Novel potential oncogenic and druggable mutations of FGFRs recur in the kinase domain across cancer types

Fibroblast growth factor receptors (FGFRs) are recurrently altered by single nucleotide variants (SNVs) in many human cancers. The prevalence of SNVs in FGFRs depends on the cancer type. In some tumors, such as the urothelial carcinoma, mutations of FGFRs occur at very high frequency (up to 60%). Many characterized mutations occur in the extracellular or transmembrane domains, while fewer known mutations are found in the kinase domain. In this study, we performed a bioinformatics analysis to identify novel putative cancer driver or therapeutically actionable mutations of the kinase domain of FGFRs. To pinpoint those mutations that may be clinically relevant, we exploited the recurrence of alterations on analogous amino acid residues within the kinase domain (PK_Tyr_Ser-Thr) of different kinases as a predictor of functional impact. By exploiting MutationAligner and LowMACA bioinformatics resources, we highlighted novel uncharacterized mutations of FGFRs which recur in other protein kinases. By revealing unanticipated correspondence with known variants, we were able to infer their functional effects, as alterations clustering on similar residues in analogous proteins have a high probability to elicit similar effects. As FGFRs represent an important class of oncogenes and drug targets, our study opens the way for further studies to validate their driver and/or actionable nature and, in the long term, for a more efficacious application of precision oncology.     

Prediction of personalized drugs based on genetic variations provided by DNA sequencing technologies

Recent reports of death and illness caused by adverse drug reactions have boosted rational drug design research. It has been shown through sequencing of the entire human genome that human genetic variations play a key role in adverse reactions to drugs as well as in differences in the effectiveness of drug treatments. The advent of high-throughput DNA sequencing technologies with bioinformatics of system biology have allowed the easy identification of genetic variations and all other pharmacogenetic variants in a single assay, thus permitting truly personalized drug treatment. This would be particularly valuable for many patients with chronic diseases who must take many medications concurrently. In this review, we have focused on pharmacogenomics for the prediction of variable drug responses between individuals with relevant genetic variations through new DNA sequencing technologies and provided directions for personalized drug therapy in the future.     

Pharmacogenomics of acute lymphoblastic leukemia

Pharmacogenomics of acute lymphoblastic leukemia (ALL) evolved rapidly in the past few years. Majority of recent findings concerns knowledge on key components of ALL treatment, 6-mercaptopurine and methotrexate. Leukemia is the most common cancer affecting children, with ALL comprising 80 % of all leukemia cases. Introduction of treatment protocols composed of several chemotherapeutic agents improved importantly survival in patients with ALL. Nevertheless, ALL is still the leading cause of cancer-related death in children. Interindividual differences in drug responses are an important cause of resistance to treatment and adverse drug reactions. Identifying pharmacogenomic determinants of drugs used in ALL treatment may allow for prospective identification of patients with suboptimal drug responses allowing for complementation of traditional treatment protocols by genotype-based drug dose adjustment.     

Polymorphic Cytochrome P450 Enzymes (CYPs) and Their Role in Personalized Therapy

The cytochrome P450 (CYP) enzymes are major players in drug metabolism. More than 2,000 mutations have been described, and certain single nucleotide polymorphisms (SNPs) have been shown to have a large impact on CYP activity. Therefore, CYPs play an important role in inter-individual drug response and their genetic variability should be factored into personalized medicine. To identify the most relevant polymorphisms in human CYPs, a text mining approach was used. We investigated their frequencies in different ethnic groups, the number of drugs that are metabolized by each CYP, the impact of CYP SNPs, as well as CYP expression patterns in different tissues. The most important polymorphic CYPs were found to be 1A2, 2D6, 2C9 and 2C19. Thirty-four common allele variants in Caucasians led to altered enzyme activity. To compare the relevant Caucasian SNPs with those of other ethnicities a search in 1,000 individual genomes was undertaken. We found 199 non-synonymous SNPs with frequencies over one percent in the 1,000 genomes, many of them not described so far. With knowledge of frequent mutations and their impact on CYP activities, it may be possible to predict patient response to certain drugs, as well as adverse side effects. With improved availability of genotyping, our data may provide a resource for an understanding of the effects of specific SNPs in CYPs, enabling the selection of a more personalized treatment regimen.     

Environmental mutagens may be implicated in the emergence of drug-resistant microorganisms

The emergence of drug-resistant microorganisms is an important medical and social problem. Drug-resistant microorganisms are thought to grow selectively in the presence of antibiotics. Most clinically isolated drug-resistant microorganisms have mutations in the target genes for the drugs. While any of the many mutagens in the environment may cause such genetic mutations, no reports have yet described whether these mutagens can confer drug resistance to clinically important microorganisms. We investigated how environmental mutagens might be implicated in acquired resistance to antibiotics in clinically important microorganisms, which causes human diseases. We selected mutagens found in the environment, in cigarette smoke, or in drugs, and then exposed Pseudomonas aeruginosa to them. After exposure, the incidence of rifampicin- and ciprofloxacin-resistant P. aeruginosa strains markedly increased, and we found mutations in genes for the antibiotic-target molecule. These mutations were similar to those found in drug-resistant microorganisms isolated from clinical samples. Our findings show that environmental mutagens, and an anticancer drug, are capable of inducing drug-resistant P. aeruginosa similar to strains found in clinical settings.     

Management of Incidental Findings in the Era of Next-generation Sequencing

Next-generation sequencing (NGS) technologies allow for the generation of whole exome or whole genome sequencing data, which can be used to identify novel genetic alterations associated with defined phenotypes or to expedite discovery of functional variants for improved patient care. Because this robust technology has the ability to identify all mutations within a genome, incidental findings (IF)- genetic alterations associated with conditions or diseases unrelated to the patient’s present condition for which current tests are being performed- may have important clinical ramifications. The current debate among genetic scientists and clinicians focuses on the following questions: 1) should any IF be disclosed to patients, and 2) which IF should be disclosed – actionable mutations, variants of unknown significance, or all IF? Policies for disclosure of IF are being developed for when and how to convey these findings and whether adults, minors, or individuals unable to provide consent have the right to refuse receipt of IF. In this review, we detail current NGS technology platforms, discuss pressing issues regarding disclosure of IF, and how IF are currently being handled in prenatal, pediatric, and adult patients.     

Towards Precision Medicine: Advances in Computational Approaches for the Analysis of Human Variants

Variations and similarities in our individual genomes are part of our history, our heritage, and our identity. Some human genomic variants are associated with common traits such as hair and eye color, while others are associated with susceptibility to disease or response to drug treatment. Identifying the human variations producing clinically relevant phenotypic changes is critical for providing accurate and personalized diagnosis, prognosis, and treatment for diseases. Furthermore, a better understanding of the molecular underpinning of disease can lead to development of new drug targets for precision medicine. Several resources have been designed for collecting and storing human genomic variations in highly structured, easily accessible databases. Unfortunately, a vast amount of information about these genetic variants and their functional and phenotypic associations is currently buried in the literature, only accessible by manual curation or sophisticated text text-mining technology to extract the relevant information. In addition, the low cost of sequencing technologies coupled with increasing computational power has enabled the development of numerous computational methodologies to predict the pathogenicity of human variants. This review provides a detailed comparison of current human variant resources, including HGMD, OMIM, ClinVar, and UniProt/Swiss-Prot, followed by an overview of the computational methods and techniques used to leverage the available data to predict novel deleterious variants. We expect these resources and tools to become the foundation for understanding the molecular details of genomic variants leading to disease, which in turn will enable the promise of precision medicine.     

The role of pharmacogenetics in treating central nervous system disorders

Symptoms of central nervous system (CNS) disorders include abnormalities in both physical and psychological domains. Many drugs indicated for the treatment of CNS disorders are fraught with side effects and/or poor efficacy which impact patients' quality of life and drives non-compliance. Moreover, for many CNS drugs such as antidepressants and antipsychotics, it takes time to determine whether a particular drug is efficacious in an individual patient. To optimize drug treatment for each patient, prescribing physicians often need to raise or lower doses, switch drug classes, or prescribe additional drugs to mitigate side effects, often in a "trial and error" fashion. Pharmacogenetic (PGx) testing, particularly in the realm of CNS therapy, can reduce the unpredictability of this process. By determining a patient's genetic profile, individual therapy parameters may be predicted pre-treatment for drug efficacy, optimal drug dose, and the risk of adverse drug reactions (ADRs). The intent of this review is to highlight the power of PGx testing to predict the likelihood of ADRs and efficacy during the treatment of the following CNS disorders: epilepsy, bipolar disorder, schizophrenia and depression.     

Differences in Efficacy and Safety of Pharmaceutical Treatments between Men and Women: An Umbrella Review

Being male or female is an important determinant of risks for certain diseases, patterns of illness and life expectancy. Although differences in risks for and prognoses of several diseases have been well documented, sex-based differences in responses to pharmaceutical treatments and accompanying risks of adverse events are less clear. The objective of this umbrella review was to determine whether clinically relevant differences in efficacy and safety of commonly prescribed medications exist between men and women. We retrieved all available systematic reviews of the Oregon Drug Effectiveness Review Project published before January 2010. Two persons independently reviewed each report to identify relevant studies. We dually abstracted data from the original publications into standardized forms. We synthesized the available evidence for each drug class and rated its quality applying the GRADE (Grading of Recommendations Assessment, Development and Evaluation) approach. Findings, based on 59 studies and data of more than 250,000 patients suggested that for the majority of drugs no substantial differences in efficacy and safety exist between men and women. Some clinically important exceptions, however, were apparent: women experienced substantially lower response rates with newer antiemetics than men (45% vs. 58%; relative risk 1.49, 95% confidence interval 1.35–1.64); men had higher rates of sexual dysfunction than women while on paroxetine for major depressive disorder; women discontinued lovastatin more frequently than men because of adverse events. Overall, for the majority of drugs sex does not appear to be a factor that has to be taken into consideration when choosing a drug treatment. The available body of evidence, however, was limited in quality and quantity, confining the range and certainty of our conclusions.     

Disparate in vivo efficacy of FTY720 in xenograft models of Philadelphia positive and negative B-lineage acute lymphoblastic leukemia

Most patients with acute lymphoblastic leukemia (ALL) respond well to standard chemotherapy-based treatments. However a significant proportion of patients, particularly adult patients, relapse with the majority dying of leukemia. FTY720 is an immunosuppressive drug that was recently approved for the treatment of multiple sclerosis and is currently under pre-clinical investigation as a therapy for a number of hematological malignancies. Using human ALL xenografts in NOD/SCIDγc(-/-) mice, we show for the first time that three Ph(+) human ALL xenografts responded to FTY720 with an 80 ± 12% (p = 0.048) reduction in overall disease when treatment was commenced early. In contrast, treatment of mice with FTY720 did not result in reduced leukemia compared to controls using four separate human Ph(-) ALL xenografts. Although FTY720 reactivated PP2A in vitro, this reactivation was not required for death of Ph(-) ALL cells. The plasma levels of FTY720 achieved in the mice were in the high nanomolar range. However, the response seen in the Ph(+) ALL xenografts when treatment was initiated early implies that in vivo efficacy may be obtained with substantially lower drug concentrations than those required in vitro. Our data suggest that while FTY720 may have potential as a treatment for Ph(+) ALL it will not be a useful agent for the treatment of Ph(-) B-ALL.     

Risk assessment and mitigation strategies for reactive metabolites in drug discovery and development

Drug toxicity is a leading cause of attrition of candidate drugs during drug development as well as of withdrawal of drugs post-licensing due to adverse drug reactions in man. These adverse drug reactions cause a broad range of clinically severe conditions including both highly reproducible and dose dependent toxicities as well as relatively infrequent and idiosyncratic adverse events. The underlying risk factors can be split into two groups: (1) drug-related and (2) patient-related. The drug-related risk factors include metabolic factors that determine the propensity of a molecule to form toxic reactive metabolites (RMs), and the RM and non-RM mediated mechanisms which cause cell and tissue injury. Patient related risk factors may vary markedly between individuals, and encompass genetic and non-genetic processes, e.g. environmental, that influence the disposition of drugs and their metabolites, the nature of the adverse responses elicited and the resulting biological consequences. We describe a new strategy, which builds upon the strategies used currently within numerous pharmaceutical companies to avoid and minimize RM formation during drug discovery, and that is intended to reduce the likelihood that candidate drugs will cause toxicity in the human population. The new strategy addresses drug-related safety hazards, but not patient-related risk factors. A common target organ of toxicity is the liver and to decrease the likelihood that candidate drugs will cause liver toxicity (both non-idiosyncratic and idiosyncratic), we propose use of an in vitro Hepatic Liability Panel alongside in vitro methods for the detection of RMs. This will enable design and selection of compounds in discovery that have reduced propensity to cause liver toxicity. In vitro Hepatic Liability is assessed using toxicity assays that quantify: CYP 450 dependent and CYP 450 independent cell toxicity; mitochondrial impairment; and inhibition of the Bile Salt Export Pump. Prior to progression into development, a Hepatotoxicity Hazard Matrix combines data from the Hepatic Liability Panel with the Estimated RM Body Burden. The latter is defined as the level of covalent binding of radiolabelled drug to human hepatocyte proteins in vitro adjusted for the predicted human dose. We exemplify the potential value of this approach by consideration of the thiazolidinedione class of drugs.     

Clinical implications of pharmacogenetics of cytochrome P450 drug metabolizing enzymes

For many drugs, pharmacogenetic polymorphisms are known affecting biotransformation and clinical outcome. The clinical importance of these variants depends on allele-frequency and the effect size of the clinical outcome parameters. Further, it depends on the therapeutic range of the drug which is affected, on predictability of drug response as well as on duration until onset of therapeutic efficacy. Consequences which arise from genotyping might be: adjustment of dose according to genotype, choice of therapeutic strategy or even choice of drug. In antidepressant drug treatment, most drugs are metabolized via the polymorphic cytochrome P450 enzyme CYP2D6. Huge differences in pharmacokinetic parameters have been consistently shown for many tricyclics, some SSRIs, and other antidepressant drugs whereas the effects on therapeutic efficacy and adverse events have been described controversially. In cardiovascular disease, oral anticoagulants, nonsteroidal anti-inflammatory drugs, oral hypoglycemic drugs and other drugs are affected by genetic polymorphisms of the cytochrome P450 drug metabolizing enzyme CYP2C9. Studies in patients or healthy volunteers revealed up to 10-fold differences in pharmacokinetic parameters due to genetic polymorphisms of CYP2C9. Pharmacogenetics based dose adjustments are one tool to individualize drug treatment according to genetic factors. They can be derived from pharmacokinetic data with the aim to obtain equal drug concentrations in each individual. Prospective validation of dose adjustments based on pharmacogenetics should be performed before routine application of such strategies. A controlled prospective clinical trial with one arm receiving genotype-based dose adjustments and the other arm receiving therapy as usual will elucidate the benefit of pharmacogenomics-based individualization of certain drug therapies.     

The Molecular Basis of Drug Resistance against Hepatitis C Virus NS3/4A Protease Inhibitors

Hepatitis C virus (HCV) infects over 170 million people worldwide and is the leading cause of chronic liver diseases, including cirrhosis, liver failure, and liver cancer. Available antiviral therapies cause severe side effects and are effective only for a subset of patients, though treatment outcomes have recently been improved by the combination therapy now including boceprevir and telaprevir, which inhibit the viral NS3/4A protease. Despite extensive efforts to develop more potent next-generation protease inhibitors, however, the long-term efficacy of this drug class is challenged by the rapid emergence of resistance. Single-site mutations at protease residues R155, A156 and D168 confer resistance to nearly all inhibitors in clinical development. Thus, developing the next-generation of drugs that retain activity against a broader spectrum of resistant viral variants requires a comprehensive understanding of the molecular basis of drug resistance. In this study, 16 high-resolution crystal structures of four representative protease inhibitors – telaprevir, danoprevir, vaniprevir and MK-5172 – in complex with the wild-type protease and three major drug-resistant variants R155K, A156T and D168A, reveal unique molecular underpinnings of resistance to each drug. The drugs exhibit differential susceptibilities to these protease variants in both enzymatic and antiviral assays. Telaprevir, danoprevir and vaniprevir interact directly with sites that confer resistance upon mutation, while MK-5172 interacts in a unique conformation with the catalytic triad. This novel mode of MK-5172 binding explains its retained potency against two multi-drug-resistant variants, R155K and D168A. These findings define the molecular basis of HCV N3/4A protease inhibitor resistance and provide potential strategies for designing robust therapies against this rapidly evolving virus.     

Molecular dynamics of G6PD variants from sub-Saharan Africa

Precision medicine uses genomic guidance to improve drug treatment safety and efficacy. Prior knowledge of genetic variant impact can enable such strategies, but current knowledge of African variants remains scarce. G6PD variants are linked to haemolytic adverse effects for a number of drugs commonly used in African populations. We have investigated a set of G6PD variants with structural bioinformatics techniques to further characterise variants with known effect, and gain insights into variants with unknown impact. We observed wide variations in patterns of root-mean-square deviation between wild-type and variant structures. Variants with known, highly deleterious impact show structural effects which may likely result in the destabilisation of the G6PD homodimer. The V68M and N126D variants (which are both common across African populations, and together form the A- haplotype) induce large conformational shifts in the catalytic NADP+ binding domain. We observed a greater impact for the haplotype than for each of the individual variants in these cases. A novel African variant (M207T) shows the potential to disrupt interactions within the protein core, urging further investigation. We explore how characterising the molecular impact of African G6PD variants can enable advanced strategies for precision medicine, as well as impact the use of novel therapeutics aiming to treat G6PD deficiency. This knowledge can assist in bridging current knowledge gaps, and aid to facilitate precision medicine applications in African populations.     

Suppression of premature termination codons as a therapeutic approach

In this review, we describe our current understanding of translation termination and pharmacological agents that influence the accuracy of this process. A number of drugs have been identified that induce suppression of translation termination at in-frame premature termination codons (PTCs; also known as nonsense mutations) in mammalian cells. We discuss efforts to utilize these drugs to suppress disease-causing PTCs that result in the loss of protein expression and function. In-frame PTCs represent a genotypic subset of mutations that make up ~11% of all known mutations that cause genetic diseases, and millions of patients have diseases attributable to PTCs. Current approaches aimed at reducing the efficiency of translation termination at PTCs (referred to as PTC suppression therapy) have the goal of alleviating the phenotypic consequences of a wide range of genetic diseases. Suppression therapy is currently in clinical trials for treatment of several genetic diseases caused by PTCs, and preliminary results suggest that some patients have shown clinical improvements. While current progress is promising, we discuss various approaches that may further enhance the efficiency of this novel therapeutic approach.     

Systematic analysis of somatic mutations in phosphorylation signaling predicts novel cancer drivers

Large‐scale cancer genome sequencing has uncovered thousands of gene mutations, but distinguishing tumor driver genes from functionally neutral passenger mutations is a major challenge. We analyzed 800 cancer genomes of eight types to find single‐nucleotide variants (SNVs) that precisely target phosphorylation machinery, important in cancer development and drug targeting. Assuming that cancer‐related biological systems involve unexpectedly frequent mutations, we used novel algorithms to identify genes with significant phosphorylation‐associated SNVs (pSNVs), phospho‐mutated pathways, kinase networks, drug targets, and clinically correlated signaling modules. We highlight increased survival of patients with TP53 pSNVs, hierarchically organized cancer kinase modules, a novel pSNV in EGFR, and an immune‐related network of pSNVs that correlates with prolonged survival in ovarian cancer. Our findings include multiple actionable cancer gene candidates (FLNB, GRM1, POU2F1), protein complexes (HCF1, ASF1), and kinases (PRKCZ). This study demonstrates new ways of interpreting cancer genomes and presents new leads for cancer research.     

Suppression of premature termination codons as a therapeutic approach

In this review, we describe our current understanding of translation termination and pharmacological agents that influence the accuracy of this process. A number of drugs have been identified that induce suppression of translation termination at in-frame premature termination codons (PTCs; also known as nonsense mutations) in mammalian cells. We discuss efforts to utilize these drugs to suppress disease-causing PTCs that result in the loss of protein expression and function. In-frame PTCs represent a genotypic subset of mutations that make up ~11% of all known mutations that cause genetic diseases, and millions of patients have diseases attributable to PTCs. Current approaches aimed at reducing the efficiency of translation termination at PTCs (referred to as PTC suppression therapy) have the goal of alleviating the phenotypic consequences of a wide range of genetic diseases. Suppression therapy is currently in clinical trials for treatment of several genetic diseases caused by PTCs, and preliminary results suggest that some patients have shown clinical improvements. While current progress is promising, we discuss various approaches that may further enhance the efficiency of this novel therapeutic approach.     

Pharmacogenomics of Drug Metabolizing Enzymes and Transporters:Relevance to Precision Medicine

The interindividual genetic variations in drug metabolizing enzymes and transporters influence the efficacy and toxicity of numerous drugs. As a fundamental element in precision med-icine, pharmacogenomics, the study of responses of individuals to medication based on their genomic information, enables the evaluation of some specific genetic variants responsible for an individual’s particular drug response. In this article, we review the contributions of genetic polymorphisms to major individual variations in drug pharmacotherapy, focusing specifically on the pharmacoge-nomics of phase-I drug metabolizing enzymes and transporters. Substantial frequency differences in key variants of drug metabolizing enzymes and transporters, as well as their possible functional consequences, have also been discussed across geographic regions. The current effort illustrates the common presence of variability in drug responses among individuals and across all geographic regions. This information will aid health-care professionals in prescribing the most appropriate treatment aimed at achieving the best possible beneficial outcomes while avoiding unwanted effects for a particular patient.