SCR gene cluster. Moreover, additional genes encoding SCR proteins appeared to be present in this region. Three distinct transcripts were detected in RNA samples from a variety of chicken organs and cell lines. Two novel genes named complement regulatory secretory protein of chicken (CRES) and complement regulatory GPI-anchored protein of chicken (CREG) besides CREM were identified by cloning corresponding cDNA. Based on the predicted primary structures and properties of the expressed molecules, CRES is a secretory protein, whereas CREG is a GPI-anchored membrane protein. CREG and CREM were protected host cells from chicken complement-mediated cytolysis. Likewise, a membrane-bound form of CRES, which was artificially generated, also protected host cells from chicken complement. Taken together, the chicken possesses an regulator of complement activation locus similar to those of the mammals, and the gene products function as complement regulators.

GRIM-19 Interacts with Nucleotide Oligomerization Domain 2 and Serves as Downstream Effector of Anti-bacterial Function in Intestinal Epithelial Cells

Nucleotide oligomerization domain 2 (NOD2) functions as a mammalian cytosolic pathogen recognition molecule, and variants have been associated with risk for Crohn disease. We recently demonstrated that NOD2 functions as an anti-bacterial factor limiting survival of intracellular invasive bacteria. To gain further insight into the mechanism of NOD2 activation and signal transduction, we performed yeast two-hybrid screening. We demonstrate that GRIM-19, a protein with homology to the NADPH dehydrogenase complex, interacts with endogenous NOD2 in HT29 cells. GRIM-19 is required for NF-kappaB activation following NOD2-mediated recognition of bacterial muramyl dipeptide. GRIM-19 also controls pathogen invasion of intestinal epithelial cells. GRIM-19 expression is decreased in inflamed mucosa of patients with inflammatory bowel diseases. GRIM-19 may be a key component in NOD2-mediated innate mucosal responses and serve to regulate intestinal epithelial cell responses to microbes.

Lentiviral siRNAs targeting multiple highly conserved RNA sequences of human immunodeficiency virus type 1

The high mutation rate of the human immunodeficiency virus (HIV) makes it difficult for any therapy employing a single anti-HIV targeting mechanism to sustain prolonged effect. In an attempt to explore novel therapy for AIDS, we developed and tested lentiviral small interfering RNA (siRNA) vectors targeting multiple highly conserved regions in the HIV type 1 (HIV-1) genome. The siRNA expression cassette was cloned into an extensively deleted HIV-1-derived lentiviral self-inactivating insulator (SIN) insulator [corrected] vector. Although some of the siRNAs targeting sites were also present in the helper construct of the vector system, the production of these lentiviral siRNA vectors were not significantly affected. When tested against different HIV-1 strains including pNL4-3 (subtype B), p89.6 (subtype B) and p90CF402.1.8 (subtype A/E recombinant), the siRNAs targeting conserved gag, pol, int and vpu, but not U3, nef or U5 regions, efficiently inhibited replication of all three viral strains. These lentiviral siRNA vectors also protected host cells from syncytium-forming macrophage- and T-cell-tropic HIV-1-induced cytotoxicity. Transduction of a long-term chronically infected human lymphoma cell line with lentiviral siRNAs resulted in stable inhibition of HIV-1 replication. Northern analysis showed that both genomic and subgenomic viral RNA species were downregulated. In addition, the viral RNA was inhibited in both the nuclear and cytoplasmic compartments of [corrected] chronically infected cells after prolonged passage, suggesting that [corrected] lentiviral siRNAs have a nuclear effect [corrected] Using these lentiviral siRNA [corrected] vectors, we further demonstrated reduced replication kinetics of HIV-1 in primary human peripheral blood lymphocytes. These results suggest that lentiviral siRNAs targeting multiple conserved HIV-1 sequences holds significant promise for the treatment of HIV-1 infections.

HGF/MET signalling protects Plasmodium-infected host cells from apoptosis

Plasmodium, the causative agent of malaria, migrates through several hepatocytes before initiating a malaria infection. We have previously shown that this process induces the secretion of hepatocyte growth factor (HGF) by traversed cells, which renders neighbour hepatocytes susceptible to infection. The signalling initiated by HGF through its receptor MET has multifunctional effects on various cell types. Our results reveal a major role for apoptosis protection of host cells by HGF/MET signalling on the host susceptibility to infection. Inhibition of HGF/MET signalling induces a specific increase in apoptosis of infected cells leading to a great reduction on infection. Since HGF/MET signalling is capable of protecting cells from apoptosis by using both PI3-kinase/Akt and, to a lesser extent, MAPK pathways, we determined the impact of these pathways on Plasmodium sporozoite infection. Although inhibition of either of these pathways leads to a reduction in infection, inhibition of PI3-kinase/Akt pathway caused a stronger effect, which correlated with a higher level of apoptosis in infected host cells. Altogether, the results show that the HGF/MET signalling requirement for infection is mediated by its anti-apoptotic signal effects. These results demonstrate for the first time that active inhibition of apoptosis in host cell during infection by Plasmodium is required for a successful infection.

Full Length and Delta Lactoferrin Display Differential Cell Localization Dynamics, but do not Act as Tumor Markers or Significantly Affect the Expression of Other Genes

Lactoferrin is a secreted protein related to transferrin. Lactoferrin indirectly protects host cells against foreign insults by killing bacteria, scavenging free iron, and binding to receptors required for viral invasion. However, lactoferrin is also proposed to act directly on cells as a transcription factor and tumor suppressor gene. In addition to full length lactoferrin, a truncated form, called delta lactoferrin, can also be produced by alternative splicing. We show here that transformed and nontransformed cells are equally able to express both full length and delta lactoferrin. Moreover, both forms of lactoferrin failed to substantially modulate the expression of other genes. Thus, lactoferrin does not seem to directly control gene expression or inhibit tumor cell growth.

A short consensus repeat-containing complement regulatory protein of lamprey that participates in cleavage of lamprey complement 3.

The prototype of the short consensus repeat (SCR)-containing C regulatory protein is of interest in view of its evolutionary significance with regard to the origin of the C regulatory system. Lamprey is an agnathan fish that belongs to the lowest class of vertebrates. Because it does not possess lymphocytes, it lacks Ig and consequently the classical C pathway. We identified an SCR-containing C regulatory protein from the lamprey. The primary structure predicted from the cDNA sequence showed that this is a secretary protein consisting of eight SCRs. This framework is similar to the alpha-chain of C4b-binding protein (C4bp). SCR2 and -3 of human C4bp are essential for C4b inactivation, and this region is fairly well conserved in the lamprey protein. However, the other SCRs of this protein are similar to those of other human C regulatory proteins. The lamprey protein binds to the previously reported lamprey C3b/C3bi deposited on yeast and cleaves lamprey C3b-like C3 together with a putative serum protease. The scheme resembles the C regulatory system of mammals, where factor I and its cofactor inactivate C3b. Unlike human cofactors, the lamprey protein requires divalent cations for C3b-like C3 cleavage. Its artificial membrane-anchored form protects host cells from lamprey C attack via the lectin pathway. Thus, the target of this protein appears to be C3b and/or its family. We named this protein Lacrep, the lamprey C regulatory protein. Lacrep is a member of SCR-containing C regulators, the first of its kind identified in the lowest vertebrates.

Functional analysis in serum from atypical Hemolytic Uremic Syndrome patients reveals impaired protection of host cells associated with mutations in factor H

A subgroup of patients with the most severe form of the Hemolytic Uremic Syndrome (HUS) presents mutations in the complement regulatory protein factor H. The functional analyses of the factor H mutant proteins purified from some of these patients have shown a specific defect in the capacity to control complement activation on cellular surfaces. Here, we show that these factor H-related complement regulatory defects can be detected in the patients’ serum with a simple hemolytic assay. Data obtained from HUS patients and control individuals indicate that this assay is a useful tool for the molecular diagnosis of factor H-related HUS.

Pharmacogenetic determinants of outcome in acute lymphoblastic leukaemia.

Present day paediatric co-operative group acute lymphoblastic leukaemia (ALL) protocols cure approximately 80% of patients, a result achieved largely through the use of risk-stratified therapies that employ multiple chemotherapy agents. These risk-based therapies utilize host and leukaemia traits to select the most appropriate therapy. However, these risk-stratified approaches predict therapy response imperfectly and an important fraction of patients experience relapse or therapy-related toxicity. Pharmacogenetics, the study of genetic variations in drug-processing genes and individual responses to drugs, may enable the improved identification of patients at higher risk for either disease relapse or chemotherapy-associated side effects. While the impact of genetic variation in the thiopurine-S-methyltransferase gene on ALL treatment outcome and toxicity has been extensively studied, the role of other polymorphisms remains less well known. This review summarizes current research on the impact of genetic variation in drug-processing genes in paediatric ALL and reviews important methodological and statistical issues presently challenging the field of pharmacogenetics.