Antidepressant Therapeutic Drug Monitoring by Minimally Invasive Techniques in Eating Disorders Patients: Preliminary Results from a Pilot Study with Focus on Vortioxetine

Patients with Eating Disorders (ED) including, among others,
Anorexia Nervosa (AN), Bulimia Nervosa (BN) and Binge-Eating...

It was suggested that SSRIs have decreased efficacy in anorexic patients with low BMI due to starvation-related biochemical changes in the brain [13] and because of inadequate concentration of nutrients, which are necessary for serotonin metabolism [14].
Nevertheless, nutritional supplements containing tryptophan and essential fatty acids do not seem to increase SSRIs efficacy in underweight patients with AN [15]: the potential reason remains uncertain, but the latest genetic findings suggest that ED may not only be seen as psychiatric, but also as metabolic and immune disorders [6,16]. In a study on patients treated by Risperidone, Paulzen M. and colleagues suggested that extreme low and high BMIs modify psychotropic drugs' metabolism, in particular pharmacokinetic parameters [17]: obese patients showed a higher plasma concentrations of the active metabolite of Risperidone when compared with cachectic patients; authors speculate that this pharmacological behaviour is due to cytochrome P450 activity alterations or differences in P-glycoprotein function. Other studies showed negative correlation between BMI and plasma concentration of psychotropic drugs and/or their metabolites [18,19] and these results are explained by the authors as linked to different distribution volumes for lipophilic substances (larger distribution volume in obese subjects leading to inadequate plasma levels and smaller therapeutic efficacy) [19]. Unterecker et al. instead point out the absence of any relationship between body weight and plasma concentrations of antidepressants, highlighting the need for further exploration studies [20]. Vortioxetine is an antidepressant with multimodal activity currently approved for the treatment of major depressive disorder that is metabolized by cytochrome P450 enzymes and subsequently by uridine diphosphate glucuronosyltransferase. Several evidences from current literature do not show clinically relevant differences in vortioxetine exposure by sex, age, race, body size, and renal or hepatic function. Dose adjustment is only recommended for cytochrome P450 2D6 poor metabolizers and when it is associated with bupropion, a strong cytochrome P450 2D6 inhibitor, and rifampin, a broad cytochrome P450 inducer.
The simultaneous administration of other drugs does not affect vortioxetine exposure or safety profile. Moreover, pharmacodynamic findings demonstrate that vortioxetine achieves high levels of serotonin transporter occupancy in relevant brain areas and modified abnormal resting state networks in the brain over the therapeutic dose range. Overall, vortioxetine can be administered without major dose adjustments [21]. Considering the altered physical and metabolic condition of patients with extremely low and high BMIs, we aimed to investigate the usefulness and feasibility of minimally-invasive and miniaturised biosampling techniques used for Therapeutic Drug Monitoring (TDM) of psychiatric Central Nervous System (CNS) drugs [22,23] in outpatients affected by ED. TDM of SSRIs has already several approved indications such as lack of clinical response despite adequate doses, adverse effects using recommended doses, and drugs' prescription in patients with pharmacokinetically relevant comorbidities (eg. extremely high or low BMI) [24,25], to obtain better therapy optimization and personalization [25,26]. Moreover, TDM can also lead to reduced healthcare expenses, due to the possibility of better efficacy, increased patient compliance and enhanced safety, leading to a reduction in hospitalizations due to unwanted effects or therapy ineffectiveness [27][28][29][30].
In this pilot study we have described the use of Volumetric Absorptive Microsampling (VAMS) in the management of pharmacological treatment of ED patients in outpatient setting in order to analyse any pharmacokinetic differences of the main antidepressants in use in patients with very varied ED and BMI. A secondary goal is to test for the first time the applicability of the VAMS technique for the analysis of antidepressants in whole blood and Oral Fluid (OF) for the purposes of TDM. The main aim could help us to better describe and understand the relationship between the BMI and biological fluid concentrations of antidepressant drugs together with their metabolites in these cohort of patient. In order to do that, we have carried out an analytical workflow based on VAMS of both whole blood and OF, followed by microextraction by packed sorbent (MEPS) [23] and Liquid Chromatographic (HPLC) analysis with spectrophotometric (UV) and spectrofluorimetric (FL) detection.

Patient's Assessment
The evaluation protocol at the outpatient unit for ED consisted of three different meetings with a psychiatrist. In the first meeting,

Hplc-Uv-Fl Instrumentation And Conditions
HPLC-UV-FL analysis was performed on a Waters Corporation

Blood-and OF-VAMS Sampling and Pretreatment
For patient sampling, IS spiking was carried out on the VAMS tip by automatic pipetting before the sampling; the tip was then left to dry for 2 h at RT before use. Mitra® VAMS microsamplers

Statistical Analysis
All the statistical analysis were performed using SPSS IBM V.24.0 (statistical Package for Social Science). Other analysis (i.e. Linear Regression) will be performed in further studies when the sample size will be compatible with statistical power.

Results
As reported in finally, one patient is treated with Escitalopram and one patient with Fluvoxamine (to treat the concomitant obsessive-compulsive disorder in comorbidity with BED).

Discussion
The main goal of this study is to highlight a possible relationship between the whole blood and salivary concentration of antidepressants and their active metabolites using minimally invasive TDM methods such as VAMS strategy in a specific population affected by ED. This subset of psychiatric patients is extremely interesting in terms of correlation of peripheral concentration of drugs with oral dosage, because patients with ED have an important variability in terms of BMI. It is well known that BMI could affect clinical response to antidepressant drugs [31,32].
TDM can give us correct information on the concentration of active metabolites of antidepressants in peripheral matrices, net of the effect of liver metabolism and BMI on the pharmacokinetics of psychotropic drugs.
VAMS technology is currently a topic of enormous interest for the bioanalytical community and is the subject of numerous studies that have grown exponentially in the last two years [23,[33][34][35][36].
VAMS devices were initially marketed for use on whole blood only; this method based on VAMS devices is applied also to the OF for the quantitative analysis of synthetic cathinones [36]. blood and salivary concentration of monohydroxycarbamazepine [37], carbamazepine, phenytoin and phenobarbital [6,38]. For amitriptyline, nortriptyline and valproic acid, however, the correlation shown was not significant [6,38,39]. For a long time the concentration of drugs in OF has been considered as a reflection of the free component of the drug in the blood, which for many psychotropic drugs is only 10% or less of the total concentration; however, the distribution of drugs between blood and OF largely depends on the pH, which increases when the secretion is stimulated. OF is considered as an interesting and advantageous matrix for TDM studies, in particular due to the non-invasiveness, but data available so far in literature show contrasting results and further studies are necessary to validate the use of OF as a matrix for the TDM of antidepressant drugs [6,40]. On the other hand, according to the current literature [32], high BMI may also affect treatment response: Khan et al., [32] have found obesity to be associated with a less vigorous treatment response, with an effect more pronounced in males. Moreover, higher BMI was associated with a slower clinical response in the Munich Antidepressant Response Study (MARS) [41]. Finally, in the Genome Based Therapeutic Drugs for Depression (GENDEP) study, obesity was associated with a poorer response to nortriptyline in men and women, and a poorer response to escitalopram in women [19]. is not effective, the therapist must empirically evaluate whether to increase the dosage or to change the type of psychiatric drug. The clinician's resistance in prescribing the highest dosages allowed for a given antidepressant to patients with very reduced BMI recalls the general principles of pharmacology according to which low distribution volumes (AN patients) will correspond to higher blood concentrations for a given dose of drug in ratio to patients with BMI in the normal range, population in which blood concentrations are studied with reference to a given oral dose.
In outpatient setting, a fortuitous trial-and-error process is used to try to reach the optimal dose for the specific patient [26,49]. In consideration of point 2., however, it is necessary to consider the pharmacokinetic variables and the absorption, distribution and elimination characteristics of the antidepressant drugs considered in this study, described in the introduction, except for vortioxetine, chosen in this study, for the peculiar pharmacokinetic profile, as well as for the uniqueness of the pharmacodynamic profile [21].
Overall, the pharmacokinetic factors to be taken into account when describing the concentrations of antidepressants in blood and OF of such patients with particular physical characteristics are numerous and intricate. Ideally, the chemical-physical properties of the drug, its absorption, first pass metabolism, bioavailability, protein binding, distribution in abnormal body volumes (in association with the study of body composition), should be taken into account.
The fat content and visceral obesity could explain some differences in the response to antidepressant drugs [19], the passage at the the whole blood concentration of the drug will be lower than in a patient with a very poor volume distribution (patient with AN).
Considering that routine blood tests are systematically prescribed during the taking in charge at the Unit for the Study and Assistance of the DCA and that patients are followed in an outpatient setting without the needing of hospitalization, it is possible to affirm with relative safety that the patients enrolled in the present study do not show significant alterations in renal and/or hepatic function, although the latter have not been thoroughly evaluated with specialized tests [53][54][55].
This pilot study reports numerous limitations for logistical reasons, having been conducted in a "real world" setting with all

Author Disclosure
This study did not have funding sources