CHAPTER I - INTRODUCTION
I.5. P HARMACOGENETICS AND PHARMACOKINETICS DETERMINANTS OF TAMOXIFEN EFFICACY
Despite the obvious benefits of this tamoxifen in the different treatment settings, the clinical outcomes of tamoxifen treatment in terms of efficacy and side effects are incomplete and inconstant, and almost 30 to 40% of patients either fail to respond or become resistant to tamoxifen [46]. One of the proposed mechanisms that may account for the impaired response to tamoxifen therapy is an altered bio-activation of the parent drug into endoxifen, and this either by genetic or environmental (non-genetic) factors [46, 47].
I.5.1. CYP2D6 genetic variability
CYP2D6 is the key enzyme responsible for the generation of endoxifen [27]. The metabolizing activity of this enzyme is highly polymorphic and varies considerably within a population and between ethnic groups. This large variability is partly determined by genetic polymorphisms in the CYP2D6 gene, with over 100 allelic variants identified to date, resulting in different phenotypic patterns [48, 49].
Currently, on the basis of CYP2D6 activity, the population is usually categorized into 4 phenotypes including ultrarapid metabolizers (UMs), extensive metabolizers (EMs), intermediate metabolizers (IMs) and poor metabolizers (PMs). Actually, CY2D6 gene polymorphisms, associated with null or reduced enzyme activity, have been reported to negatively influence (in a gene-dose manner) the blood level of endoxifen in numerous prospective pharmacokinetic studies [28, 29, 50-54]. Some
Page | 7 retrospective and prospective studies have shown that CYP2D6 polymorphism was associated with worse clinical outcomes in PMs and IMs patients in terms of recurrence, disease free survival and overall survival or breast cancer development in the prevention setting [50, 52, 53, 55-63].
Several studies sought for the relevance of CYP2D6 genotyping as a biomarker of tamoxifen efficacy.
As much as 25 published reports, essentially retrospectives studies and retrospective analyses of prospective cohort trials, have addressed this issue. The report by Schroth et al. [60] is one of the largest cohort study, in postmenopausal women with early breast cancer and under adjuvant tamoxifen monotherapy, that observed a significantly higher risk of breast cancer recurrence in patient with impaired CYP2D6 activity (HR of 2.12 for PM and 1.49 for IM for time to recurrence, P <
0.006) than in EM patients. Other studies in Caucasians and Asian populations have reported similar worse clinical outcomes in PM and IM than EM patients in terms of recurrence- and disease-free survival and overall survival [52, 53, 59, 61, 62, 64-67]. Conversely, several studies [68-74] and most notably two recent retrospective analyses of large prospective trials, the ATAC (Arimidex, Tamoxifen, Alone or in Combination) [75] and the BIG 1-98 (Breast International Group 1-98) [76]
trials failed to show any significant relationship between CYP2D6 phenotypic groups and recurrence, however the validity of genotype data in these studies have been questioned.
In the metastatic treatment setting, 2 studies observed a shorter time to progression and worse overall survival in IM CYP2D6 patients heterozygous for the reduced CYP2D6 allele *10 [50] and in patients poor CYP2D6 metabolizers or under strong CYP2D6 inhibitors [77]. In the prevention setting, an Italian prevention trial showed a higher breast cancer incidence in women with the PM phenotype [63]. However, 2 other larger prevention trials failed to demonstrate an impact of CYP2D6 metabolism or potent CYP2D6 inhibitor on tamoxifen efficacy [78, 79].
The great heterogeneity and inconsistency in study results have been attributed to different confounding factors and critical errors in experimental and study designs [80-82]. The major identified factors are inherent to the quality genotype data including:
The lack of comprehensive CYP2D6 genotyping with limited allelic coverage and consequently misclassification of patients [83].
Heterogeneity in alleles grouping and phenotypes definition or scoring.
The use of low-quality genetic DNA extracted from somatic tissues and especially breast tumor tissues frequently affected ( 35% of ER + tumors) by a loss of heterozygosity (LOH) at the chromosome 22q13 harboring the CYP2D6 gene. This deletion in the CYP2D6 genes leads to strong allelic imbalance, deviation for Hardy-Weinberg Equilibrium (a measure of expected allele and genotype frequencies in the population and an indicator of genotyping quality and
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uniformity of the population under investigation) and erroneous CYP2D6 genotyping and misinterpretation of pharmacogenetics study results.
Other confounders identified from study comparisons may explain theses discrepant data. These confounders include lack of information or adjustment for CYP2D6 inhibitors co-administration, lack of information on tamoxifen adherence and the use of combination therapy. Actually, most of the studies that failed to show any association between CYP2D6 genotype polymorphism and tamoxifen efficacy, included patients with concomitant use of tamoxifen and chemotherapy. Besides, heterogeneity in size of the study population, disease stage, end-point definition (i.e recurrence-free survival, event-free survival, disease-free survival, overall survival) and length of follow-up (particularly in case of a switch to an aromatase inhibitor) may contribute to explain controversial findings of these studies [80, 81, 84].
I.5.2.Influence of other genetic variations of metabolizing enzymes and transporters
As previously indicated, other CYPs (CYP2C9, 2C19, 3A4, 3A5), UGTs and SUTs are also involved in the metabolism of tamoxifen. For CYP2C19, Schroth et al. [57] found that carriers of the CYP2C19*17 variant (homozygous or heterozygous) had a lower risk for relapse than patient carriers of *1 (wild type), *2 or *3 (defectives) alleles. Other group either failed to confirm such results or found opposite results suggesting a longer breast cancer survival in carriers of the defective CYP2C19*2 variant [63, 71, 85, 86]. Conversely, for CYP3A5 (*3 null variant), 2B6, 2C9 (*2, *3 null variant), SULT1A1 (gene copy number and *2 reduced activity variant), UGT1A2B7 (*2 reduced activity allele) and UGT2B15 (*2, increased activity alleles) no association between genotype and clinical outcome have been demonstrated [55, 60, 68-70, 75, 87, 88]. In addition, tamoxifen and its active metabolites 4-hydroxy-tamoxifen and endoxifen are substrate of P-glycoprotein (P-gp), however, this enzyme do not seem to play a significant role in the drug pharmacokinetics and pharmacodynamics [66, 89, 90].
The study by Kiyotani et al. [53] have shown no effect of ABCB1 gene polymorphism (coding for the P-gp) on tamoxifen outcome, however, they observed that patient with reduced activity of the transporter MRP2 (multidrug resistance associated-protein 2 coded by the ABCC2 gene) have shorter recurrence-free survival.
I.5.3. Effect of non-pharmacogenetics factors I.5.3.1. CYP2D6 inhibitors
Amongst patient under tamoxifen treatment, approximately 14 to 30% receive an antidepressant, such as selective serotonin reuptake inhibitors (SSRIs) or selective serotonin and norepinephrine reuptake inhibitors (SNRIs), to treat depression or to alleviate tamoxifen-induced vasomotor
Page | 9 symptoms (i.e. hot flushes) [91-93]. Some of these SSRIs, such as paroxetine and fluoxetine, are strong CYP2D6 inhibitors resulting in a reduced to null CYP2D6 activity and impairment in tamoxifen metabolism with a significant reduction (up to 70%) in the main active tamoxifen metabolite, endoxifen concentration to levels comparable to those in CYP2D6 PM patients [28, 29, 94].
Such phenocopying to apparent CYP2D6 PM status, is expected to hamper treatment effectiveness.
However, the epidemiologic studies that have attempted to correlate concomitant CYP2D6 inhibitor use to tamoxifen outcome have also reported mixed conclusions. One large cohort study by Kelly et al. [95] reported an increase in breast cancer mortality risk in patient using paroxetine and this increase in mortality was closely related to the duration of overlap use with tamoxifen. They estimated that for a 41% overlap-time use with tamoxifen, 1 additional breast cancer death over 20 women occur within 5 years after tamoxifen cessation. This study failed to demonstrate the same association for fluoxetine, sertraline, fluvoxamine, venlafaxine and citalopram. Other studies reported an increased risk of recurrence and lower overall survival in patients under strong and moderate CYP2D6 inhibitors [55, 77, 96]. By contrast, in a population-based case-control study in Denmark, Lash et al. find no evidence of a relationship between concurrent use of (es)citalopram and tamoxifen and risk of breast cancer recurrence [97]. The same observation was also reported for more potent CYP2D6 inhibitors (paroxetine, fluoxetine, sertraline) [74, 98-101].
As for pharmacogenetics studies, results from different retrospective heterogeneous cohort and case-control studies were conflicting and failed to confirm the observed association even between strong CYP2D6 inhibitors and tamoxifen outcome. Although, these findings were inconclusive, present recommendations are to avoid whenever possible the use of potent CYP2D6 inhibitors in tamoxifen treated breast cancer patients and to consider rather the use of medications with little CYP2D6 inhibitory potential such as venlafaxine and citalopram [12, 81, 102, 103].
I.5.3.2. Compliance
Non-adherence to tamoxifen may have played a non negligible role as confounding factor in pharmacogenetics studies as well as in trials investigating the effect of CYP2D6 inhibitors on tamoxifen effectiveness. Early treatment discontinuation and non-adherence to treatment are a major concern for adjuvant endocrine therapies. Among patients under tamoxifen, 15 to 20%
discontinued their treatment by the 1st year and 31 to 61% at the end of 5 years. Among patients who continued their treatment, adherence to tamoxifen declined over-time and ranged from 41 to 88% [104-107]. Most notably, poor adherence to tamoxifen has been linked to a lower breast cancer recurrence or event-free time [65, 101]. Poor adherence and early discontinuation have also been linked to an increase in all-cause mortality in breast cancer patients [108, 109]. It has been reported
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that tamoxifen side effects such as hot flashes are major determinant for adherence and persistence to the adjuvant hormonal therapy [110, 111]. Rae et al. [111] also observed that patients with higher CYP2D6 activity presented increased rate of treatment discontinuation and postulated that this could be related to the reported higher burden of side effects such as hot flashes in CYP2D6 EM and IM than PM patients [55].