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Impact of alprazolam in allostatic load, neurocognition and quality of life of patients with neuroticism, anxiety
disorders and chronic stress (GEMA): Observational study protocol for a phase IV clinical trial.
Journal: BMJ Open
Manuscript ID: bmjopen-2014-007231
Article Type: Protocol
Date Submitted by the Author: 17-Nov-2014
Complete List of Authors: Soria, Carlos; Institute of Biosciences Henri Laborit, Córdoba Remedi, Carolina; Institute of Biosciences Henri Laborit, Córdoba Nuñez, Daniel; GADOR SA, Scientific Direction D'Alessio, Luciana; Cell Biology and Neuroscience Institute, CONICET Roldán, Emilio; GADOR SA, Scientific Direction
<b>Primary Subject Heading</b>:
Mental health
Secondary Subject Heading: Mental health
Keywords: CLINICAL PHARMACOLOGY, Adult psychiatry < PSYCHIATRY, Anxiety disorders < PSYCHIATRY
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Impact of alprazolam in allostatic load, neurocognition and quality of life of patients with
neuroticism, anxiety disorders and chronic stress (GEMA): Observational study protocol for a
phase IV clinical trial.
Authors
Carlos A. Soria, Carolina Remedi, Daniel A. Núñez, Luciana D’Alessio, Emilio J. A. Roldán.
Institution
Institute of Biosciences Henri Laborit, Córdoba, Argentina.
Scientific Direction Gador SA, Buenos Aires, Argentina.
Corresponding Author:
Luciana D’Alessio, Phd, MD
email: luladalessio@gmail.com
Darwin 429 (C1414CUI) CABA
Buenos Aires, Argentina
Phone 00541169518500
Background
Benzodiazepines are used as anxiolytics in interventions planned for a few weeks. However a
number of symptomatic patients with chronic stress and allostatic loads frequently demand to
continue their use because they cannot solve the underlying morbidity with two consequences:
poor results and risks due to prolonged consumption. Nevertheless, chronic stress and
exposure to allostatic load may induce riskier cognitive alterations. Hence, the use of
alprazolam in low doses might favor improvements in allostatic load and cognitive variables
beyond anxiolysis.
Methods/analysis: This study, GEMA (Gador study of Estres Modulation by Alprazolam), is an
ongoing prospective phase IV study, which aims to determine the impact of medication on
neurobiochemical variables related to chronic stress, metabolic syndrome, neurocognition and
quality of life in patients with anxiety, allostatic load and neuroticism. The study is an on demand
assignment of alprazolam doses in a wide range, in patients with anxiety disorders (six or more
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points in the Hamilton-A scale), neuroticism (a score of more than 18 points in the NEO-FFI
scale), neurocognitive performance (more than 25 points in the Folstein’s Mini-Mental State
Examination) and an allostatic load expressed by at least 3 positive clinical or biochemical items
(Crimmins and Seeman criteria).
Study participants will be assessed prior to treatment (week-1), on the day treatment started
baseline (day 0), and during treatment on weeks 1, 2, 4, 8 and 12. Clinical variables of anxiety,
neuroticism, allostatic load, neurobiochemical studies, neurocognition and quality of life will be
determine.
Ethics and dissemination: This study protocol was approved by an Independent Ethics
Committee of FEFyM (Fundación de Estudios Farmacológicos y Medicamentos/Foundation for
Pharmacological Studies and Drugs, Buenos Aires) and by regulatory authorities of Argentina
(ANMAT, Dossier # 61409-8 of April 20th, 2009), following the law of Habeas Data and
psychotherapeutic drug control. Trial Registration: GEMA - 20811
Limitations:
One of the important biases of the project is the lack of a parallel control group, which is not
regulatory allowed in our area, in a phase IV clinical trial with highly symptomatic patients with
inherent risks. As the project moves forward, however, the study itself can be justified in a more
advanced stage, for example, in a face-to-face comparison of different schemes.
Another difficulty is the existence of knowledge gaps among the clinical features, cognitive
observations and variables measured in laboratories. So, we assume that as we move forward
in the associations among them, other hypotheses on mechanism of action may arise that can
be formulated and tested by other investigators.
Key Words: Alprazolam, benzodiazepines, anxiety disorders, allostatic load, chronic stress,
cognition.
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1- Background
Stress related mental condition such as anxiety and neuroticism 123
are frequently
associated with higher allostatic loads and chronic stress, and overall all of these factors
increase the vulnerability to diseases and reduce resilience45678
.
Anxiety disorders have been linked to disturbances of hypotalamo- pituitary-adrenal
(HPA) axis with controversial results9 10
11
. Hormones such as cortisol and CRH (corticotrophin
releasing hormone) have been associated to states of fear and anxiety in experimental models
1213. Furthermore, increased basal levels of cortisol were found in patients with panic disorders
and in patients with GAD (general anxiety disorders) 14 15
. Nevertheless others authors did not
find any association in cortisol levels and anxiety scale measures 11
or as well found a reduction
in cortisol awakening response among older patients with anxiety disorders 16
. On the contrary
studies performed among posttraumatic stress disorder patients showed lower levels of cortisol
17. High levels of cortisol were also found in patiens with neuroticism (a predisposition to
respond with intense emotional reaction to psychosocial stressors) 12
.
In stress conditions both HPA axis and the noradrenergic (NA) system seem to function
in a synchronous fashion and these two systems are crucial for stress response in mammals 67
.
There is substancial evidence that anxiety in humans is associated with high levels of 3-
methoxy-4-hydroxyphenilglycol (MHPG), a principal NA metabolite, and salivary determination
of MHPG may be a useful marker for detecting changes in catecholamine metabolism 18
19
20
.
1.1 Does allostatic load and chronic stress affect cognition process?
The allostatic load model has been particularly informative in specifying the additive
effects of multiple biological processes that contribute to wear and tear on tissues and
accelerate pathophysiology related to stress6 57
21
. Stressful experiences have functionally-
relevant effects on dendritic arbor and spine/synapse number in many brain regions, including
the hippocampus, amygdala and the prefrontal cortex (PFC) with effects not only on cognitive
function but also on emotional regulation and other self-regulatory behaviors and upon
neuroendocrine and autonomic function 22
. By compromising hippocampal function, allostatic
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load and elevated levels of glucocorticoids impair normal hippocampal function as well memory
processes (declarative memory) 7 23
24
. Furthermore, stress-related effects upon the PFC has an
importance in working memory process (the ability to keep events in mind)25
and self regulatory
behaviors. The dorso lateral PFC is responsible for planning approaches and sequences of
behavior that are required for goal-directed behavior. This process is critical of executive
function 7 25
.
In experimental models, chronic restraint stress caused spine loss and debranching of dendrites
on PFC neurons and impaired working memory7 22
26
. In humans, adverse childhood
experiences were associated with smaller volume of the PFC greater activation of the HPA axis,
and elevation in inflammation levels 27
. Furthermore, it has been shown that stress and allostatic
load increases interference from irrelevant information and thus impairs selective attention 28
,
and high levels of cortisol in response to stressors showed lower performance in working
memory tasks in normal adults 29
. Also, it has been found that anxiety disorders affect cognitive
style of functioning, reversed by alprazolam treatment 30
.
1.2 How treatment anxiety with benzodiazepines impacts on allostatic load?
Benzodiazepines (BZD) are GABAergic substances with anxiolytic and sedative activity
available since the sixties. The therapeutic use to depress acute and subacute symptoms of
anxiety disorders has gained great popular acceptance up to the present 31
. Stress related
mental conditions such as anxiety disorders and neuroticism are treated with psychotherapy
and psychopharmacologic drugs, which are primarily prescribed to reduce pathological
symptoms but the impact on the underlying disorder is generally less understood. Furthermore,
many patients persist with symptoms and consequently require the use of BZD for longer
periods of time. Nevertheless, this indication is an off-label use is therefore regarded with
caution because of the possibility of favoring dependence on medication and other undesired
events such as cognitive impairment 31
32
33
.
In clinical practice prolonged prescription is described when there is recurrence of a
clinical situation and due to a positive perception of both patients and prescribers, there is a
favorable risk/benefit ratio because of the presence of a chronic condition which has not been
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completely resolved. This demand for medication could be significantly reduced if basic
phenomena that trigger anxiety, such as persistence of a harmful stress state, allostatic charge
accumulation and/or decrease of individual resilience level, could be efficiently modulated or
abolished. When it dosen’t happen and the stressors persist, cognitive variables could be
affected, making it difficult for the patient to take decisions that are favorable for QOL (quality of
life)33
34
35 36
Following the NICE (National Institute for Health and Clinical Excellence, UK)
guidelines, treatment of a person with GAD (general anxiety disorder) and chronic stress
consists of psychotherapy (cognitive behavioral therapy), but when symptoms induce a
significant functional impairment, it should be resorted with drugs, generally antidepressant and
benzodiazepines 36
. The objective of treatment is not only to suppress symptoms of anxiety, but
also to prevent a chronic outcome, and to guide patients to positive behaviors “decisions
making” for a better QOL. This involves strengthening native resilience mechanisms, balancing
the impact of individual allostatic loads and promoting an efficient use of cognitive resources
through supportive therapies. The result of this effort, however, is not always satisfactory or it
demands a long time, especially when the neurobiochemical function of the affected patient is
unbalanced by a chronic and unresolved morbid condition5 .
As BZD have anxioliytic and sedating properties, it is expected that they influence the
stress system. As well most studies showed a decrease in cortisol levels 37
38
39
40
41
, some
studies reported mixed results 41
42
43
, and these inconsistencies may be explained by
differences in patients groups, dosages, and BZD used among studies 41
. Some studies yield a
reduction of variations in noradrenaline (NA) metabolite levels (MHPG) "noradrenergic
volatility"44
, and in circulating and salivary cortisol levels using alprazolam in anxious patients 18
39 37
and in normal subjects 45
or as well an increase levels of dehydroepiandrostrerone (a anti-
cortisol mediator)46
. Nevertheless many controversial findings were found depending on age
and gender: younger anxious patients showed higher levels of salivary cortisol awakening
response 11
16
, but other authors found high salivary cortisol levels associated with severity of
late-life generalized anxiety disorders in older adults 15
. Furthermore women seem to yield a
different profile in the cortisol response to stress, in relation to men on response to alprazolam
treatment 40
. Previous data from our clinical practice showed that the administration of
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alprazolam results in a fast and significative decrease of MHPG concentrations and a decrease
of noradrenergic volatility accompanied by a slower but equally sustained decrease in salivary
cortisol levels 47
(Figure 1).
The direct action of alprazolam on GABAergic neurons, and secondarily, on
catecholaminergic and serotoninergic neurotransmitters may induce the indirect anti-stress
target reducing cortisol plasma levels. The reduction in "noradrenergic volatility" defined by the
sudden variations in the levels of NA (noradrenaline) and its metabolite MHPG observed in
patients with GAD, stress-related diseases and depressive states, may also be involved 44
48
49
.
In stress conditions both the NA system and the HPA axis seem to function in a synchronous
fashion. Increased activity in one system goes along with parallel variations in the other and
vice versa and the coordinated interaction of these two systems is a unique element for stress
response in mammals 50
51
52
. The noradrenergic function is hyperactive as it has been
documented in anxiety states, indeed the question is how the HPA axis responds, and if the
physiological synchronicity of both systems remains intact or not, during treatment.
1.3 Do benzodiazepines affect cognitive function positively in stressed patients?
Identification of drugs with adverse cognitive effects, requires dedicated monitoring of cognitive
function, which is often absent from routine hospital and ambulatory clinical care 53
. The short
term effects of BZD on cognition are well known and they are mediated through an agonist
action on receptors of γ aminobutyric acid (GABA A)54
53
. A systematic review of the association
between benzodiazepines and cognition found that cognitive impairment was presumed to be
transient and reversible in most studies 34
53
. Nevertheless the long term adverse effects of
BZD on cognition are still debated 32
53
53
and available data shows conflicting results. Some
authors found an improvement in memory and an increased risk of dementia in prospective
population based studies, performed in elderly patients with long term use of BZD32
55
56
57
58
.
Furthermore longer acting BZD were associated with greater risk of developing dementia
compared with shorter acting benzodiazepines 32
. Nevertheless, other concomitant factors could
be also responsible of dementia in such studies. Indeed, a higher degree of neuroticism in midlife
was associated with increased risk of dementia and long-standing distress, in a population-based study. 59
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Other studies showed not conclusive results 60
61
or even reported a potential protective effect
on behavioral disturbances associated to dementia: the role of GABA in dementia has yet to be
fully elucidated and behavioural symptoms associated with dementia may be a direct or indirect
result of GABAergic dysfunction 62
63
. Furthermore, in young adults, BZD may affects cognition
differentially depending on gender: cognitive alteration of long-term memory have been reported
in women, whereas there was no significant affectation in men 61
.
In this study we selected alprazolam (a triazolo-benzodiazepine) for different reasons: is
a high-potency benzodiazepines with an indirect activity through 5-HT1A serotonergic receptors,
which would produce a regulation of the hyperactive noradrenergic pathways and, through
them, a reduction of noradrenergic volatility. The reduction of noradrenergic volatility by high-
potency benzodiazepines would have a preventive effect via the noradrenergic system, beyond
the central GABAergic anxiolytic effects. The level of concentration and distribution of
alprazolam within the CNS is effective when it is administered in relatively low doses, with a
differential impact on GABAA receptors, upregulated in individuals under chronic stress, and
with less impact on cognitive functions because of the shorter acting 32
49
64
58
65
(Table 1)
1.4 General Hypothesis
In the present study we aim to investigate the intermediate term (12 weeks) biochemical and
neurocognitive effects of treatment with low doses of alprazolam in young adults with anxiety
and chronic stress. The question attempted to be solved is whether anxiolytic pharmacology
intervention with alprazolam, provided with secure schemes, modify allostatic load,
neurocognition performance and quality of life, beyond the known anxiolytic effect 49
66
67
. The
global hypothesis under investigation is if neurobiochemical indicators of allostatic load
(noradrenergic metabolites and cortisol levels), are modulated during treatment with alprazolam,
and if there are neurocognitive consequences when treatment maintains neurobiochemical
parameters within the normal range. A pharmacological modulation of such mediators may
contribute to reduce the impact of allostatic load, promoting resilience and better biochemical
and neurocognitive performance translated in a better quality of life.
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This study is based on the hypothesis that high potency benzodiazepines such as
alprazolam may serve as more than single anxiety suppressive agents reducing allostatic load
and facilitating adaptative responses. GEMA project proposes an intervention designed to
determine if minimum doses of alprazolam adjust neurobiochemical mediators, neurocognition
and affect quality of life, altered by stress (Figure 2).
1.5 Objectives (End-points): The project consists of three complementary stages (Figure 3).
1.5.1 To determine whether neurobiological and biochemical markers of allostatic load, tend to
normalize with 12 weeks of therapeutic intervention.
1.5.2 To determine if neurocognitive performance is modified after treatment and if there is a
positive correlation with biochemical variables of allostatic load and with therapeutic levels.
1.5.3 To assess QOL before and after treatment
Secondarily, the purpose of this study is to report conditions of failure due to lack of response or
emergence of undesired events, including adverse events, and possible situations of
habituation to or dependence on alprazolam.
2- Methods and analysis:
2.1 Protocol Design:
The GEMA project (Gador study of Estres Modulation by Alplax), is a prospective phase IV
clinical trial. The study began in 2010 and is still in advance. This is a prospective observational
study, controlled with on demand assignment of alprazolam doses in variable doses. It is a
phase IV study reported to regulatory authorities in compliance with the rules in force and
consequently enclosed for the use of experimental controls or interventions.
The presence of variable doses will allow us to establish a continuous range of dose-response.
2.2 Inclusion Criteria
Patients over than 21 years, who have at the time of admission anxiety symptoms (six or more
points in the Hamilton-A scale),68
neuroticism (18 or more points in the NEO-FFI scale),
69cognitive performance (25 or more points in the Folstein’s Mini-Mental State Examination)
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70and allostatic load expressed by at least 3 positive clinical or biochemical items (Crimmins and
Seeman criteria). 71
2.3 Exclusion Criteria
Other psychiatric disorders listed under Axis I of the DSM-IV (Diagnostic and Statistical Manual
of Mental Disorders, Fourth Edition)72
. Patients requiring other psychotropics added-on to
alprazolam, sympathicomimetics, corticosteroids or another medication that might kinetically or
dynamically interfere with alprazolam. Patients who had received fluoxetine in the previous 4
weeks or MAO (monoamine oxidase) inhibitors in the previous 2 weeks, or showing a history of
hypersensitivity to alprazolam. Women with confirmed or suspected pregnancy, or subjects
affected with concomitant conditions, severe or unstable, in the cardiovascular system,
gastrointestinal, hepatic, renal, neurological and other systems.
Patients will be withdrawn from this study if dose is prematurely modified, or if patient decides
not to continue with the administered drug, has a related adverse event that modifies his/her
daily performance, or if the investigator considers that the scheme is not safe or beneficial for
the patient according to his/her current clinical status.
2.4
Sample Size: Changes in variables higher than 25% (pre-treatment versus post-treatment or
significant correlations) with a 1-ß power of 0-80 are estimated to be detected. For the first
phase, an n = 29-55 cases, for the second phase an n = 112 and for the third an extension of
the sample to 182 assessable patients, according to a PC program (Statistica, Tulsa) will be
considered enough.
2. 5
Therapeutic Scheme and Dose Assignment
Patients will be extensively assessed by a team of medical doctors, clinicians and psychiatrists,
who decide the most appropriate approach for each case. The principal investigator will
determine the initial dose of alprazolam, usually from 0.75 mg/day to 3.0 mg/day, according to
the presence of anxiety symptoms and according to patient’s tolerance. The ongoing scheme of
alprazolam will be maintained for 3 months. Thereafter the investigator can decide to continue
or discontinue treatment gradually, by reducing 0.5 mg every 3 days.
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2.6 Patient Assessment.
Study participants will be evaluated prior to treatment (week-1), on the day treatment started
(day 0), and during treatment on weeks 1, 2, 4, 8 and 12. Clinical variables of anxiety,
neuroticism and allostatic load, neurocognitive and neurobiochemical studies will be
determined.
Anxiety assessment: Two scales will be rated: 1 - The Hamilton Anxiety Rating Scale (HARS) of
14 items. 68
The 0-5 score indicates absence of anxiety, 6-14 mild anxiety and more than 15
moderate to severe anxiety. 2 - The State-Trait Anxiety Inventory (STAI) of 40 items73
, which
assesses the current state and the anxiety trait as a stable tendency. The scale scores a
maximum of 60 points and percentiles are described for gender and age.
Neuroticism assessment: The NEO five factor inventory (NEO FFI) 69
is an abbreviated
measure of personality factors related to neuroticism, extraversion, openness, kindness and
responsibility.
Allostatic load determination: Clinical and biochemical evaluations will be determined: 1-
Systolic blood pressure will be measured in the supine and standing position, with a previous 2-
3 minutes rest A systolic pressure > 140mmHg and/or a diastolic pressure < 90 mmHg will be
considered a positive criteria.74
2- Heart frequency: a frequency exceeding 90 bpm (beats per
minute) in supine or standing position with a previous 2-3 minutes rest is considered a positive
pulse. 3- BMI (body mass index) and waist-hip ratio are measured while patient is standing and
breathing normally, horizontal over the edge of iliac crests, using a tight tape without
compressing the skin. 4- Laboratory biochemical tests in serum samples include total
cholesterol, and HDL and LDL fractions, glycosylated hemoglobin, fibrinogen, albumin,
andreactive protein. Furthermore, plasma noradrenaline and cortisol and salivary MHPG (3-
methoxy 4 hydroxyphenilglycol) and cortisol will be determined.
General condition and safety: 1-Clinical Global Impression Scale (National Institute of Mental
Health 1976) (CGI), which includes two subscales, 1a - clinical profile severity (CGI-S) and 1b-
clinical profile improvement (CGI-MJ) with 8 points each one. 2-The Mini-Mental State (MEC)
quantifies cognitive impairment in time-space orientation, immediate memory, concentration and
calculation, long-term memory, language and praxis.70
3- UKU scale (Udvalg fur klinische
undersogelser)75
to determine side effects; with 56 items: the first 9 are for psychic events, 8 for
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neurologic, 11 for autonomic and 25 for others. The last two items evaluate side events
interference with the patient's daily performance and their impact on medication intake (none,
dose reduction, withdrawal or change). In each one, the investigator assesses whether the
event was related to drug treatment or not. 4- Additional safety assessments include respiratory
rate and laboratory tests: complete blood count, creatinine, BUN (blood urea nitrogen), total
bilirrubin, alkaline phosphatase, liver enzymes. Human chorionic gonadotropin (hCG-B) will be
determined when appropriate. All patients will be informed about any eventual depressant effect
of alprazolam and risks in activities that require alert and the consumption of alcohol will be not
recommended. They will be also monitored to detect potential risks of suicide or overdose. All
adverse events are monitored, classified and reported according to GCP (good clinical practice)
standards of the local regulatory authority (ANMAT and Ministry of Health of the Province of
Córdoba, Argentina).
Cognitive assessment: To quantify the neurocognitive variables; attention, working memory,
executive functions, impulsiveness, verbal and nonverbal fluency the following tests will be
determined. 1- QPSS continuous performance test (CPT)76
; 2- Digit symbol test (DST) 77
; 3-
Digit span test (DST) 78
; 4- Verbal fluency test (VFT)79
; 5-Five point test (FPT)80
; 6-revised
Taylor complex test (RTCT)81
and 7- Stroop test (ST)82
. All of them will be applied after 6 hours
since the last administration of alprazolam (nadir plasma levels). Cognitive tests and laboratory
tests are blinded to the analyst, the remaining tests are open.
Quality of life (QOL) assessment: The health and quality of life scale, health questionnaire SF-
36 validated Spanish version will be determined. 83
Statistical assessment:
The mean, minimum and maximum values of quantitative variables will be determined. Pre- and
post-treatment values will be compared using t-test and/or non parametric test. Statistical
significance will be fixed at p<0.05 (2-sided; β-1 power≥0.80). Correlation analysis will be
performed by calculating the Pearson Test correlation coefficient for ungrouped data; with r>
0.80 and p<0.05 (2-sided) strong correlation and 0.80<r>0.50 and p<0.05 (2-sided), moderate
correlation.
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Treatment safety: Regarding patient safety, the use of alprazolam, a well known, low action,
nervous system depressant drug, at minimum doses, even when no symptoms palliation is
required, guarantees good tolerability, and no drawbacks are expected as a result of direct
effects. It is worth emphasizing that the proposed intervention tends to reduce long and/or very
frequently repeated demands for medication and with that improve the global safety rate and
QOL.
3. Ethics and dissemination:
The protocol was approved by an Independent Ethics Committee of FEFyM (Fundación de
Estudios Farmacológicos y Medicamentos/Foundation for Pharmacological Studies and Drugs,
Buenos Aires) and by regulatory authorities of Argentina (ANMAT, Dossier # 61409-8 of April
20th, 2009), following the law of Habeas Data and psychotherapeutic drug control. All
participants will agree voluntarily to participate, after having obtained the informed consent and
signed it, in accordance with local regulations and standards of bioethics. The study is
developed under Phase IV conditions, but the full therapeutic intervention has no cost for the
patient under the study protocol. The present study protocol is still ongoing and patient
recruitment is in process.
It is our expectation to see a decline within anxiety scores and allostatic load, reflected by
biochemical variables and an improvement in cognitive performance with a better quality of life,
even in patients treated with low doses of alprazolam. We expect to elucidate improvements in
cognitive variables, such as time-space orientation, immediate memory, concentration and
calculation, long-term memory, language and praxis, as well as a tendency to an overall
improvement of QOL scores.
If the GEMA project fulfills its basic purposes, an original road would open for the search of
specific modulators for the treatment of mental illnesses. It has been stated by health
economists, the “macro” conditions of the cost/health ratio are given more by classic or vintage
drugs rather than by innovations, many of which disappear after a promotional period. In this
sense alprazolam has persisted as a medical tool for several years, and regardless of what
formal pharmaceutical clinical trials and the data that investigators might justify their
permanence. The GEMA project is also an update in the knowledge of the impact of this
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compound for patients with anxiety and allostatic load, and is also a project designed to define a
new profile for the biomedical use of alprazolam.
4. Discussion
GEMA project proposes an intervention, designed to determine if minimum doses of alprazolam
treatment ameliorates additional neurobiochemical variables altered by stress as usually are
detected in anxiety patients, and if it contributes to prevent the impact of allostatic loads,
promoting resilience, better cognitive performance and making healthier decisions translated
into a better quality of life.
If the hypothesis is confirmed, a new paradigm would come into being, which would renew
interest in the clinical management of these compounds, beyond their typical use as anxiolytic
agents. In short, the proposal is to treat each episode more efficiently in order to reduce the
time required to achieve positive results (bene facere) while decreasing the tendency to
excessive consumption of drugs (non nocere).
5- List of abbreviations
HPA (hypotalamo pituitary adrenal)
CRH (corticotrophin releasing hormone)
GAD (general anxiety disorders)
PFC (prefrontal cortex)
MHPG (3-methoxy- 4- hydroxyphenilglycol)
BZD (benzodiazepines)
NICE (National Institute for Health and Clinical Excellence, UK)
NA (noradrenaline)
STH (somatotropic hormone)
GHRH (growth-hormone-releasing hormone)
MDD (major depressive disorder)
CRH (corticotropin releasing hormone)
MAO (monoamine oxidase)
STAI (state -trait anxiety inventory)
HARS (Hamilton anxiety rating scale)
NEO FFI (NEO five factor inventory for neuroticism)
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BMI (body mass index)
HDL (high density lipoprotein)
LDL (low density lipoprotein)
CGI (Clinical Global Impression Scale)
MEC (The Mini-Mental State)
UKU (Udvalg fur klinische undersogelser)
GCP (good clinical practice)
QOL (quality of life)
BUN (blood urea nitrogen)
CPT (continuous performance test
DST (digit symbol test)
DST (digit span test)
VFT (verbal fluency test
FPT (five point test)
RTCT (revised Taylor complex test)
ST (stoop test).
6- Competing interests
This work is carried out under a grant awarded by Gador SA, Buenos Aires (2010-2012) and
funds of the Instituto de Biociencias Henri Laborit, Córdoba. CS and LD are consultants at
Gador SA, and DN and EJAR are investigators employed by Gador SA, and CR does not
declare any conflict of interest. Gador SA, Darwin 429, Buenos Aires City, Argentina.
7- Authors’ contributions
CS, CR, DN and EJAR have made substantial contributions to conception and design. LD has
been involved in drafting and revising the manuscript for intellectual content.
EJAR has given final approval of the version to be published.
8- Author’ information
CS and CR: Institute of Biosciences Henri Laborit, Córdoba
DN and EJAR: Scientific Direction, Gador SA, Buenos Aires, Argentina.
LD: Institute of Biology and Neuroscience E de Robertis, CONICET, Argentina.
9- Acknowledgments
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Authors thank the participation and contributions of the work team involved in GEMA.
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12. Figure Legends
Table 1 Data come from different experimental models and do not represent human SNC condition, either in healthy or under allostatic loads. Gray boxes presume positive for the cognitive development under drug therapy. Adapted from references
4964
Figure 1 Percentage of reduction of salivary cortisol and MHPG levels among alprazolam (0.75 to 3.0 mg/day) responder patients. Modified from [47]
Figure 2 Upper panel: Repeated stress (S-blue) conditions plus accumulation of allostatic loads (AL-red) result in chronic stress, and with time loss of the individual resilience (R-green) levels. Anxiety (A-yellow) emerges as a reaction of an excessive stress applied to the body. Under chronic stress, cognitive resources are impaired and with it the possibilities of the affected subject to maintain its quality of life. Consequently, the affected live with a neurobiological environment poorly efficient to solve the daily challenges of life. Middle panel: With a short-term anxiolytic use of alprazolam, symptoms are reduced but demands of treatment persist, as well as the impact of the AL, and the decay of R, are not being significantly modified. Lower panel: With prolonged use of low-dose alprazolam anti-stress biochemical effects, plus a consequent increase of cognitive functions, may allow the progressive recovering of R, shorten S duration, and may further reduce the impact of AL. Under these new biological conditions the subject increased its chances to seek for an improved quality of life and may there of shorten the demand of palliative medication. Figure 3 Stage I: Short term evaluation of 0.75 to 3.0 mg/day alprazolam on biochemical variables related to chronic stress. Stage II: Short term evaluation of 0.75 to 3.0 mg/day alprazolam on cognitive and clinical variables related to chronic stress. Stage III: Intermediate term evaluation of low alprazolam doses in cognitive, clinical and quality of life indicators.
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Figure 1. MHPG and cortisol salivary levels
0 20 40 60 80 100
5% of reduction (salivary levels)
up to 5% of reduction (salivary levels)
58.33
41.67
95.65
4.35
Cortisol
MHPG
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Figure 2. Hypothesis of the therapeutic intervention postulated at the GEMA project.
No adequate therapy
Traditional use of
benzodiazepines
The GEMA project
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Figure 3. Development stages of the GEMA Project
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Table 1 Comparative affinity of different anxiolytics benzodiazepines to GABAA subtypes receptors. GABA receptor subtype
α1 α2 α3 α5 β3 γ2 SNC
depression
Motor
impairment
amnesia
Anxiolysis Disinhibition
Anticonvulsive Mio-relaxation
Anxiolysis
Learning memory
Anticonvulsive Muscle relaxation
sedation
Addiction respiratory
depression
Alprazolam Low High Moderate Low? Low High
Clonazepam Moderate High High Low? Moderate High
Diazepam Moderate High High Moderate? Moderate Moderate
Lorazepam Low High High Moderate? Low High
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SPIRIT 2013 Checklist: Recommended items to address in a clinical trial protocol and
related documents*
Section/item ItemNo
Description
Administrative information
Title 1 Descriptive title identifying the study design, population, interventions,
and, if applicable, trial acronym
Trial registration 2a Trial identifier and registry name. If not yet registered, name of
intended registry (page 1- abstract)
2b All items from the World Health Organization Trial Registration Data
Set (attached as additional information)
Protocol version 3 Date and version identifier( attached as additional information)
Funding 4 Sources and types of financial, material, and other support (pag 16 )
Roles and
responsibilities
5a Names, affiliations, and roles of protocol contributors (page 1 and 16)
5b Name and contact information for the trial sponsor (page 16)
5c Role of study sponsor and funders, if any, in study design; collection,
management, analysis, and interpretation of data; writing of the report;
and the decision to submit the report for publication, including whether
they will have ultimate authority over any of these activities (page 16)
5d Composition, roles, and responsibilities of the coordinating centre,
steering committee, endpoint adjudication committee, data
management team, and other individuals or groups overseeing the
trial, if applicable (see Item 21a for data monitoring committee)
(page 16)
Introduction
Background and
rationale
6a Description of research question and justification for undertaking the
trial, including summary of relevant studies (published and
unpublished) examining benefits and harms for each intervention
(point 1: pages 2-6)
6b Explanation for choice of comparators
Objectives 7 Specific objectives or hypotheses (point 1: page 7)
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Trial design 8 Description of trial design including type of trial (eg, parallel group,
crossover, factorial, single group), allocation ratio, and framework (eg,
superiority, equivalence, noninferiority, exploratory) (point 2 page 8)
Methods: Participants, interventions, and outcomes
Study setting 9 Description of study settings (eg, community clinic, academic hospital)
and list of countries where data will be collected. Reference to where
list of study sites can be obtained (point 2 page 8-9)
Eligibility criteria 10 Inclusion and exclusion criteria for participants. If applicable, eligibility
criteria for study centres and individuals who will perform the
interventions (eg, surgeons, psychotherapists) (point 2 page 8-9)
Interventions 11a Interventions for each group with sufficient detail to allow replication,
including how and when they will be administered (point 2.5 page 10)
11b Criteria for discontinuing or modifying allocated interventions for a
given trial participant (eg, drug dose change in response to harms,
participant request, or improving/worsening disease) (point 2.2 pag 9)
11c Strategies to improve adherence to intervention protocols, and any
procedures for monitoring adherence (eg, drug tablet return,
laboratory tests)
11d Relevant concomitant care and interventions that are permitted or
prohibited during the trial) (point 2.2 pag 9)
Outcomes 12 Primary, secondary, and other outcomes, including the specific
measurement variable (eg, systolic blood pressure), analysis metric
(eg, change from baseline, final value, time to event), method of
aggregation (eg, median, proportion), and time point for each
outcome. Explanation of the clinical relevance of chosen efficacy and
harm outcomes is strongly recommended (point 2.6 pages 10-12)
Participant
timeline
13 Time schedule of enrolment, interventions (including any run-ins and
washouts), assessments, and visits for participants. A schematic
diagram is highly recommended (see Figure) (point 2.5 page 10)
Sample size 14 Estimated number of participants needed to achieve study objectives
and how it was determined, including clinical and statistical
assumptions supporting any sample size calculations (point 2.4 page
10)
Recruitment 15 Strategies for achieving adequate participant enrolment to reach
target sample size
Methods: Assignment of interventions (for controlled trials)
Allocation:
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Sequence
generation
16a Method of generating the allocation sequence (eg, computer-
generated random numbers), and list of any factors for stratification.
To reduce predictability of a random sequence, details of any planned
restriction (eg, blocking) should be provided in a separate document
that is unavailable to those who enrol participants or assign
interventions
Allocation
concealment
mechanism
16b Mechanism of implementing the allocation sequence (eg, central
telephone; sequentially numbered, opaque, sealed envelopes),
describing any steps to conceal the sequence until interventions are
assigned
Implementation 16c Who will generate the allocation sequence, who will enrol participants,
and who will assign participants to interventions
Blinding
(masking)
17a Who will be blinded after assignment to interventions (eg, trial
participants, care providers, outcome assessors, data analysts), and
how
17b If blinded, circumstances under which unblinding is permissible, and
procedure for revealing a participant’s allocated intervention during
the trial
Methods: Data collection, management, and analysis
Data collection
methods
18a Plans for assessment and collection of outcome, baseline, and other
trial data, including any related processes to promote data quality (eg,
duplicate measurements, training of assessors) and a description of
study instruments (eg, questionnaires, laboratory tests) along with
their reliability and validity, if known. Reference to where data
collection forms can be found, if not in the protocol
18b Plans to promote participant retention and complete follow-up,
including list of any outcome data to be collected for participants who
discontinue or deviate from intervention protocols
Data
management
19 Plans for data entry, coding, security, and storage, including any
related processes to promote data quality (eg, double data entry;
range checks for data values). Reference to where details of data
management procedures can be found, if not in the protocol
Statistical
methods
20a Statistical methods for analysing primary and secondary outcomes.
Reference to where other details of the statistical analysis plan can be
found, if not in the protocol
20b Methods for any additional analyses (eg, subgroup and adjusted
analyses)
20c Definition of analysis population relating to protocol non-adherence
(eg, as randomised analysis), and any statistical methods to handle
missing data (eg, multiple imputation)
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Methods: Monitoring
Data monitoring 21a Composition of data monitoring committee (DMC); summary of its role
and reporting structure; statement of whether it is independent from
the sponsor and competing interests; and reference to where further
details about its charter can be found, if not in the protocol.
Alternatively, an explanation of why a DMC is not needed
21b Description of any interim analyses and stopping guidelines, including
who will have access to these interim results and make the final
decision to terminate the trial
Harms 22 Plans for collecting, assessing, reporting, and managing solicited and
spontaneously reported adverse events and other unintended effects
of trial interventions or trial conduct
Auditing 23 Frequency and procedures for auditing trial conduct, if any, and
whether the process will be independent from investigators and the
sponsor
Ethics and dissemination
Research ethics
approval
24 Plans for seeking research ethics committee/institutional review board
(REC/IRB) approval (point 2.3 page 9)
Protocol
amendments
25 Plans for communicating important protocol modifications (eg,
changes to eligibility criteria, outcomes, analyses) to relevant parties
(eg, investigators, REC/IRBs, trial participants, trial registries, journals,
regulators)
Consent or assent 26a Who will obtain informed consent or assent from potential trial
participants or authorised surrogates, and how (see Item 32)
26b Additional consent provisions for collection and use of participant data
and biological specimens in ancillary studies, if applicable
Confidentiality 27 How personal information about potential and enrolled participants will
be collected, shared, and maintained in order to protect confidentiality
before, during, and after the trial
Declaration of
interests
28 Financial and other competing interests for principal investigators for
the overall trial and each study site
Access to data 29 Statement of who will have access to the final trial dataset, and
disclosure of contractual agreements that limit such access for
investigators
Ancillary and
post-trial care
30 Provisions, if any, for ancillary and post-trial care, and for
compensation to those who suffer harm from trial participation
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Dissemination
policy
31a Plans for investigators and sponsor to communicate trial results to
participants, healthcare professionals, the public, and other relevant
groups (eg, via publication, reporting in results databases, or other
data sharing arrangements), including any publication restrictions
31b Authorship eligibility guidelines and any intended use of professional
writers
31c Plans, if any, for granting public access to the full protocol, participant-
level dataset, and statistical code
Appendices
Informed consent
materials
32 Model consent form and other related documentation given to
participants and authorised surrogates
Biological
specimens
33 Plans for collection, laboratory evaluation, and storage of biological
specimens for genetic or molecular analysis in the current trial and for
future use in ancillary studies, if applicable
*It is strongly recommended that this checklist be read in conjunction with the SPIRIT 2013
Explanation & Elaboration for important clarification on the items. Amendments to the
protocol should be tracked and dated. The SPIRIT checklist is copyrighted by the SPIRIT
Group under the Creative Commons “Attribution-NonCommercial-NoDerivs 3.0 Unported”
license.
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Impact of alprazolam in allostatic load and neurocognition of patients with anxiety disorders and chronic stress
(GEMA): Observational study protocol
Journal: BMJ Open
Manuscript ID: bmjopen-2014-007231.R1
Article Type: Protocol
Date Submitted by the Author: 21-May-2015
Complete List of Authors: Soria, Carlos; Institute of Biosciences Henri Laborit, Córdoba Remedi, Carolina; Institute of Biosciences Henri Laborit, Córdoba Nuñez, Daniel; GADOR SA, Scientific Direction D'Alessio, Luciana; Cell Biology and Neuroscience Institute, CONICET Roldán, Emilio; GADOR SA, Scientific Direction
<b>Primary Subject Heading</b>:
Mental health
Secondary Subject Heading: Mental health
Keywords: CLINICAL PHARMACOLOGY, Adult psychiatry < PSYCHIATRY, Anxiety disorders < PSYCHIATRY
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Impact of alprazolam in allostatic load and neurocognition of patients with anxiety
disorders and chronic stress (GEMA): Observational study protocol
Authors
Carlos A. Soria, Carolina Remedi, Daniel A. Núñez, Luciana D’Alessio, Emilio J. A.
Roldán.
Institutions
Institute of Biosciences Henri Laborit, Córdoba, Argentina.
Scientific Direction Gador SA, Buenos Aires, Argentina.
Corresponding Author:
Luciana D’Alessio, Phd, MD
email: luladalessio@gmail.com , dcientifica@gador.com.ar
Darwin 429 (C1414CUI) CABA
Buenos Aires, Argentina
Phone 00541169518500
Abstract
Introduction
The allostatic load model explains the additive effects of multiple biological
processes that accelerate pathophysiology related to stress, particularly in central
nervous system. Stress related mental condition such as anxiety disorders and
neuroticism (a well-known stress vulnerability factor), have been linked to
disturbances of hypotalamo-pituitary-adrenal (HPA) with cognitive implications.
Nevertheless there are controversial results in the literature and there is a need to
determine the impact of the psychopharmacological treatment on allostatic load
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parameters and in cognitive functions. GEMA (Gador study of Estres Modulation by
Alprazolam), aims to determine the impact of medication on neurobiochemical
variables related to chronic stress, metabolic syndrome, neurocognition and quality
of life in patients with anxiety, allostatic load and neuroticism.
Methods/analysis: In this observational prospective phase IV study, highly
sympthomatic patients with anxiety disorders (six or more points in the Hamilton-A
scale), neuroticism (more than 18 points in the NEO-FFI scale), an allostatic load
(three positive clinical or biochemical items at Crimmins and Seeman criteria) will
be included. Clinical variables of anxiety, neuroticism, allostatic load,
neurobiochemical studies, neurocognition and quality of life will be determine prior
and periodically (one, two, four, eight, and twelve weeks) after treatment (on
demand of alprazolam from 0.75 mg/day to 3.0 mg/day). A sample of n =55/182
patients will be consider enough to detect variables higher than 25% (pre-
treatment versus post-treatment or significant correlations) with a 1-ß power of 0-
80. T-test and/or non parametric test, and Pearson Test for correlation analysis will
be determine.
Ethics and dissemination: This study protocol was approved by an Independent
Ethics Committee of FEFyM (Foundation for Pharmacological Studies and Drugs,
Buenos Aires) and by regulatory authorities of Argentina (ANMAT, Dossier # 61409-
8 of April 20th, 2009), following the law of Habeas Data and psychotherapeutic drug
control.
Limitations of this study
One of the important biases of the project is the lack of a parallel control group,
which is not regulatory allowed in our area, in a phase IV trial with highly
symptomatic patients with inherent risks. As the project moves forward, however,
the project itself can be justified in a more advanced stage, for example, in a face-
to-face comparison of different schemes.
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Another difficulty is the existence of knowledge gaps among the clinical features,
cognitive observations and variables measured in laboratories. So, we assume that
as we move forward in the associations among them, other hypotheses on
mechanism of action may arise that can be formulated and tested by other
investigators.
Key Words: Alprazolam, benzodiazepines, anxiety disorders, allostatic load, chronic
stress, cognition.
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1- Introduction
Stress related mental condition such as anxiety disorders are frequently
associated with chronic stress [1,2,3]. If the stress response is excessive and
prolonged, the cost of reinstating homeostasis become too high, a condition that is
termed allostatic load (an indicator of the of adverse health outcomes because of
physiologic dysregulation) [4-10].
Anxiety disorders have been linked to disturbances of hypotalamo-pituitary-
adrenal (HPA) axis with controversial results[11-13]. Hormones such as cortisol and
CRH (corticotrophin releasing hormone) have been associated to states of fear and
anxiety in experimental models [14,15]. Furthermore, increased basal levels of
cortisol were found in patients with panic disorders and in patients with GAD
(general anxiety disorders) [16,17]. Nevertheless others authors did not find any
association in cortisol levels and anxiety scale measures[13] or as well found a
reduction in cortisol awakening response among older patients with anxiety
disorders[18]. On the contrary studies performed among posttraumatic stress
disorder patients showed lower levels of cortisol[19].
Similar pattern of cortisol levels were also found in patients with neuroticism
(a predisposition to respond with intense emotional reaction to psychosocial
stressors) [1,2]. Neuroticism is a well-known stress vulnerability factor that is
robustly associated with psychiatric disorders such as anxiety and depression [20].
Neuroticism and stress domains do not measure an identical construct
nevertheless, it has been reported as that cumulative stress was more likely to
precipitate psychiatric symptoms in vulnerable individuals with high levels of
neuroticism [20] and furthermore, these individuals perceive and have more
stressors, respond exaggeratedly to them, and require more time to recover from
them [1,2,21].
During stress activation both HPA axis and the noradrenergic (NA) system
seem to function in a synchronous fashion and these two systems are crucial for
stress response in mammals [6,7]. There is substancial evidence that anxiety in
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humans is associated with high levels of 3-methoxy-4-hydroxyphenilglycol (MHPG),
a principal NA metabolite, and salivary determination of MHPG may be a useful
marker for detecting changes in catecholamine metabolism [22-24], however there
is a need for determine by objective measures the impact of stress mediators on
allostatic load and cognitive functioning in these patient and the impact of the
psychopharmacological treatment on these parameters.
The allostatic load model has been particularly informative in specifying the
additive effects of multiple biological processes that contribute to wear and tear on
tissues and accelerate pathophysiology related to stress, particularly in central
nervous system [5,6,7,25,10]. These allodynamic processes can be adaptive in the
short term (allostasis) and maladaptive in the long term (allostatic load). The brain
is the central organ of stress processes and allodynamic adaptation and it is a key
target of allostasis, thus enable protection and adaptation. Consequently allostatic
load can affect brain plasticity since adjustments of biological systems [9].
It has been demonstrated that stressful experiences have functionally-
relevant effects on dendritic arbor and spine/synapse number in many brain
regions, including the hippocampus, amygdala and the prefrontal cortex (PFC) with
effects not only on cognitive function but also on emotional regulation and other
self-regulatory behaviors and upon neuroendocrine and autonomic function[9]. By
compromising hippocampal function, allostatic load and elevated levels of
glucocorticoids impair normal hippocampal function as well memory processes
(declarative memory)[7,26,27]. Furthermore, stress-related effects upon the PFC
has an importance in working memory process (the ability to keep events in
mind)[28] and self regulatory behaviors. The dorso lateral PFC is responsible for
planning approaches and sequences of behavior that are required for goal-directed
behavior. This process is critical of executive function[7,28].
In experimental models, chronic restraint stress caused spine loss and
debranching of dendrites on PFC neurons and impaired working memory[7,9,29]. In
humans, adverse childhood experiences were associated with smaller volume of the
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PFC greater activation of the HPA axis, and elevation in inflammation levels [30].
Furthermore, it has been shown that stress and allostatic load increases
interference from irrelevant information and thus impairs selective attention
[31],and high levels of cortisol in response to stressors showed lower performance
in working memory tasks in normal adults [32].Also, it has been found that anxiety
disorders affect cognitive style of functioning, reversed by alprazolam treatment
[33].
Following the NICE (National Institute for Health and Clinical Excellence, UK)
guidelines, treatment of a person with GAD (general anxiety disorder) and chronic
stress consists of psychotherapy (cognitive behavioral therapy), but when
symptoms induce a significant functional impairment, it should be resorted with
drugs, generally antidepressant and benzodiazepines [34]). Benzodiazepines (BZD)
are GABAergic substances with anxiolytic and sedative activity available since the
sixties. The therapeutic use to depress acute and subacute symptoms of anxiety
disorders has gained great popular acceptance up to the present[35].Stress related
mental conditions such as anxiety disorders and neuroticism are treated with
psychotherapy and psychopharmacologic drugs, which are primarily prescribed to
reduce pathological symptoms but the impact on the underlying disorder is
generally less understood. Furthermore, many patients persist with symptoms and
consequently require the use of BZD for longer periods of time. Nevertheless, this
indication is an off-label use is therefore regarded with caution because of the
possibility of favoring dependence on medication and other undesired events such
as cognitive impairment[35,36,37].However when the stressors persist, cognitive
variables could be affected, making it difficult for the patient to take decisions that
are favorable for QOL (quality of life)[34,37,38]
The objective of treatment is not only to suppress symptoms of anxiety, but
also to prevent a chronic outcome, and to guide patients to positive behaviors
“decisions making” for a better QOL[5].
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As BZD have anxioliytic and sedating properties, it is expected that they
influence the stress system. As well most studies showed a decrease in cortisol
levels [39-43], some studies reported mixed results [43-45], and these
inconsistencies may be explained by differences in patients groups, dosages, and
BZD used among studies [43]. Some studies yield a reduction of variations in
noradrenaline (NA) metabolite levels (MHPG) "noradrenergic volatility"[46], and in
circulating and salivary cortisol levels using alprazolam in anxious patients
[22,39,41] and in normal subjects [47] or as well an increase levels of
dehydroepiandrostrerone (an anti-cortisol mediator)[48]. Nevertheless many
controversial findings were found depending on age and gender: younger anxious
patients showed higher levels of salivary cortisol awakening response [13,18], but
other authors found high salivary cortisol levels associated with severity of late-life
generalized anxiety disorders in older adults [17]. Furthermore women seem to
yield a different profile in the cortisol response to stress, in relation to men on
response to alprazolam treatment [42]. Previous data from our clinical practice
showed that the administration of alprazolam results in a fast and significative
decrease of MHPG concentrations and a decrease of noradrenergic volatility
accompanied by a slower but equally sustained decrease in salivary cortisol
levels[49](Figure 1).The direct action of alprazolam on GABAergic neurons, and
secondarily, on catecholaminergic and serotoninergic neurotransmitters may induce
the indirect anti-stress target reducing cortisol plasma levels. The reduction in
"noradrenergic volatility" defined by the sudden variations in the levels of NA
(noradrenaline) and its metabolite MHPG observed in patients with GAD, stress-
related diseases and depressive states, may also be involved [46,50-54].
Nevertheless the other aspect of benzodiazepines treatment are cognitive
adverse effects, and identification of them requires dedicated monitoring of
cognitive function, which is often absent from routine hospital and ambulatory
clinical care [55].The short term effects of BZD on cognition are well known and
they are mediated through an agonist action on receptors of γ aminobutyric acid
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(GABA A)[55,56]. A systematic review of the association between benzodiazepines
and cognition found that cognitive impairment was presumed to be transient and
reversible in most studies [38,55]. Nevertheless the long term adverse effects of
BZD on cognition are still debated[36,55] and available data shows conflicting
results. Some authors found an improvement in memory and an increased risk of
dementia in prospective population based studies, performed in elderly patients
with long term use of BZD[36,57-60]. Furthermore longer acting BZD were
associated with greater risk of developing dementia compared with shorter acting
benzodiazepines [36]. Nevertheless, other concomitant factors could be also
responsible of dementia in such studies. Indeed, a higher degree of neuroticism in
midlife was associated with increased risk of dementia and long-standing distress,
in a population-based study [61]. Other studies showed not conclusive
results[62,63] or even reported a potential protective effect on behavioral
disturbances associated to dementia: the role of GABA in dementia has yet to be
fully elucidated and behavioral symptoms associated with dementia may be a direct
or indirect result of GABAergic dysfunction[64,65]. Furthermore, in young adults,
BZD may affects cognition differentially depending on gender: cognitive alteration
of long-term memory have been reported in women, whereas there was no
significant affectation in men [63].
In this study we selected alprazolam (a triazolo-benzodiazepine) for different
reasons: is a high-potency benzodiazepines with an indirect activity through 5-HT1A
serotonergic receptors, which would produce a regulation of the hyperactive
noradrenergic pathways and, through them, a reduction of noradrenergic volatility.
The reduction of noradrenergic volatility by high-potency benzodiazepines would
have a preventive effect via the noradrenergic system, beyond the central
GABAergic anxiolytic effects. The level of concentration and distribution of
alprazolam within the CNS is effective when it is administered in relatively low
doses, with a differential impact on GABAA receptors, upregulated in individuals
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under chronic stress, and with less impact on cognitive functions because of the
shorter acting [36,51,60,66,67](Table 1).
In the present study GEMA (Gador study of Estres Modulation by Alprazolam), the
following research questions were formulated: How treatment anxiety with
benzodiazepines impacts on allostatic load and neuroticism? Does allostatic load
and chronic stress affect cognition process? Do benzodiazepines affect cognitive
function positively in stressed patients?
1.4 General Hypothesis
In the present study we aim to investigate biochemical and neurocognitive
variables in patients with high anxiety levels associated to chronic stress, and
neuroticism (a clinical condition that indicate an exaggeratedly response to the
stressors that require more time to recover from them) [51,68,69] and the impact
of intermediate term treatment (12 weeks) with low doses of alprazolam.
In clinical practice high symptomatic anxiety disorders are frequently associated
with chronic stress and neuroticism (a well-known stress vulnerability factor) [20]
and overall all of these factors increase the vulnerability to diseases and reduce
resilience. In these high symptomatic patients the use of benzodiazepines is
required for longer periods of time. Nevertheless, this indication is an off-label use,
and is therefore regarded with caution because of the possibility of favoring
dependence on medication and other undesired events such as cognitive functions,
reflexes and alert states [35].
The global hypothesis under investigation is if neurobiochemical indicators of
allostatic load (noradrenergic metabolites and cortisol levels) are modulated during
treatment with alprazolam, and if there are neurocognitive consequences when
treatment maintains neurobiochemical parameters within the normal range. A
pharmacological modulation of such mediators may contribute to reduce the impact
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of allostatic load, promoting resilience and better biochemical and neurocognitive
performance translated in a better quality of life.
This study is based on the hypothesis that high potency benzodiazepines such as
alprazolam may serve as more than single anxiety suppressive agents reducing
allostatic load and facilitating adaptative responses. GEMA project proposes an
intervention designed to determine if minimum doses of alprazolam adjust
neurobiochemical mediators, neurocognition and affect quality of life, altered by
stress (Figure 2)
The question attempted to be solved is, whether anxiolytic pharmacology
intervention with alprazolam in these high symptomatic patients modify allostatic
load, neurocognition performance and quality of life, beyond the known anxiolytic
effect.
1.5 Objectives (End-points): The project consists of three complementary stages
(Figure 3).
1.5.1 To determine whether clinical and biochemical markers of allostatic load, tend
to normalize with 12 weeks of therapeutic intervention.
1.5.2 To determine if neurocognitive performance is modified after treatment and if
there is a positive correlation with biochemical variables of allostatic load and with
therapeutic levels.
1.5.3 To assess QOL before and after treatment
Secondarily, the purpose of this study is to report conditions of failure due to lack
of response or emergence of undesired events, including adverse events, and
possible situations of habituation to or dependence on alprazolam.
2- Methods and analysis:
2.1 Protocol Design:
The GEMA project (Gador study of Estres Modulation by Alplax), is a prospective
phase IV clinical trial. The study began in 2010 and is still in advance. This is a
prospective observational study, controlled with on demand assignment of
alprazolam doses in variable doses. It is a phase IV study reported to regulatory
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authorities in compliance with the rules in force and consequently enclosed for the
use of experimental controls or interventions.
The presence of variable doses will allow us to establish a continuous range of
dose-response.
2.2 Inclusion Criteria
In this study highly sympthomatic patients over than 21 years, who have at the
time of admission anxiety symptoms (six or more points in the Hamilton-A scale),
[70] neuroticism (18 or more points in the NEO-FFI scale), [71] allostatic load
expressed by at least 3 positive clinical or biochemical items (Crimmins and
Seeman criteria) with normal cognitive performance (25 or more points in the
Folstein’s Mini-Mental State Examination)[72] were included. Allostatic load
measurement included markers of hypothalmic pituitary axis, sympathetic nervous
system, cardio-vascular system, metabolic system, renal function, lung function,
and markers of inflammation [10].
2.3 Exclusion Criteria
Other psychiatric disorders listed under Axis I of the DSM-IV(Diagnostic and
Statistical Manual of Mental Disorders, Fourth Edition)[73]. Patients requiring other
psychotropics added-on to alprazolam, sympathicomimetics, corticosteroids or
another medication that might kinetically or dynamically interfere with alprazolam.
Patients who had received fluoxetine in the previous 4 weeks or MAO (monoamine
oxidase) inhibitors in the previous 2 weeks, or showing a history of hypersensitivity
to alprazolam. Women with confirmed or suspected pregnancy, or subjects affected
with concomitant conditions, severe or unstable, in the cardiovascular system,
gastrointestinal, hepatic, renal, neurological and other systems.
Patients will be withdrawn from this study if dose is prematurely modified, or if
patient decides not to continue with the administered drug, has a related adverse
event that modifies his/her daily performance, or if the investigator considers that
the scheme is not safe or beneficial for the patient according to his/her current
clinical status.
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2.4
Sample Size: Changes in variables higher than 25% (pre-treatment versus post-
treatment or significant correlations) with a 1-ß power of 0-80 are estimated to be
detected. For the first phase, an n = 29-55 cases, for the second phase an n = 112
and for the third an extension of the sample to 182 assessable patients, according
to a PC program (Statistica, Tulsa) will be considered enough.
2.5 Therapeutic Scheme and Dose Assignment
Patients will be extensively assessed by a team of medical doctors, clinicians and
psychiatrists, who decide the most appropriate approach for each case. The
principal investigator will determine the initial dose of alprazolam, usually from 0.75
mg/day to 3.0 mg/day, according to the presence of anxiety symptoms and
according to patient’s tolerance. The ongoing scheme of alprazolam will be
maintained for 3 months. Thereafter the investigator can decide to continue or
discontinue treatment gradually, by reducing 0.5 mg every 3 days.
2.6 Patient Assessment.
Study participants will be evaluated prior to treatment (week-1), on the day
treatment started (day 0), and during treatment on weeks 1, 2, 4, 8 and 12.
Clinical variables of anxiety, neuroticism and allostatic load, neurocognitive and
neurobiochemical studies will be determined.
Anxiety assessment: Two scales will be rated: 1 - The Hamilton Anxiety Rating
Scale (HARS) of 14 items. [70] The 0-5 score indicates absence of anxiety, 6-14
mild anxiety and more than 15 moderate to severe anxiety. 2 - The State-Trait
Anxiety Inventory (STAI) of 40 items[74], which assesses the current state and the
anxiety trait as a stable tendency. The scale scores a maximum of 60 points and
percentiles are described for gender and age.
Neuroticism assessment: The NEO five factor inventory (NEO FFI)[71] is an
abbreviated measure of personality factors related to neuroticism, extraversion,
openness, agreeableness, and conscientiousness. Allostatic load determination:
Clinical and biochemical evaluations will be determined: 1- Systolic blood pressure
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will be measured in the supine and standing position, with a previous 2-3 minutes
rest A systolic pressure > 140mmHg and/or a diastolic pressure < 90 mmHg will be
considered a positive criteria.[75]2- Heart frequency: a frequency exceeding 90
bpm (beats per minute) in supine or standing position with a previous 2-3 minutes
rest is considered a positive pulse. 3- BMI (body mass index) and waist-hip ratio
are measured while patient is standing and breathing normally, horizontal over the
edge of iliac crests, using a tight tape without compressing the skin. 4- Laboratory
biochemical tests in serum samples include total cholesterol, and HDL and LDL
fractions, glycosylated hemoglobin, fibrinogen, albumin, and C reactive protein.
Furthermore, plasma noradrenaline and cortisol and salivary MHPG (3-methoxy 4
hydroxyphenilglycol) and cortisol will be determined.
General condition and safety: 1-Clinical Global Impression Scale (National Institute
of Mental Health 1976) (CGI), which includes two subscales, 1a - clinical profile
severity (CGI-S) and 1b-clinical profile improvement (CGI-MJ) with 8 points each
one. 2-The Mini-Mental State (MEC) quantifies cognitive impairment in time-space
orientation, immediate memory, concentration and calculation, long-term memory,
language and praxis [72] 3- UKU scale (Udvalg fur klinische undersogelser)[76] to
determine side effects; with 56 items: the first 9 are for psychic events, 8 for
neurologic, 11 for autonomic and 25 for others. The last two items evaluate side
events interference with the patient's daily performance and their impact on
medication intake (none, dose reduction, withdrawal or change). In each one, the
investigator assesses whether the event was related to drug treatment or not. 4-
Additional safety assessments include respiratory rate and laboratory tests:
complete blood count, creatinine, BUN (blood urea nitrogen), total bilirrubin,
alkaline phosphatase, liver enzymes and human chorionic gonadotropin (hCG-B)
will be determined when appropriate. All patients will be informed about any
eventual depressant effect of alprazolam and risks in activities that require alert
and the consumption of alcohol will be not recommended. They will be also
monitored to detect potential risks of suicide or overdose. All adverse events are
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monitored, classified and reported according to GCP (good clinical practice)
standards of the local regulatory authority (ANMAT and Ministry of Health of the
Province of Córdoba, Argentina).
Cognitive assessment: To quantify the neurocognitive variables; attention, working
memory, executive functions, impulsiveness, verbal and nonverbal fluency the
following tests will be determined. 1- QPSS continuous performance test (CPT)[77];
2- Digit symbol test (DST)[78]; 3- Digit span test (DST)[79]; 4- Verbal fluency test
(VFT)[80]; 5-Five point test (FPT)[81]; 6-revised Taylor complex test (RTCT)[82]
and 7- Stroop test (ST)[83]. All of them will be applied after 6 hours since the last
administration of alprazolam (nadir plasma levels). Cognitive tests and laboratory
tests are blinded to the analyst, the remaining tests are open.
Quality of life (QOL) assessment: The health and quality of life scale, health
questionnaire SF-36 validated Spanish version will be determined. [84]
2.7 Statistical assessment:
The mean, minimum and maximum values of quantitative variables will be
determined. Pre- and post-treatment values will be compared using t-test and/or
non parametric test. Statistical significance will be fixed at p<0.05 (2-sided; β-1
power≥0.80). Correlation analysis will be performed by calculating the Pearson Test
correlation coefficient for ungrouped data; with r>0.80 and p<0.05 (2-sided) strong
correlation and 0.80<r>0.50 and p<0.05 (2-sided), moderate correlation.
Treatment safety: Regarding patient safety, the use of alprazolam, a well known,
low action, nervous system depressant drug, at minimum doses, even when no
symptoms palliation is required, guarantees good tolerability, and no drawbacks are
expected as a result of direct effects. It is worth emphasizing that the proposed
intervention tends to reduce long and/or very frequently repeated demands for
medication and with that improve the global safety rate and QOL.
3. Ethics and dissemination: Approval and Informed Consent
The protocol was approved by an Independent Ethics Committee of FEFyM
(Fundación de Estudios Farmacológicos y Medicamentos/Foundation for
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Pharmacological Studies and Drugs, Buenos Aires) and by regulatory authorities of
Argentina (ANMAT, Dossier # 61409-8 of April 20th, 2009), following the law of
Habeas Data and psychotherapeutic drug control. All participants will agree
voluntarily to participate, after having obtained the informed consent and signed it,
in accordance with local regulations and standards of bioethics. The study is
developed under Phase IV conditions, but the full therapeutic intervention has no
cost for the patient under the study protocol. The present study protocol is still
ongoing and patient recruitment is in process. Trial Registration: GEMA - 20811
4. Expected results
It is our expectation to see a decline within anxiety scores and allostatic load,
reflected by biochemical variables and an improvement in cognitive performance
with a better quality of life, even in patients treated with low doses of alprazolam.
We expect to elucidate improvements in cognitive variables, such as time-space
orientation, immediate memory, concentration and calculation, long-term memory,
language and praxis, as well as a tendency to an overall improvement of QOL
scores. If the GEMA project fulfills its basic purposes, an original road would open
for the search of specific modulators for the treatment of mental illnesses. It has
been stated by health economists, the “macro” conditions of the cost/health ratio
are given more by classic or vintage drugs rather than by innovations, many of
which disappear after a promotional period. In this sense alprazolam has persisted
as a medical tool for several years, and regardless of what formal pharmaceutical
clinical trials and the data that investigators might justify their permanence. The
GEMA project is also an update in the knowledge of the impact of this compound for
patients with anxiety and allostatic load, and is also a project designed to define a
new profile for the biomedical use of alprazolam.
5. Discussion
GEMA project constitute an observational clinical trial designed to determine
objective measures of stress pathophysiology in high symptomatic patients with
comorbid anxiety, allostatic load and neuroticism treated with minimum doses of
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alprazolam, with the aim to determine neurobiochemical and neuropsychological
variables altered by stress.
Treatment aims to contribute to prevent the impact of allostatic load, promoting
resilience, better cognitive performance and making healthier decisions translated
into a better quality of life. If the hypothesis is confirmed, a new paradigm would
come into being, which would renew interest in the clinical management of these
compounds, beyond their typical use as anxiolytic agents. In short, the proposal is
to treat each episode more efficiently in order to reduce the time required to
achieve positive results (bene facere) while decreasing the tendency to excessive
consumption of drugs (non nocere).
6- List of abbreviations
HPA (hypotalamo pituitary adrenal)
CRH (corticotrophin releasing hormone)
GAD (general anxiety disorders)
PFC (prefrontal cortex)
MHPG (3-methoxy- 4- hydroxyphenilglycol)
BZD (benzodiazepines)
NICE (National Institute for Health and Clinical Excellence, UK)
NA (noradrenaline)
STH (somatotropic hormone)
GHRH (growth-hormone-releasing hormone)
MDD (major depressive disorder)
CRH (corticotropin releasing hormone)
MAO (monoamine oxidase)
STAI (state -trait anxiety inventory)
HARS (Hamilton anxiety rating scale)
NEO FFI (NEO five factor inventory for neuroticism)
BMI (body mass index)
HDL (high density lipoprotein)
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LDL (low density lipoprotein)
CGI (Clinical Global Impression Scale)
MEC (The Mini-Mental State)
UKU (Udvalg fur klinische undersogelser)
GCP (good clinical practice)
QOL (quality of life)
BUN (blood urea nitrogen)
CPT (continuous performance test
DST (digit symbol test)
DST (digit span test)
VFT (verbal fluency test
FPT (five point test)
RTCT (revised Taylor complex test)
ST (stoop test).
7- Competing interests
This work is carried out under a grant awarded by Gador SA, Buenos Aires (2010-
2012) and funds of the Instituto de Biociencias Henri Laborit, Córdoba. CS and LD
are consultants at Gador SA, and DN and EJAR are investigators employed by
Gador SA, and CR does not declare any conflict of interest. Gador SA, Darwin 429,
Buenos Aires City, Argentina.
8- Authors’ contributions
CS, CR, DN and EJAR have made substantial contributions to conception and
design. LD has been involved in drafting and revising the manuscript for intellectual
content.EJAR has given final approval of the version to be published.
9- Author’ information
CS and CR: Institute of Biosciences Henri Laborit, Córdoba
DN and EJAR: Scientific Direction, Gador SA, Buenos Aires, Argentina.
LD: Institute of Biology and Neuroscience E de Robertis, CONICET, Argentina.
10- Acknowledgments
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Authors thank the participation and contributions of the work team involved in
GEMA.
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37. Barker MJ, Greenwood KM, Jackson M et al. Persistence of cognitive effects after withdrawal from long-term benzodiazepine use: a meta-analysis. Arch Clin Neuropsychol 2004;19(3):437–54.
38. Wein AJ. A systematic review of amnestic and non-amnestic mild cognitive impairment induced by anticholinergic, antihistamine, GABAergic and opioid drugs. J Urol 2013;190(6):2168.
39. Curtis GC, Abelson JL, Gold PW. Adrenocorticotropic hormone and cortisol responses to corticotropin-releasing hormone: changes in panic disorder and effects of alprazolam treatment. Biol Psychiatry 1997;41(1):76–85.
40. Abelson JL, Curtis GC, Cameron OG. Hypothalamic-pituitary-adrenal axis activity in panic disorder: effects of alprazolam on 24 h secretion of adrenocorticotropin and cortisol. J Psychiatr Res. 1996;30 (2):79-93.
41. Fries E, Hellhammer DH, Hellhammer J. Attenuation of the hypothalamic-pituitary-adrenal axis responsivity to the Trier Social Stress Test by the benzodiazepine alprazolam. Psychoneuroendocrinology 2006;31(10):1278–88.
42. Pomara N, Willoughby LM, Ritchie JC et al. Sex-related elevation in cortisol during chronic treatment with alprazolam associated with enhanced cognitive performance. Psychopharmacology 2005;182(3):414–9.
43. Manthey L, Giltay EJ, van Veen T, Neven AK, Vreeburg S a, Penninx BWJH, et al. Long-term benzodiazepine use and salivary cortisol: the Netherlands Study of Depression and Anxiety (NESDA). J Clin Psychopharmacol 2010;30(2):160–8.
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44. Laakmann G, Wittmann M, Gugath M et al. Effects of psychotropic drugs (desimipramine, chlorimipramine, sulpiride and diazepam) on the human HPA axis. Psychopharmacology 1984;84(1):66–70.
45. Pomara N, Willoughby LM, Ritchie JC et al. Interdose elevation in plasma cortisol during chronic treatment with alprazolam but not lorazepam in the elderly. Neuropsychopharmacology 2004;29(3):605–11.
46. Coplan JD. Clinical Improvement With Fluoxetine Therapy and Noradrenergic Function in Patients With Panic Disorder. Arch Gen Psychiatry [Internet]. American Medical Association 1997;54(7):643.
7. Zemishlany Z, McQueeney R, Gabriel SM DM. Neuroendocrine and monoaminergic responses to acute administration of alprazolam in normal subjects. Neuropsychobiology. 1990;23(3):124–8.
48. Kroboth PD, Salek FS, Stone RA BR. Alprazolam increases dehydroepiandrosterone concentrations. J Clin Psychopharmacol. 1999;19(2):114–24.
49. Soria CA, Remedi C, Muñóz MS et al. Translational assessment of alprazolam on MHPG, cortisol and cognitive function in patients with neuroticism, allostatic load and anxiety disorders. Int J Neuropsychopharmacol. 2010;13:268.
50. Möhler H. Role of GABAA receptors in cognition. Biochem Soc Trans 2009;37:1328–33.
51. Maubach K. GABAA Receptor Subtype Selective Cognition Enhancers. Curr Drug Targets-CNS Neurol Disord. 2003;2(4):233–9(7).
52. Vasa RA, Pine DS, Masten CL et al. Effects of yohimbine and hydrocortisone on panic symptoms, autonomic responses, and attention to threat in healthy
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54. Juster R, Mcewen BS, Lupien SJ. Neuroscience and Biobehavioral Reviews Allostatic load biomarkers of chronic stress and impact on health and cognition. Neurosci Biobehav Rev 2010;35(1):2–16.
55. Yaffe K, Boustani M. Benzodiazepines and risk of Alzheimer’s disease. BMJ 2014;349:g5312–g5312.
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58. Paterniti S, Dufouil C AA. Long-term benzodiazepine use and cognitive decline in the elderly: the Epidemiology of Vascular Aging Study. J Clin Psychopharmacol. 2002;22(3):285–93.
59. Gallacher J, Elwood P, Pickering J et al. Benzodiazepine use and risk of dementia: evidence from the Caerphilly Prospective Study (CaPS). J Epidemiol Community Heal. 2012;66(10):869–73.
60. Wu CS, Wang SC, Chang IS LK. The association between dementia and long-term use of benzodiazepine in the elderly: nested case-control study using claims data. Am J Geriatr Psychiatry. 2009;17(7):614–20.
61. Johansson L, Duberstein PR, Hällström T. Midlife personality and risk of Alzheimer disease and distress. Neurology. 2014;1–7.
62. Allard J, Artero S RK. Consumption of psychotropic medication in the elderly: a re-evaluation of its effect on cognitive performance. Int J Geriatr Psychiatry. 2004;Oct;18(10).
63. Boeuf-Cazou O, Bongue B, Ansiau D, Marquié J-C, Lapeyre-Mestre M. Impact of long-term benzodiazepine use on cognitive functioning in young adults: the VISAT cohort. Eur J Clin Pharmacol 2011;67(10):1045–52.
64. Fastbom J, Forsell Y WB. Benzodiazepines may have protective effects against Alzheimer disease. Alzheimer Dis Assoc Disord. 1998;12(1):14–7.
65. Sultan S, Gebara EG, Moullec K et al. D-serine increases adult hippocampal neurogenesis. Front Neurosci 2013;7:155.
66. Ellinwood EH, Heatherly DG, Nikaido AM et al. Psychopharmacology Comparative pharmacokinetics and pharmacodynamics of lorazepam, alprazolam and diazepam. Intelligence. 1985;392–9.
67. Tan KR, Rudolph U LC. Hooke don benzodiazepines: GABAA receptor subtypes and addiction. Trends in Neurosci. 2011;34:188–97.
68. Norton PJ PE. A meta-analytic review of adult cognitive-behavioral treatment outcome across the anxiety disorders. J Nerv Ment Dis. 2007;195(6):521–31.
69. Fleishaker JC, Garzone PD, Chambers JH, Sirocco K, Weingartner H. Comparison of the spectrum of cognitive effects of alprazolam and adinazolam after single doses in healthy subjects. Psychopharmacology 1995;120(2):169–76.
70. Hamilton M. Hamilton anxiety rating scale (HAM-A) Br J Med Psychol 1959; 32:50-55.
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12. Figure Legends
Table 1 Data come from different experimental models and do not represent human SNC condition, either in healthy or under allostatic loads. Gray boxes presume positive for the cognitive development under drug therapy. Adapted from references [51][66] Figure 1 Percentage of reduction of salivary cortisol and MHPG levels among alprazolam(0.75 to 3.0 mg/day) responder patients.Modified from [47]
Figure 2
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Upper panel: Repeated stress (S-blue) conditions plus accumulation of allostatic loads (AL-red) result in chronic stress, and with time loss of the individual resilience (R-green) levels. Anxiety (A-yellow) emerges as a reaction of an excessive stress applied to the body. Under chronic stress, cognitive resources are impaired and with it the possibilities of the affected subject to maintain its quality of life. Consequently, the affected live with a neurobiological environment poorly efficient to solve the daily challenges of life. Middle panel: With a short-term anxiolytic use of alprazolam, symptoms are reduced but demands of treatment persist, as well as the impact of the AL, and the decay of R, are not being significantly modified. Lower panel: With prolonged use of low-dose alprazolam anti-stress biochemical effects, plus a consequent increase of cognitive functions, may allow the progressive recovering of R, shorten S duration, and may further reduce the impact of AL. Under these new biological conditions the subject increased its chances to seek for an improved quality of life and may thereof shorten the demand of palliative medication. Figure 3 Stage I: Short term evaluation of 0.75 to 3.0 mg/day alprazolam on biochemical variables related to chronic stress. Stage II: Short term evaluation of 0.75 to 3.0 mg/day alprazolam on cognitive and clinical variables related to chronic stress. Stage III: Intermediate term evaluation of low alprazolam doses in cognitive, clinical and quality of life indicators.
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SPIRIT 2013 Checklist: Recommended items to address in a clinical trial protocol and
related documents*
Section/item ItemNo
Description
Administrative information
Title 1 Descriptive title identifying the study design, population, interventions,
and, if applicable, trial acronym
Trial registration 2a Trial identifier and registry name. If not yet registered, name of
intended registry (page 1- abstract)
2b All items from the World Health Organization Trial Registration Data
Set (attached as additional information)
Protocol version 3 Date and version identifier( attached as additional information)
Funding 4 Sources and types of financial, material, and other support (pag 16 )
Roles and
responsibilities
5a Names, affiliations, and roles of protocol contributors (page 1 and 16)
5b Name and contact information for the trial sponsor (page 16)
5c Role of study sponsor and funders, if any, in study design; collection,
management, analysis, and interpretation of data; writing of the report;
and the decision to submit the report for publication, including whether
they will have ultimate authority over any of these activities (page 16)
5d Composition, roles, and responsibilities of the coordinating centre,
steering committee, endpoint adjudication committee, data
management team, and other individuals or groups overseeing the
trial, if applicable (see Item 21a for data monitoring committee)
(page 16)
Introduction
Background and
rationale
6a Description of research question and justification for undertaking the
trial, including summary of relevant studies (published and
unpublished) examining benefits and harms for each intervention
(point 1: pages 2-6)
6b Explanation for choice of comparators
Objectives 7 Specific objectives or hypotheses (point 1: page 7)
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Trial design 8 Description of trial design including type of trial (eg, parallel group,
crossover, factorial, single group), allocation ratio, and framework (eg,
superiority, equivalence, noninferiority, exploratory) (point 2 page 8)
Methods: Participants, interventions, and outcomes
Study setting 9 Description of study settings (eg, community clinic, academic hospital)
and list of countries where data will be collected. Reference to where
list of study sites can be obtained (point 2 page 8-9)
Eligibility criteria 10 Inclusion and exclusion criteria for participants. If applicable, eligibility
criteria for study centres and individuals who will perform the
interventions (eg, surgeons, psychotherapists) (point 2 page 8-9)
Interventions 11a Interventions for each group with sufficient detail to allow replication,
including how and when they will be administered (point 2.5 page 10)
11b Criteria for discontinuing or modifying allocated interventions for a
given trial participant (eg, drug dose change in response to harms,
participant request, or improving/worsening disease) (point 2.2 pag 9)
11c Strategies to improve adherence to intervention protocols, and any
procedures for monitoring adherence (eg, drug tablet return,
laboratory tests)
11d Relevant concomitant care and interventions that are permitted or
prohibited during the trial) (point 2.2 pag 9)
Outcomes 12 Primary, secondary, and other outcomes, including the specific
measurement variable (eg, systolic blood pressure), analysis metric
(eg, change from baseline, final value, time to event), method of
aggregation (eg, median, proportion), and time point for each
outcome. Explanation of the clinical relevance of chosen efficacy and
harm outcomes is strongly recommended (point 2.6 pages 10-12)
Participant
timeline
13 Time schedule of enrolment, interventions (including any run-ins and
washouts), assessments, and visits for participants. A schematic
diagram is highly recommended (see Figure) (point 2.5 page 10)
Sample size 14 Estimated number of participants needed to achieve study objectives
and how it was determined, including clinical and statistical
assumptions supporting any sample size calculations (point 2.4 page
10)
Recruitment 15 Strategies for achieving adequate participant enrolment to reach
target sample size
Methods: Assignment of interventions (for controlled trials)
Allocation:
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Sequence
generation
16a Method of generating the allocation sequence (eg, computer-
generated random numbers), and list of any factors for stratification.
To reduce predictability of a random sequence, details of any planned
restriction (eg, blocking) should be provided in a separate document
that is unavailable to those who enrol participants or assign
interventions
Allocation
concealment
mechanism
16b Mechanism of implementing the allocation sequence (eg, central
telephone; sequentially numbered, opaque, sealed envelopes),
describing any steps to conceal the sequence until interventions are
assigned
Implementation 16c Who will generate the allocation sequence, who will enrol participants,
and who will assign participants to interventions
Blinding
(masking)
17a Who will be blinded after assignment to interventions (eg, trial
participants, care providers, outcome assessors, data analysts), and
how
17b If blinded, circumstances under which unblinding is permissible, and
procedure for revealing a participant’s allocated intervention during
the trial
Methods: Data collection, management, and analysis
Data collection
methods
18a Plans for assessment and collection of outcome, baseline, and other
trial data, including any related processes to promote data quality (eg,
duplicate measurements, training of assessors) and a description of
study instruments (eg, questionnaires, laboratory tests) along with
their reliability and validity, if known. Reference to where data
collection forms can be found, if not in the protocol
18b Plans to promote participant retention and complete follow-up,
including list of any outcome data to be collected for participants who
discontinue or deviate from intervention protocols
Data
management
19 Plans for data entry, coding, security, and storage, including any
related processes to promote data quality (eg, double data entry;
range checks for data values). Reference to where details of data
management procedures can be found, if not in the protocol
Statistical
methods
20a Statistical methods for analysing primary and secondary outcomes.
Reference to where other details of the statistical analysis plan can be
found, if not in the protocol
20b Methods for any additional analyses (eg, subgroup and adjusted
analyses)
20c Definition of analysis population relating to protocol non-adherence
(eg, as randomised analysis), and any statistical methods to handle
missing data (eg, multiple imputation)
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Methods: Monitoring
Data monitoring 21a Composition of data monitoring committee (DMC); summary of its role
and reporting structure; statement of whether it is independent from
the sponsor and competing interests; and reference to where further
details about its charter can be found, if not in the protocol.
Alternatively, an explanation of why a DMC is not needed
21b Description of any interim analyses and stopping guidelines, including
who will have access to these interim results and make the final
decision to terminate the trial
Harms 22 Plans for collecting, assessing, reporting, and managing solicited and
spontaneously reported adverse events and other unintended effects
of trial interventions or trial conduct
Auditing 23 Frequency and procedures for auditing trial conduct, if any, and
whether the process will be independent from investigators and the
sponsor
Ethics and dissemination
Research ethics
approval
24 Plans for seeking research ethics committee/institutional review board
(REC/IRB) approval (point 2.3 page 9)
Protocol
amendments
25 Plans for communicating important protocol modifications (eg,
changes to eligibility criteria, outcomes, analyses) to relevant parties
(eg, investigators, REC/IRBs, trial participants, trial registries, journals,
regulators)
Consent or assent 26a Who will obtain informed consent or assent from potential trial
participants or authorised surrogates, and how (see Item 32)
26b Additional consent provisions for collection and use of participant data
and biological specimens in ancillary studies, if applicable
Confidentiality 27 How personal information about potential and enrolled participants will
be collected, shared, and maintained in order to protect confidentiality
before, during, and after the trial
Declaration of
interests
28 Financial and other competing interests for principal investigators for
the overall trial and each study site
Access to data 29 Statement of who will have access to the final trial dataset, and
disclosure of contractual agreements that limit such access for
investigators
Ancillary and
post-trial care
30 Provisions, if any, for ancillary and post-trial care, and for
compensation to those who suffer harm from trial participation
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Dissemination
policy
31a Plans for investigators and sponsor to communicate trial results to
participants, healthcare professionals, the public, and other relevant
groups (eg, via publication, reporting in results databases, or other
data sharing arrangements), including any publication restrictions
31b Authorship eligibility guidelines and any intended use of professional
writers
31c Plans, if any, for granting public access to the full protocol, participant-
level dataset, and statistical code
Appendices
Informed consent
materials
32 Model consent form and other related documentation given to
participants and authorised surrogates
Biological
specimens
33 Plans for collection, laboratory evaluation, and storage of biological
specimens for genetic or molecular analysis in the current trial and for
future use in ancillary studies, if applicable
*It is strongly recommended that this checklist be read in conjunction with the SPIRIT 2013
Explanation & Elaboration for important clarification on the items. Amendments to the
protocol should be tracked and dated. The SPIRIT checklist is copyrighted by the SPIRIT
Group under the Creative Commons “Attribution-NonCommercial-NoDerivs 3.0 Unported”
license.
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Recommended