Keywords

9.1 Introduction

Pharmacogenetics is the study of heritable diversified traits on subject’s genome that explicate their pharmacological variability in drug response and toxicity. Pharmacogenetics became major bioscience of concern in the 1950s when clinical observations of tangled drug response in individuals were found in the relevance of inheritance and were postulated on simple Mendelian inheritance patterns [76]. Friedrich Vogel in 1959 was the first who coined the term pharmacogenetics [107]. Genetic polymorphism is the basic unit of pharmacogenetics. Over the past two decades, plenty of research and discoveries have been made since DNA sequencing has gotten persuasive and generally accessible. Around 30–90% of DNA in the human genome discovered has regions of repetitive DNA that are highly polymorphic in nature [116]. It is estimated that there are 3 billion nucleotides pairs on DNA in the double helix of 23 pairs of chromosomes, the likelihood of polymorphic DNA and diversity in people is so obvious.

Although the distinct drug responses among individuals are multifactorial and include both genetic and environmental factors which were remarkably found to be linked with genetic polymorphisms in drug metabolizing enzymes, drug-molecule transporters, drug receptor as well as target sites [26]. Since the launch of the human genome project, a massive amount of data on the occurrence of various polymorphisms on the human genome has been obtained [142].

9.2 Genetic Polymorphism

Polymorphism is a Greek word comprised of “poly” means many/multiple and “morph” means forms/variety. The term “polymorphism” was used during the early stage of clinical observations as altered behavior of drug and was explained as sub-groups of the population which are displaying a distinct drug metabolic behavior from the majority and encompassing at least 1% or more in the given population [62, 100]. Phenotypes are observable differences in organisms controlled by DNA. These variations in DNA sequence or polymorphism (genotypes) make the individuals unique in appearance, risk of development of various diseases, response to drugs (xenobiotics), and the pathogens, etc. [135]. The altered gene expression may produce proteins that may correlate to individual’s phenotypic status, disease idiopathy and a different response to drugs.

The human genome is 99.9% identical with minute difference of 0.1%. This difference is accountable for the adapted phenotypic diversity within the geo-environmental range. Single nucleotide polymorphisms (SNPs) are most frequent type of polymorphism, involving variations in a single base pair. Other polymorphic variants can be substantially bigger, involving extensive regions of DNA [1].

9.2.1 SNPs and Polymorphism

SNPs are the simplest form of genetic difference among individuals with an alteration in a DNA structure building block unit, the nucleotide; i.e., A, T, C, or G (adenine, thymine, cytosine, and guanine, respectively). More than 325 million SNPs are known on the human DNA strands out of which 15 million are present at frequencies of 1% or higher across the populace of the world [135]. SNPs are used for gene discovery and mapping, evolutionary biology studies, prediction of drug activity, diagnostic tests, and pharmacogenomic studies [57, 121]. Example of SNP-induced diseases is sickle cell anemia and cystic fibrosis [72]. Due to complex inheritance patterns of many genetic diseases like diabetes, cardiovascular and many cancers, many geneticists like to focus on SNPs in order to understand the inheritance pattern and pathogenic pathway of the disease as well as involvement of genetics in drugs responses which allows liberty to design therapeutic strategies with respect to genetic differences at individual level. SNPs in the regulatory and coding regions of a gene shown functional consequences. Linkage between a phenotype and a functional variant in individuals can be identified directly through SNP markers. This characteristic makes SNPs ideal for high-density genetic-markers maps essentially to separate complex genetic traits. SNPs occur in the human genome at the frequency of every 300–2000 base pairs where every individual carries two copies of each gene [90]. Most of the SNPs in non-coding regions of the genome are functionally silent. However, few SNPs are biologically functional which can alter protein structure or expression during sequencing. Such SNPs are the soul of human diversity in both health and disease. The identification procedure of these SNPs is an ongoing process that particularly are relevant to any disease. Genetic profiling based on such SNPs may act as a fingerprint that gives complete information about a subject’s susceptibility to different diseases and response to different drugs [7].

9.2.1.1 SNPs in Pharmacogenetics

SNPs are key factors in determining an individual's vulnerability to a variety of diseases like high blood pressure, diabetes, cardiovascular diseases, neurological disorders, and especially cancers, despite the fact that these disorders are multifactorial and include genetic and nongenetic factors [78]. Individual responses to drugs, including efficacy and adverse drug reactions (ADRs), are also highly variable, however, these variations may be influenced by the age, nutritional state, pathophysiology and severity of the disease, renal/hepatic function, medication interactions, and coexisting disorders [6]. Since the first documented occurrence in the 1950s, it has been well established that changes in drug metabolism and disposition at drug target receptors are caused by genetic polymorphism and are evidently inherited [48]. SNPs are abundant on the human genome and have become significant biological markers for mapping human maladies, heritable traits of population, diseases, and drug disposition as well as in developmental investigations.

9.2.1.2 Importance of SNPs in Cancer Pharmacogenetics

Cancer is a disease notorious for abnormal cell growth. The cancer patients show a heterogeneous pattern of response to many of the chemotherapeutic agents [47]. Scrutinized personalized anticancer therapy paired with biomarker-based regimen has promising potential for optimal cure from chemotherapy with reduced chemotherapy-related fatality [119]. Clinically, prognostic and predictive biomarkers are used simultaneously for the facilitation of cancer diagnosis, cancer treatment, and determination of possible toxicity [8]. For example, pharmacogenetic markers are extensively used for predicting therapeutic responses to methotrexate and 5-fluorouracil, the drugs that target folate metabolism [146]. Many cancer biomarkers depicted predictive and prognostic characteristics but with the variety of prognostic, predictive, toxicity, or pharmacological biomarkers, there are fewer clinically useful predictive biomarkers for solid tumors [25]. Different researches found a correlation between SNPs and cancer. Gemignani et al. studies show that polymorphism in the dopamine receptor gene, i.e., DRD2 is linked with an increased risk of colorectal carcinoma [52]. Some other examples include response to gefitinib treatment in non-small cell lung cancer patients with an EGFR mutation, ERCC1 polymorphisms and cisplatin activity, UGT1A1 gene polymorphic forms and irinotecan neutropenia, thymidylate synthase (TS) gene polymorphisms and 5-FU sensitivity, and cytidine deaminase (CDA) genotype and response to gemcitabine [32].

9.2.2 Somatic Versus Germline Mutations in Oncology

Cancer is a diversified by-product of mutations occurring in the germline and somatic (tumor) genome. The germline and somatic cells’ genetic variations are the main factors for the prognosis of cancers [2]. Genetic factors influencing the response to a drug include both pharmacokinetic and pharmacodynamics factors (Fig. 9.1). Variations in tumor DNA influence the drug choice for chemotherapy and germline mutations influence the efficacy of a drug chiefly through alteration of pharmacokinetic parameters [112, 153].

Fig. 9.1
A flowchart. Genetic factors influencing the drug response lead to pharmacokinetics and pharmacodynamics. Pharmacokinetics leads to drug metabolizing enzymes and drug transporters. Pharmacodynamics leads to Drug therapeutic targets. They have efficacy and adverse effects.

Genetic factors influencing the efficacy and toxicity of the drugs

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Any cell in the body other than the germ cell (i.e., sperm and egg) is a somatic cell. Mutations in the somatic cells are acquired mutations that can be caused by intrinsic factors such as DNA damage, oxidative stress, replication errors, or mutations in proto-oncogenes or oncogenes, or extrinsic factors such as direct DNA gene damage by ultraviolet rays, X-rays, radiation exposure, toxic chemicals, heat variability, and/or further environmental variables. These changes are non-inheritable and are useful in research studies of different disease conditions, e.g., cancer or neurological disorder. Some somatic mutations damage the DNA and accumulate in the cancerous cells which serve as drug targets or prognostic markers [2, 113]. Somatic cell mutations are extremely rare as they do not provide atmosphere or enhance cell division or expansion to new genotype [28].

Somatic mutations affect the response of drugs as they keep changing due to pressure from cytotoxic therapy and genetic instability. For the assessment of drug efficacy, genetic sequencing for somatic mutations from tumor cells is regularly done throughout the treatment span as a distinct mechanism of resistance emerges from these mutations that may require the recommendation of different therapy.

Germline variations are constitutional mutations. These mutations, particularly SNPs, are valuable biomarkers for predicting drug-induced toxicity and response to medication. Tumor cell markers may identify possible germline mutations for inheritable cancer, however, confirmatory germline testing is indicated for genetic cancer prediction [21].

Cancer genetics hold a risk of inheritance and is also an important preventive factor. The oncologists and cancer geneticists utilize the information generated from genomic testing for risk reduction and prevention strategies. Currently, parallel sequencing of tumor and germline DNA is performed, which can provide information about therapeutic options and cancer predisposition [149].

9.2.3 Pharmacogenetic Biomarkers and Their Significance

Biomarkers are check-points or indicators in the biological system that are powerful tools to portray the entire picture of disease from the earliest manifestation to the terminal stages [98]. They are either predictive, prognostic, or surrogate biomarkers. Predictive and surrogate biomarkers are important in decision-making strategies of treatment [9]. Predictive and prognostic biomarkers are proteins or bio-molecules, that are measured in body fluids, used as indicators that help in the diagnosis, progression, or reappearance of cancer, and are important for designing personalized chemotherapy regimen by identifying response to therapy and progression of the disease [94]. Biomarkers are usually well categorized according to the demand. Biomarkers used during the drug discovery stages for the assessment of drug effects in preclinical and/or early clinical studies are known as pharmacological biomarkers [8]. SNPs act as genetic biomarkers because they are polymorphic alleles on DNA sequencing which are useful for pinpointing a progression of disease instead the cause of disease [135].

The main purpose of PGx knowledge is to optimize drug therapies. For this purpose, many DNA-based pharmacogenetic tests are performed for the detection of genetic variations associated with the risk of drug disposition. Biomarkers counterparts for therapy selection in targeted chemotherapeutics which are intended to attack tumor cells with specific identifiable protein structure considerably different from the normal cells. As a result, individual-specified chemotherapy which is carefully guided by PGx biomarkers is more selective for cancer cells than normal cells which results an improvement in prognosis of disease condition in cancer patients’ and reduce toxic effect events from chemotherapy in normal cells. Epidermal growth factor receptor EGFR, KRAS, HER2, and c-Kit are some of the examples [10]. The FDA labeled alleles that influence drug efficacy and toxicity as pharmacogenetics biomarkers [66]. The FDA has incorporated pharmacogenetics information into drug labels and provided a detailed list of drugs entitled with genetic biomarkers. The FDA has also emphasized the importance of pharmacogenetics for personalized medicine. For example, dosing recommendations based on CYP2D6 and TPMT polymorphism have been given for mercaptopurine, as these enzymes affect the drug metabolism [110].

PGx (pharmacogenomic and pharmacogenetic) labeling in FDA-approved drugs includes biomarkers as biomarker-drug pairs as either predictive or prognostic markers. These PGx biomarkers can be labeled according to their nomenclature systems, for example, cytochrome P450 (CYP) may be labeled main family of enzymes, subfamily, gene levels, and alleles variants, etc. including wild-type and mutant one with the allele also labeled for functional status (i.e., decreased, increased, normal or nonfunctional) [99].

9.2.4 Anticancer Drugs and Pharmacogenetics of Enzymes

Alteration in proteins or enzymes involved in drug targets (i.e., pharmacodynamics–PD) and drug metabolism and transport (i.e., pharmacokinetics–PK) are key factors for drug response variation [33, 45].

All drugs undergo pharmacokinetic changes before imparting pharmacodynamic effects. The drug pharmacokinetics primarily involves phase I modification oxidation, reduction, and hydrolysis reactions primarily by CYP-450 family of enzymes, conjugation in phase II modification occurs through enzymes like glutathione-S-transferases (GSTs) and uridine diphosphate glucuronosyltransferases (UGTs), and elimination usually through bile or urine [74]. SNPs in drug-metabolizing enzymes can be classified as the phenotype of the extensive-metabolizer to the poor-metabolizer which either intensifies the toxicity or nullifies the drug response [5].

Enzymes of anabolic and catabolic pathways for purine and pyrimidine analogues, such as TPMT and DPD, and drug transporters are important targets in cancer therapy. Polymorphism in these targets greatly influences the pharmacokinetics of drugs by affecting the enzyme activity and may alter the drug response and extent of toxicity [15].

Genotyping an individual allows selecting the effective drug and dose for prophylactic or therapeutic use to optimize and enhance the effectiveness of therapy. Pharmacogenetics by identifying the hereditary variabilities that influence the drug responses helps to create proper phenotyping and genotyping tests. Utilization of this information is helpful to anticipate the effective and positive clinical results from dose-related toxicity [110, 136].

Adverse drug reactions are therapy-limiting factors that are due to interference with the target protein or due to activation or inhibition of therapeutic drug targets. A proper screening method and diagnostic biomarker can work wonders to control individual variability of drug toxicity and drug hypersensitivity [92].

9.3 Applying Pharmacogenetic Knowledge in Clinical Oncology

PGx research mainly emphasizes the genetic variations found in germline genome as well as in tumor genome. Germline variations help to forecast drug efficacy and toxic effects while somatic mutations help in selection of a better chemotherapeutic drug with improved therapeutic efficacy. Mainly these genetic variations are studied relative to gene expression for chemotherapy response and tumorigenesis [153]. Contemplative genetic and pharmacological knowledge has given profound benefits in cancer therapy by increasing therapeutic index and reducing toxicity and tumor response [23]. It is evidently well-recognized that DNA damage is a vital factor for cancer prognosis. Damaged DNA during repairing procedure leads to mutations that affect oncogenes and tumor suppressor genes. These genetic defects predispose various cancer types. However, this damaged DNA also provides an important avenue for chemotherapy [144]. Patient-genome and tumor-genome polymorphic variations not only affects the regulation of transport, retention, efflux of drugs, also determine the penetration into tumor tissue. Hence, drug-related toxicities depend on the germline mutations whereas tumor mutations play key role in dose-limiting factors in cancer management [79].

In clinical oncology, PGx information about genetic variations and polymorphism that affects the drug related toxicity, treatment response and survival from chemotherapy is utilized to develop safer and effective patient-tailored or targeted therapy. Various of the targeted patient-tailored approaches include hormone treatments, inhibitors of signal transduction, gene expression modulators, apoptosis enhancers, angiogenesis inhibitors, immune-therapies, and toxin delivery agents [46]. However, drug response alteration is not monogenic polymorphisms that encode for PK- or PD-related proteins, it is in fact a complex multifactorial and multigenetic process. For example, patients with wild-type KRAS tumors show response to EGFR-targeted therapies while mutated KRAS tumors and nonresponsive to monoclonal antibodies targeting EGFR receptors [37]. FDA has included gene expression studies in PGx studies in relevancy of tumorigenesis and chemotherapy response [153].

Pharmacogenetics is gene-drug interaction and pharmacogenomic is all-genes in genome-to-drug response which imparts great significance in oncology. Chemotherapeutic agents have a narrow therapeutic index and a high risk of the development of drug toxicity. Both somatic and germline genetic variations have role in cancer treatment however germline variations impart a key role in cancer risk and treatment outcome [102].

Genetic biomarkers (SNPs from somatic or germline mutations) are the alterations in the nucleotide sequences of DNA. These genetic traits are readily identifiable and are helpful to detect predisposed risk of heritable diseases in individuals of populace, ethnically different populations, and/or different species [135]. Some examples from somatic and germline cells are available in Table 9.1 with their respective pharmacogenetic biomarkers.

Table 9.1 Cancer pharmacogenetic biomarkers in FDA drug labeling
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9.4 Tools and Techniques for Pharmacogenetics Studies

To identify the genetic variations and understand their significance in tumorigenesis and treatment response several techniques and tools are available. Genome-wide association studies (GWAS) of cancer have databased huge number of variants associated with cancer susceptibility and/or genetics of cancer risk [133]. Similarly, advancement in genetic and molecular technologies helped to understand the molecular basis of genetic alteration and tumorigenesis. The Cancer Genome Atlas (TCGA) and the International Cancer Genome Consortium (ICGC) are serving well by providing comprehensive data on the nature of tumors from different cancers [152]. Dynamic and explorative efforts are being made from the data of both types of mutagenic risk-alleles to identify the associative link in carcinogenetic risk and/or may influence the development of any cancerous tumor or cancer type. Preliminarily, the identification, association, and influential relation of cancer and tumor are done on basic principles that analyze tumor data with germline cancer risk variant, the relationship of the patient genome to particular tumor type, and finally relationship between the tumor molecular data for germline association studies [50].

The Next Generation Sequencing (NGS) has performed sequencing of millions of DNA fragments and identified novel and rare mutants of cancer, also detects the carrier of cancer mutation in family, and provides the molecular rationale of precisely targeted therapy [58]. NGS technologies provide economical testing for genetic sequencing and are now widely applied in the diagnosis of genetic disorders, especially in cancer [95]. Germline variants are mostly either identified through candidate-gene approaches or genome-wide association studies (GWAS). The Cancer Genome Atlas (TCGA) and International Cancer Genome Consortium (ICGC) are vigilant in the identification of somatic mutations from genetic materials of cancer cells. NGS has enabled the incorporation of genomic data into the clinical utility [24]. For example, high-throughput genotypic technology can identify genes with germline mutations associated with susceptibility of cancer, e.g., there is elevated risk of breast and ovarian cancers with BRCA1 and BRCA2 genes inheritance. Genetic testing for mutations in these genes is done for the identification of the individuals susceptible to heritable cancers.

Cancer is also known as a disease of altered pathways. Mutations show inconsistent behavior, as sometimes high-impact coding site mutations show no functional significance while some somatic mutations from high-ratio of rare variants to common ones show clinical functional impact. Differential mutation analysis, another method to identify the significance of mutant genes in cancer, analyzes the significant driver mutations along with pathway analysis which potentially can uncover novel cancer pathways instead of addressing known cancer pathways [114].

Catalogue of Somatic Mutations in Cancer (COSMIC) provides a comprehensive database for studying the consequences of somatic cell mutations in human malignancies (https://cancer.sanger.ac.uk). It encompasses all genetic pathways by which somatic mutations cause cancer, including non-coding mutations, gene fusions, copy-number variants, and drug-resistant mutations, in addition to coding mutations [140].

9.5 Pharmacogenetics and Drug Development

The annotation of the human genome sequence with concomitant advancement in genomic technologies has facilitated the drug discovery and development process along with precise genetic data availability which can outline individual risks and benefits for specific therapeutic strategies. The clinical observational data, molecular information of targets and drugs, and understanding of biological pathways of disease are driven together to design and develop a better therapeutic regimen [118]. This knowledge is applied within the pharmaceutical industry along with research engines, in the relevance of genetic tools for the drug discovery process. The goal of developing a beneficial drug in the pharmaceutical industry is a well-outlined process and is termed the pharmaceutical pipeline [70]. SNPs are favorite molecular-markers in pharmacogenomics investigations because they affect drug-metabolizing pathways and/or ADRs, as well as, are linked to a variety of disorders, including cancer, autoimmune diseases, neuropsychiatry, and infectious diseases [7].

Pharmacological understanding of the drug disposition, i.e., pharmacokinetic parameters (PK) and pharmacodynamic parameters (PD) along with SNPs (pharmacogenetics; drug transporters and metabolizers) and advancement in gene expression technologies has opened new doors for drug development that can have high efficacy and selectivity to target and low toxicity. This is especially significant in cancers where both the germline and somatic genotypic variations that correlate with adverse events and drug efficacy profiles have been identified [49]. The new drug discovery process has evolved drastically over the past half-century from pharmacological effects process to chemically-modulated drugs to more precisely biologically-driven process, i.e., identification of biological pathway of disease as well as drug responses toward the development of the new drug as a forward drug discovery process [44].

9.5.1 Pharmacogenetics in Anticancer Drug Development

Cancer drugs development also follows the same traditional pathway which approximately takes 15 years although considerable efforts have been made by the research industry and FDA to reduce the timeline and cost of this procedure. Almost 90% of the potential new drugs that have passed through discovery and preclinical phase fail in clinical trials due to efficacy, safety, and toxicity issues. Implementation of PGx (pharmacogenomics) biomarkers in drug development have progressed the oncology therapeutics to more tailored treatment from the paradigm of fit-for-all to a more targeted and histology-oriented approach toward, namely Precision Medicine (PM) [139].

The application of PGx principles in treating certain cancers has beneficial outcomes and has a crucial role in cancer drug development [41, 154]. An integration of PGx principles with various phases of drug development is shown in Fig. 9.2.

Fig. 9.2
An illustration depicts cancer drug development. 1. Target discovery. 2. Pre-clinical phase. 3. Clinical development. 4. F D A approval and phase I V. 5. P G x biomarker identification. 6. Clinical qualification. 7. Clinical use. 8. Identification and development of tools to find the best candidate. 9. Phase 1, Screening of prognostic or predictive biomarkers. Phase 2, Screening of biomarkers with drug activity. Phase 3, Validation of biomarker. Clinical practice with further validation.

PGx application in each R&D phase of drug development

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There is a major discordance in phase II and phase III results in terms of efficacy, safety, and toxicity which imparts huge time and money losses. In 2014, AstraZeneca’s scientists proposed five “R’s”, as shown in Fig. 9.3, being the most important technical determinants for drug discovery and development success and pipeline quality which recently has shown considerably improved success rates in phase III trials [104].

Fig. 9.3
An illustration depicts the proposed 5 R's. 1. Right target. 2. Right patient. 3. Right tissue. 4. Right safety. 5. Right commercial potential.

Proposal of a “5-Rs” framework applied to cancer drug development

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Imatinib, a tyrosine kinase inhibitor used to treat chronic myelogenous leukemia, is a classic example of anticancer drug development utilizing PGx knowledge [153]. Similarly, crizotinib which is a tyrosine kinase inhibitor targeting ALK, demonstrated substantial results in phase I trials. This granted its accelerated approval to phase II trials by the FDA and is recently approved for ALK-positive relapsed or refractory, systemic anaplastic large cell lymphoma (ALCL) in pediatric patients of 1-year and older (www.fda.gov).

The advancement and development in strategic drug discovery technologies including high-throughput screening and structure-based design strategies has made it possible to design small-molecule targets that are target-based therapeutic agents. Such agents are expected to have high efficacy, sensitive to target selectivity, and exhibits less toxicity than the previous versions of anticancer agents [138].

Identification of prognostic, diagnostic, and predictive biomarkers will accelerate the process of drug development and approval with the possibility of post-approval risks management [86]. However, PGx incorporation along with drug development has added costs burdens to the already high-budget drug development process [40]. However, drug regularity authorities realized the potential benefits of PGx technologies, encourage the pharmaceutical industry to integrate PGx knowledge into drug development [87].

9.6 Genetic Factors Influencing Efficacy and Toxicity of Individual Chemotherapeutic Agents

The concept of “Chemical individuality of Man” was given by Sir Archibald Garrod that’s based on clinical observation and the concept of erring in the drug-metabolizing pathways that alters pharmacological effects of the drugs are inherited [101]. The potential overlapped linkage of metabolic pathways of disease (receptors), genetics (SNPs), and pharmacological effects (i.e., pharmacokinetic and pharmacodynamic effects) can be utilized to determine the peculiarities of a disease based on the differential response of drugs,as the drug targets the disease-causing gene [27].

In cancer, the remarkable inter-individual unpredictability in drug response was found to be linked with polymorphisms in drug-metabolizing enzymes, drug-molecule transporters, drug receptors sites, and drug target sites [26, 118]. Although inter-individual drug responses are influenced by numerous factors including genetic and environmental aspects. With the advancement of the Human Genome Project, substantial information, and data has been compiled for multiple polymorphic alleles and heritable variables on the human genomic DNA [142].

In the GWAS catalogue, about 93% of registered SNPs are in the non-coding regions and are known as regulatory SNPs that can affect gene splicing and transcription factor binding. Approximately 10 M SNPs are in coding or non-coding regions. For example, elevated levels of HDL are associated with a defect in cholesteryl ester transfer protein (CEPT), deactivating mutations in the Janus kinase 3 (JAK 3) gene results in the severe combination of the immune-deficient syndrome [106, 126].

Pharmacogenetic factors influencing pharmacokinetic and pharmacodynamics properties of anticancer drugs are summarized in Table 9.2.

Table 9.2 Genetic polymorphisms altering PK and PD of anticancer drugs
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9.6.1 Tamoxifen and Polymorphic CYP2D6

Tamoxifen, a selective estrogen-receptor modulator, has successfully been used for treatment and prevention of breast cancer. Oral tamoxifen citrate has been approved as endocrine therapy of breast cancer and has sustained since for over 25 years in estrogen-receptor (ER) positive breast cancer treatment and related conditions. Tamoxifen binds and thus block the ligand-binding domain of an ER from interaction with estrogen. This inhibits the ER associated transcriptional activation and subsequent tumor growth [96].

Hepatic phase I and II enzymes are responsible for the complex metabolism of tamoxifen. Tamoxifen is metabolized to its major primary metabolites: N-desmethyltamoxifen and 4-hydroxytamoxifen by hepatic cytochrome P450 enzymes, (CYPs) CYP3A4 (and CYP3A5), and highly polymorphic CYP2D6, respectively [39]. When the primary metabolites are oxidized, a pharmacologically active metabolite called 4-hydroxy-N-desmethyltamoxifen (endoxifen) is formed in large amounts. The binding affinities of endoxifen and 4-hydroxytamoxifen to ER and ER, as well as the reduction of ER-dependent gene expression and breast cancer cell proliferation, are similar. Tamoxifen and 4-hydroxytamoxifen stabilize ERα, and endoxifen reduces ERα protein levels in breast cancer cells by tagging it for degradation by proteasomes. These data, together with the fact that endoxifen plasma concentrations are 5–10 times higher than those of 4-hydroxytamoxifen, indicate that endoxifen is an important metabolite having approximately 50% high affinity for the estrogen receptors of tamoxifen [17, 89]. The efficacy of tamoxifen can be affected by the genetic polymorphism in CYP2D6 exhibiting two major alleles for poor metabolizers (PM of drug and extensive metabolizers (EM, where PM’s have low levels of endoxifen and need dose adjustments [71]. Hence, pharmacogenetics testing with pharmacokinetic consideration is required for achieving tamoxifen efficacy in breast cancer, however, there is a controversy for pre-emptive CYP2D6 genotyping due to conflicting results of the studies concerning the association between CYP2D6 genotype and tamoxifen-related clinical outcome [20].

Alternative anti-estrogen medication, such as letrozole, anastrozole, or exemestane, could be administered to postmenopausal women with early stage ER positive breast cancer who are anticipated to not react to tamoxifen based on their cytochrome P450 2D6 (CYP2D6) genotype. By blocking the aromatase enzyme CYP19A1, these drugs restrict the production of estrogenin extragonadal tissues [67].

9.6.2 Irinotecan and Polymorphic UDT1A1

Irinotecan is a topoisomerase inhibitor that has displayed anticancer effect in a range of solid tumors. It is a first-hand choice to treat colorectal, pancreatic, and small cell lung cancer [68]. Carboxylesterase metabolize irinotecan to its active form of 7-ethyl-10-hydroxy-camptothecin (SN-38). Elevated levels of SN-38 cause the side effects of irinotecan, such as diarrhea and leucopenia. Allelic variations on the genomic structures cause divergent expression of enzymatic functional capacity as metabolizers and transporter proteins which may interfere with the pharmacokinetics and pharmacodynamics of irinotecan [35]. SN-38 glucuronide is a polar product of SN-38 which is excreted in urine and bile after being inactivated by UDP-glucuronosyltransferase 1A1 (UGT1A1) in the liver. Thus, the rate of glucuronidation is a vital predictive factor in the development of toxicity with irinotecan. The enzyme UGT1A1 is highly polymorphic. UGT1A1∗6 and UGT1A1*28 are the most commonly investigated polymorphisms in relation to irinotecan metabolism. In variant allele UGT1A1*28, an extra TA repeat sequence is present within the T-A-T-A box of the CGT1A1 gene promotor. Allelic form UGT1A1*28 is linked with reduced transcription of UGT1A1 protein with lower titer of SN-38 glucuronidation. The patients homozygous for UgT1A1*28 variant have significant high levels of the active metabolite SA-38, and consequently are at elevated risk to present with adverse events like diarrhea and leucopenia with irinotecan therapy. Similarly, homozygous patients with UGT1A1*6 mutant allele are on the higher risk zone for adverse events with an increased systemic exposure to irinotecan and SN-38 [73, 151].

Irinotecan is a hydrophilic molecule with a large volume of distribution (Vd) in steady state, estimated over 400 L/m2 [157]. The lactone ring of 7-ethyl-10-hydroxy-camptothecin (SN-38) can be hydrolyzed to a carboxylate isoform at physiological pH. As a result, there is a pH-dependent equilibrium between these types [35]. Because only the lactone form of irinotecan has anticancer activity, a modest alteration in physiological pH could affect the drug’s pharmacokinetics and effectiveness [111].

9.6.3 6-Mercaptopurine and TPMT

6-Mercaptopurine (6-MP) is a very well-known cytotoxic, purine analogue agent from thiopurine family, frequently used in conditions like autoimmune diseases, inflammatory bowel disease as well as certain cancers like acute lymphoblastic leukemia (ALL) especially in children and chronic myeloid leukemia (CML). 6-MP is an inactive-prodrugs that requires activation to thioguanine nucleotides (TGN) that incorporate into DNA to cause cytotoxicity. 6-MP undergo conversion to 6-thioguanine nucleotides (6-TGN) by hypoxanthine guanine phosphoribosyl transferase enzyme (HGPRT) once it enters leukemic cells. The integration of 6-TGN into DNA or RNA causes the cytotoxic and immunosuppressive effects of 6-MP. The other two pathways involved in the inactivation of 6-MP are oxidation to thiouric acid catalyzed by xanthine oxidase enzyme and S-methylation of thiol moiety of 6-MP by thiopurine methyltransferase (TPMT), thus opposing the action of HGPRT.

Intracellular metabolism is key factor to keep a balance between efficacy and toxicity. Inter individual variation in clinical response and toxicity of 6-MP may result from variability in intracellular 6-MP disposition due to DNA point mutations affecting gene expression and/or protein functions of the genes involved in 6-MP metabolism.

The TPMT polymorphism is important in inter-individual variability in 6-MP response, with over 90% of population exhibiting high activity with only about 10% having intermediate, and 0.3% having low-to-undetectable enzyme activity. The important genetic variants accounted for the majority of the intermediate and low enzyme activity cases include TPMT*2 (239G > C), *3A (460G > A and 719A > G, *3B (460G > A). Traditional thiopurine doses have been linked to a high risk of developing hematological toxicity in TPMT-deficient patients, according to studies. Heterozygous patients for TPMT mutant variant require 30–50% lower thiopurine dosages than standard doses, and homozygous patients for TPMT deficient mutation require up to tenfold lower doses or the use of alternate medicines [134]. TMPT-deficient patients are at elevated risk of suffering from hematological toxicity if given frequent thiopurine dosages, according to several studies. The efflux transporters of the ATP-binding cassette superfamily, particularly the multidrug resistance associated protein 4 and 5, are another genetic component responsible for inter-individual variability in response to thiopurines treatment (MRP4 and MRP5). These two proteins regulate the intracellular quantities of cyclic nucleotides at the physiological level [3]. Overexpression of these transporters has been linked to thiopurine treatment resistance [155].

9.6.4 5-Flurouracil and TYMS, MTHFR, DPD

5-fluorouracil (5-FU), a fluorinated analog of uracil, is a key component of chemotherapeutic medicines used to treat solid tumors, such as colorectal (CRC), gastric cancer, and breast cancer, for palliative and adjuvant purposes [19, 56]. 5-FU, as well as its oral prodrugs capecitabine and tegafur have narrow therapeutic index, although tolerable develop severe toxicities like neutropenia, stomatitis, diarrhea, and hand-foot syndrome which may result in prolonged hospital stay and death may result in 0.5–1% of patients. The cytotoxic action of 5-FU requires its activation to 5-fluoro-2-deoxyuridine monophosphate (5-FdUMP). By inhibiting thymidylate synthase (TS), an enzyme essential for de novo pyrimidine production, 5-FdUMP reduces tumor cell proliferation. Many patients experience poor outcome from 5-FU treatment as a result of tumor recurrence after 5-FU therapy [38]. 5-FU is concomitantly used as 5-FU-based therapy like in FOLFOX therapy which includes 5-FU, leucovorin, and oxaliplatin or FOLFIRI consisting of 5-FU, leucovorin, and irinotecan, although empirically has provided boosted response rates to therapy to about 40–50% in CRC patients but unfortunately clinical studies showed this has not effectively prolonged disease-free survival in such patients [42, 55].

9.6.4.1 5-Fluorouracil and DPD

In the liver, the enzyme dihydropyrimidine dehydrogenase (DPD) converts at least 85 percent of 5-FU to dihydrofluorouracil. Several allelic variants in the DPD gene have been associated with reduced DPD enzymatic activity. Generally, 3–5% of carriers are heterozygous individuals and 0.1% are carriers are homozygous individuals of allelic variants of DPD gene that are probably associated with inactivation of enzyme function. A deficiency of DPD causes increased levels of 5-FdUMP leading to hematopoietic, gastrointestinal (GI) and neurological toxic events with standard doses of 5-FU. Previous investigations have shown significant results in determining the relationship of DPD enzyme deficiency and 5-FU toxicity; where 55% of patients with DPD-deficient enzyme suffered from grade-4 neutropenia compared to 13% of patients with a normal DPD activity (P = 0.01). Reduced DPD activity was also identified in peripheral blood mononuclear (PBM) cells in 39–59% of these patients. Furthermore, individuals with low DPD activity developed toxicity symptoms 10-days sooner than patients with normal DPD activity [22]. Studies on DPYD, the gene that encodes DPD, from the patients who developed severe 5FU-associated toxicity have identified 11 mutations including a splice-site mutation, a nonsense mutation, i.e., E386X, four missense mutations and five polymorphic alleles. The most common mutation altering DPD activity is the splice-site mutation which is IVS14 + 1G → A. As 5-FU is a widely used chemotherapeutic agent in cancer patients and the associated toxicity is severe; analysis of the DPD activity in PBM cells or screening for the IVS14 + 1G → A mutation has been routinely suggested as pharmacogenetic protocol prior to the start of 5-FU treatment [147].

9.6.4.2 5-Fluorouracil and TYMS

The cytotoxic effect of 5-FU is due to an impairment of de novo nucleotide synthesis. In the presence of methylene tetrahydrofolate reductase, 5-FdUMP inhibits the activity of thymidylate synthase (TS) enzyme by inhibiting the methylation of deoxyuridylic acid (dUMP) to deoxythymidylic acid (dTMP) by creating a stable covalent complex. 5-FU limits purine synthesis via decreasing TS activity, resulting in decreased DNA replication and repair and tumor cell growth suppression. Nucleotides can be synthesized by cells in two ways: de novo synthesis and salvage synthesis. Nucleosides and nucleobases must first be transferred across the cell membrane by nucleoside transporter proteins before they can be salvaged. Human equilibrative nucleoside transporter 1 (hENT1) is the most common nucleoside transporter which is also closely linked to transport and pharmacological activity of numerous other drugs [123]. The in vitro cell viability studies on colorectal tissues have exhibited high expression of hENT1 demonstrated and resistance to 5-FU treatment [132].

9.6.4.3 5-Fluorouracil and MTHFR

Methylenetetrahydrofolate (i.e., MTHFR) is an enzyme pivotal for the regulation of intracellular pool of folate for nuclei acid and protein synthesis. It irreversibly catalyzes the conversion of 5,10-methylenetetrahydrofolate (5,10-MTHF) to 5-methyltetrahydrofolate, which assists in homocysteine re-methylation to methionine. The pyrimidine and DNA synthesis are halted by the complex produced by FdUMP which is an active metabolite of fluorouracil, thymidylate synthetase, and 5,10-MTHF. It is hypothesized that MTHFR enzyme activity is indirect proportional to 5,10-MTHF and fluorouracil cytotoxicity. 677C > T variant of MTHFR polymorphic alleles is predominantly linked to clinical outcome where 677TT genotype showed more toxicity to 5-FU in advanced colorectal cancer [36]. According to studies, advanced gastric cancer patients receiving fluorouracil-based therapy should have their TS and MTHFR polymorphisms genotyped as pharmacogenetic protocols. Individuals with MTHFR 677TT had a much greater survival rate than patients with CT or CC, according to the findings [125]. Another study demonstrated that MTHFR polymorphisms are not clinically relevant for cancer treatment as only homozygous carriers of MTHFR polymorphisms probably exhibit functional variations in folate metabolism if folate levels are low in vivo [4].

9.6.5 Methotrexate and SLCO1B1, TYMS

For the treatment of rheumatoid arthritis (RA) and acute lymphoblastic leukemia (ALL), methotrexate (MTX) is the most commonly utilized medication. Nonetheless, MTX can cause serious dose-limiting side effects and organ toxicity. The reduced folate carrier (SLC19A1) or the solute carrier organic anion transporter B1 (SLCO1B1) transports MTX, a structural analogue of folic acid, into the cell [53]. A polymorphism (rs1051266) at codon 27 of the gene coding for SLC19A1 protein with resultant substitution of arginine for histidine has been identified, although the functional significance of this SNP is not yet established.

Homozygous patients for the rs1051266 variation exhibited greater plasma MTX levels compared to patients of other genotypes in one trial of 204 children with leukemia receiving MTX [85]. Despite this, genome-wide association studies have found no evidence of a link between the SLC19A1 polymorphism and MTX pharmacokinetics.

The SLCO1B1 gene is almost entirely expressed in the liver. SLCO1B1 has been identified as the only gene linked with MTX clearance in genome-wide association studies. Endogenous chemicals like bilirubin and estrogens, as well as medicines like statins and MTX, are substrates of the SLCO1B1 transporter, which is situated at the basolateral membrane of the hepatocyte. The SLCO1B1 SNP rs4149056, which encodes a T521C change, has been linked to lower MTX clearance across the board. The rs4149056 common variant is found in two ∗ alleles: *5 (no additional coding variants) and *15 (with rs2306283 A > G). *23 and *31 are two further reduced function alleles linked to poor MTX clearance and in vitro transport. *14 and *35, respectively, were alleles linked to enhanced MTX clearance and transporter expression. Many ∗ alleles carry the rs2306283 A > G mutation, which is linked to enhanced transporter expression [141]. Thymidylate synthase (TYMS) is a crucial enzyme in DNA synthesis that produces purines de novo. The enzyme is inhibited by MTX, which hinders the synthesis of deoxythymidine monophosphate. Low TYMS expression in lymphoblastic leukemia cells is linked to lessened MTX’s antileukemic effect and an increased risk of relapse. Within the TYMS gene’s 5′-untranslated region,a tandem repeat sequence with a variable number of 28 base pair repeats has been discovered [95]. These repeats appear to function as enhancers, as more repeat sequences boost both mRNA expression and enzyme activity [115]. Individuals homozygous for three copies of the repeat required larger doses of MTX (>6 mg/week) than patients homozygous for two copies, according to a Japanese study [93].

9.6.6 Platinum Compounds and ERCC1,2, ATP7B

Platinum compounds, i.e., cisplatin, carboplatin, and oxaliplatin are popular antitumor drugs. They are standard first-line choice-of-chemotherapy agents for advanced lung cancer, breast cancer, and colorectal cancer, etc. They cause cell apoptosis through formation of DNA adducts by crosslinking DNA strand and abates cell growth in active tumors. The cell replication system has an active DNA repair system. The genes responsible for DNA repair show genetic variations. Due to presence of polymorphic genes, this DNA repair pathway is important and promising predictor to determine the effective treatment outcomes from these agents as well as the various pathways responsible in drug resistance development. As for instance, excision repair cross-complementing enzyme group 1, i.e., ERCC1, compared with other nucleotide excision repair, i.e., NER genes, is prominently involved in developing cisplatin resistance. As ERCC1 is responsible in NER pathway for DNA repair through single-stranded DNA breakage and forms a complex which is structure-specific endonuclease with xeroderma pigmentosum complementation group-F (XPF, also known as ERCC4) [158]. ERCC1 has been identified with several polymorphic forms with varied DNA repair capacities.

Platinum compounds are standard chemotherapy regimen for advanced non-small cell lung cancer (i.e., NSCLC). The patients having K751Q-ERCC2 genotype showed slower disease progression with cisplatin-treated NSCLC than those harbored the K751Q-ERCC2 genotype. The KK-type homozygous patients showed significant chemotherapy benefits and significant longer survival than those patients with heterozygous KK-type or QQ-homozygous genotype. However, the reported data shows confliction of ERCC2/XPD variant alleles responsible for decreased overall survival in NSCLC patients treated with cisplatin. Similarly, a silent C118T SNP is reportedly associated with lowered mRNA production in ovarian carcinoma cell lines. Studies have reported that homozygous carrier patients of ERCC1-118C allele have shown suggestively better survival rate. Also in colorectal carcinoma, SNP-K751Q of the ERCC2-XPD (xeroderma pigmentosum group D gene) in peripheral blood lymphocytes was of prognostic significance in patients treated with combination of 5-FU and oxaliplatin. Also the presence of a nonsynonymous SNP, replacing a lysine with glutamine at codon 751 of the XPD protein, was reportedly associated with treatment outcome in patients with metastatic colorectal cancer [120].

ATP7A and ATP7B are important copper-transporting ATPases that contribute to platinum compounds resistance by regulating drug efflux. ATP7B overexpression show poor prognosis of platinum-agent treated patients in different cancerous conditions including epidermoid carcinoma cells due to increased rate of cisplatin efflux and cisplatin resistance [122].

Mutations in the genes of DNA repair system leads to defective mismatch repair (i.e., MMR). This also hinders with cisplatin binding to DNA. Also, this MMR defectiveness causes microsatellite instability (i.e., MSI) which is useful to demonstrate genome instability. The MSI + positive shows significant resistance to chemotherapy and poor prognosis of disease in metastatic germ cell tumors [65].

Tumor protein 53 (TP53) gene makes a tumor suppressor protein. Better expression of p53 protein (wild-type) is directly related to the level of chemotherapeutic response from DNA damaging agents. Mutations in TP53 weaken the of apoptotic pathways and significantly decrease the levels of the BAX gene (Bcl-2 associated X-protein, the apoptosis agonist), with the resultant inability to commence programmed cell death by damaging cell DNA and develops chemo-resistant phenotype. Whereas TP53/MDM2 alterations reportedly coexisted frequently in patients with adverse clinical outcomes from GCT tumor tissue [97].

9.6.7 Vemurafenib and BRAF V600E

Vemurafenib is a BRAF inhibitor which was approved in 2011 by FDA in United States for the treatment of unresectable or metastatic melanoma. A combination of vemurafenib and MEK inhibitor cobimetinib is given to the patients with BRAF mutant V600E. Compared with vemurafenib monotherapy, its combination therapy with cobimetinib showed significant promising progression-free survival chances in patients along with decreased incidence of secondary cutaneous cancers [84]. The coefficients of variation of systemic exposure of vemurafenib-treated patient is reported from 32 to 55% which is significant for determining the exposure-response relationships with regards to toxicity and response [80].

Interpatient pharmacokinetic variations of vemurafenib are considerably due to polymorphic drug-metabolizing enzymes and drug transporters hence provides sound grounds to consider pharmacogenetic protocols by genotyping the patients and dose adjustments as an effective approach to optimize the interpatient variability and drug response and combat drug toxicity. CYP3A4, enzyme of cytochrome family readily metabolizes vemurafenib. The drug efflux transporters system of ATP-binding cassette subfamily B member 1 (ABCB1), P-glycoprotein and ATP-binding cassette subfamily G member 2 (ABCG2) have reportedly polymorphic forms that are potentially responsible for acquired resistance against vemurafenib in breast cancer and BRAF (v600E) mutant cancers [69, 156]. Polymorphic forms of CYP3A4, ABCB1, and ABCG2 hinders the protein expression and clarify the significant interpatient differences of vemurafenib bioavailability and response to toxicity relationship. As CYP3A4*22 variant (rs35599367, 15389C > T) reportedly reduced the CYP3A4 mRNA expression and functional enzyme activity [150]. This was postulated due t observable reduced pazopanib clearance in heterozygous patients for CYP3A4*22 allele [16] as well as also in patients treated for breast cancer with drug docetaxel, a CYP3A4 *22 positive allelic status was showing increased events of grade 3/4 toxic effects [128]. Increased exemestane plasma concentrations are reported for patients with positive CYP3A4*22 allele status with early stage breast cancer [63]. Furthermore, peripheral neurotoxicity either acute or chronic is more often common in patients carrying the CYP3A4* 22 allele on paclitaxel exposure [34].

9.6.8 Crizotinib and ALK

Crizotinib was approved in 2011 by FDA and is a first clinically designed and synthesized as an antitumor agent with multiple targets that inhibits of receptor tyrosine kinase including hepatocyte growth factor receptor (HGFR, MET) and Recepteur d'Origine Nantais (RON) [159]. Mutations in the ALK gene can result in altered expression of oncogene fusion proteins. This agent showed marked antitumor activity in patients with advanced, ALK-positive NSCLC [18, 124].

Oral dose of crizotinib is 250 mg given twice daily. The maximum tolerated dose with a steady-state concentration reached in 15 days when given to 167 patients with cancer [60]. Average bioavailability is 43% (range: 32–66%) with minimal influence of other factors like ethnicity, food, age, sex or body weight on the single-dose crizotinib [88]. PK parameters for crizotinib treated ALK-positive NSCLC patients including anaplastic large cell lymphoma, neuroblastoma, and inflammatory myofibroblastic tumor ware similar. In Asian ethnic patients, peak plasma concentrations (C-max) and area under the plasma concentration-time curve of crizotinib was greater than patients with non-Asian ethnicity [109].

Mutant ALK gene fusion plays an active role for underlying development of NSCLC. ALK gene fusion defines a subgroup of tumors that are susceptible to targeted molecular therapy. It is extremely essential to precisely identify ALK gene fusion for the use of ALK inhibitors for NSCLC treatment. Crizotinib has shown significant reduction in the proliferation of cells carrying genetic alterations in ALK in phase I and II clinical trials [82].

Crizotinib is primary metabolized by highly polymorphic CYP3A4/5 enzymes. Drug interactions and resistance is most likely. Crizotinib like small-molecule tyrosine kinase inhibitors (TKIs) including imatinib, erlotinib, and gefitinib have low penetration of the cerebrospinal fluid and requires alternative treatment regimen. Studies on cell lines overexpressing showed that a novel agent PF-06463922 which is a multitargeted ALK and ROS1-TKI, a low-efflux substrate potentially had better CNS penetration in advanced ALK-positive NSCLC [11]. PF-06463922 also exhibited better efficacy and potency against brain metastases compared with crizotinib and alectinib in Phase I and II clinical trials. This agent can have promising results in terms of treatment effectiveness in ALK-rearranged NSCLC patients with CNS disease in the crizotinib resistance [30].

9.6.9 Cetuximab, Panitumumab and KRAS, BRAF

Cetuximab and panitumumab are commonly used monoclonal antibodies for the treatment of metastatic colorectal cancer as well as head and neck cancers. These agents bind to extracellular EGFR. However, cancer patients with mutation of KRAS oncogene do not benefit from this treatment which also act as powerful negative predictive biomarker for resistance to EGFR-inhibitory therapy [30]. KRAS gene is a member of RAS family of oncogenes that activate proteins for intracellular EGFR signaling pathways which plays an important role in cell proliferation, differentiation, and apoptosis. KRAS mutation downregulates the signaling pathways. Hence, treatment regimen with cetuximab or panitumumab will have no combined benefit in curing KRAS-mutated tumors [77].

Many retrospective researches and assessments have shown that clinical benefits from EGFR-inhibitor treatment benefits the cancer patients with unmutated KRAS oncogene. The 60–65% of the population with wild-type or neutral KRAS oncogene respond well with cetuximab or panitumumab while rest of 35–40% of the population with cancers are poor responders to this regimen [127]. Therefore, FDA has recommended a pharmacogenetic testing on the KRAS gene before advising cetuximab and panitumumab in the treatment of colon, lung as well as head and neck cancers and is indicated only for patients with KRAS mutant negative, i.e., the wild-type KRAS [81].

In addition to KRAF-mutation, BRAF mutations also show predictive biomarker characteristics for EGFR-inhibitors as serine-threonine-kinase BRAF is major effector of KRAS. KRAS mutation resistant patient were also non-responders to treatment with BRAF mutation. However, patients of colorectal cancer cells with mutant V600E allele showed response to BRAF inhibitor sorafenib [108].

9.6.10 Trastuzumab and HER2/neu

Trastuzumab humanized monoclonal antibody which are highly effective in breast cancers. These agents target the human epidermal growth factor receptor type-2, i.e., HER2. It is registered U.S. and European Union (EU) for the curing of human epidermal growth factor receptor HER2 positive metastatic breast cancer, for adjuvant treatment of localized HER2 breast cancer as well as for rare type of gastroesophageal junction adenocarcinoma. In patients with breast cancers, HER-2 gene amplification is significantly related to poor disease prognosis, overall cancer-free periods, and survival of patients [129, 130]. However, it is noted that only 25–30% of HER-2/neu + breast cancer patients are responsive to this agent [131]. In addition to its direct anti-proliferative and pro-apoptotic effects, various immune mechanisms including antibody-dependent cell-mediated cytotoxicity (ADCC) and complement-mediated cytotoxicity (CMC) along with antiangiogenic activity are contributor mechanisms [131]. There is a considerable improvement in prognosis of HER2 + breast cancers since the development of targeted therapies such as trastuzumab. Lapatinib is the other approved regimen other than trastuzumab for HER2 + breast cancer. It works intracellularly against trastuzumab-resistant patients [75].

The trastuzumab showed induced cardiotoxicity in clinical trials. The combination therapy trastuzumab with anthracycline have showed increased incidences of cardiac toxicities [130]. Several explanations are postulated to determine the trastuzumab-induced cardiac toxicity. Trastuzumab is a monoclonal antibody initiates a mechanism of antibody-dependent cell cytotoxicity and complement-dependent cytotoxicity which may result in induced cardiac cell toxicity [54]. Other debatable explanations is evidence-based experimental data which supports a major role for HER2 signaling in cardiac myopathies [64]. Various HER2 gene polymorphic variants have been reported [51]. At clinical levels, the extensively investigated germline polymorphism is codon 655A > G (rs1136201, Ile/Val) of transmembrane domain of the HER2 protein associated with a high risk for breast cancer. The presence of polymorphic Val allele may act as predictive biomarker to determine trastuzumab-induced cardiac toxicity that possibly render cardiomyocytes dependent upon HER2 signaling pathway [13, 14, 61].

9.6.11 Imatinib and BCR-ABL, C-Kit

The discovery of molecular mechanisms and chromosomal anomaly responsible for chronic myeloid leukemia was a hallmark achievement. This also promised advancement and development of more targeted therapies. The Philadelphia chromosome abnormality was due to translocation at chromosome 22 and 9. This lead to juxtaposition of protooncogene between the breakpoint cluster region (BCR) and the c-ABL oncogene [31]. This additionally triggered numerous other signal transduction pathways associated with cell proliferation, differentiation, survival and resistance to programed cell death or apoptosis [145].

Imatinib is an original tyrosine kinase inhibitor (TKI) widely used for the targeted treatment of chronic myeloid leukemia (CML) as well as a first-line treatment for multiple other BCR-ABL-, c-KIT-, and PDGFR-driven cancers.

There is interpatient variability in plasma concentrations after oral administration of imatinib is associated with drug transporters. These drug transporters may act as liable predictive biomarkers for imatinib distribution into CML cells. Therefore, PGx parameters expected to play potentially important role in personalized imatinib dosing to improve treatment outcomes. Imatinib is metabolized by CYP2C8 and CYP3A4 in vitro studies, however, genotype CYP2C8 showed significant effect on imatinib metabolism in CML patients when exposed systemically. There is no considerable clinical evidence for CYP3A4 or CYP3A5 genotypes to modify imatinib metabolism and pharmacokinetics [12]. Imatinib has showed a promising efficacy in CML patients with failed interferon-alfa therapy in phase I-II trials [43].

Imatinib is a substrate for several drug influx and efflux transporters in liver and/or CML cells. Polymorphic forms in genes of drug transporters expressed in CML cells may influence intracellular distribution of drug as well as plasma concentration to drug response relationship [12]. Imatinib being a potent selective inhibitor of BCR-ABL, also targets c-kit and the platelet-derived growth factor receptor. Mutations in c-kit are found to be quite common in Asian ethnics therefore c-kit inhibitors are priority-based investigational material in this population. The phase II, open-label, single-arm trial studies for disease progression free survival, overall response to drug rate, and survival of the patient were measured. These patients with metastatic melanoma were screened for c-kit mutation. The study advocated treatment of metastatic melanoma with imatinib was especially favorable in patients with genetic variances in c-kit gene [59].

Imatinib is a first-line of choice tyrosine kinase inhibitor (TKI) for the treating of CML, which has improved overall survival (OS) in these patients. The discovery of newer TKIs for Ph + chronic-phase (CP) CML prognosis was compared for imatinib versus second-generation dasatinib, nilotinib, bosutinib, and third-generation TKIs ponatinib for efficacy and safety in terms of overall survival period, progression-free survival period of patient, hematological and nonhematological toxicities [148]. Among all of these agents, imatinib showed advantage for patients with comorbidities. However, ponatinib is an oral TKI showed potent in vitro activity against natural BCR-ABL as well as mutant BCR-ABL, including T315I in phase II trials. This can potentially be a drug-of-choice for patients who experience failure to TKI therapy and with T315I BCR-ABL mutation [29].

9.6.12 Erlotinib and EGFR

Erlotinib is a small-molecule tyrosine kinase inhibitor (TKI) that specifically binds to the epidermal growth factor receptor (EGFR) approved as a drug-of-choice in the second-line treatment setting. However, phase III trials made it the backbone for the treatment of mutated-EGFR NSCLC patients. Exon-19 deletion and at exon-21 L858R are most common EGFR mutations that show higher response to EGFR-TKIs [103]. Erlotinib at standard daily oral dose is well tolerated in NSCLC patients. The reported adverse effects are skin rash and diarrhea generally. Others are mild, reversible, and manageable [117, 143]. Erlotinib has showed clear evidence to benefit the younger Asian nonsmoker patients with adenocarcinoma of the lung especially patients carrying mutated EGFR [137].

Rhabdomyolysis syndrome is muscle injury due to myoglobinuria, electrolyte abnormalities, and acute nephrotoxicity. This can be associated with the use of lipid-lowering agents and alcohol consumption but still it is an uncommon complication in chemotherapeutic treatment [91]. Only one case is reported till now with erlotinib treatment as single-agent for occurrence of rhabdomyolysis as an adverse event [105]. Erlotinib is mainly metabolized by CYP3A4 isoenzyme and follows P-glycoprotein (MDR1/ABCB1) and ABCG2 drug transporter system. This system potentially interacts transporter-mediated drug–drug interactions especially with CYP3A4, P-glycoprotein, and ABCG2 inhibitors. Therefore, inter-individual pharmacokinetic variability due to genetic variance at drug targets affects erlotinib’s disposition [83].

9.7 Conclusion

Treatment of cancer is challenged by drug resistance, moderate tumor selectivity, narrow therapeutic index, severe side effects, significant inter-individual variability to drug response, and frequent relapse. Genetic polymorphism predominately involving SNPs has largely explained the inter-individual differences in drug response. Therefore, the knowledge of pharmacogenetics has the potential to address the issues of variable drug efficacy and toxicity by individualizing cancer therapy based on a patient’s genetic profile to improve treatment outcomes and reduce adverse events. Genotyping for critical SNPs should be an integral part of clinical workup to identify the patients who could benefit from a specific regimen without developing extensive toxicities. Controlling the genetic factors responsible for treatment failure and side effects has also been one of the main goals in the search for new anticancer drugs tailored to individual genetic profiles for personalized medicine.