Transcript (AIMSS): Identification of genetic predictors and

Aromatase Inhibitor-induced musculoskeletal Symptoms (AIMSS):
Identification of genetic predictors and causative mechanisms
Jason Robarge M.S.1, Todd Skaar Ph.D.1,2, Djane Duarte Ph.D.1, Michael Vasko Ph.D.1, David Flockhart M.D. Ph.D.1,2
of Pharmacology and Toxicology, 2Department of Medicine, Division of Clinical Pharmacology
Results: Our preliminary studies indicate letrozole reduces nociceptive thresholds to
mechanical stimuli. Experiments are underway to determine the dose-dependence of this
effect, the effect of acute versus chronic dosing, and the influence of genetic background on
the response.
INTRODUCTION
Third generation aromatase inhibitors (AIs) - anastrozole (Arimidex), letrozole (Femara), and
exemestane (Aromasin) - are commonly used for the treatment of hormone receptor-positive
breast cancer. Although they are highly effective, their side-effects often limit their usefulness;
one of the most significant is the musculoskeletal arthralgia. The arthralgia developed on AIs
presents clinically as non-inflammatory regional musculoskeletal disorder and has been
termed “aromatase inhibitor-induced musculoskeletal symptoms” or AIMSS. Patients often
complain of stiff or painful joints, morning stiffness, and tendonitis.
AIMSS are frequent, and develop soon after initiating therapy. An analysis conducted by the
COnsortium on BReast cancer phArmacogenomics (COBRA) of breast cancer patients
receiving daily exemestane or letrozole showed an incidence of 45.4%; a rate now
corroborated by others.1,2 The median time to onset was 1.6 months (range 0.4 – 10 months).
13% of the patients to discontinue therapy because of the AIMSS.1
Unfortunately, AIMSS also appear to be associated with improved survival; consequently, our
inability to manage the AIMSS means that breast cancer patients are often not able to take a
drug that would help them survive longer.3 Therefore, understanding the mechanisms
contributing to or predictors of AIMSS may help us to improve drug tolerability in those patients
who develop symptoms, potentially improving their long term prognosis.
While AIMSS may limit tolerance to AIs in some patients, there are no clear predictors of who
may get the symptoms, and the causative mechanism remains unknown. The mechanism
seems to be a result of the estrogen deprivation; however, the specific target is unknown. One
hint may come from previous in-vitro and rodent studies that have implicated estrogen as a
neuroendocrine modulator of pain processing; its action is thought to occur via a number of
mechanisms within the CNS and in peripheral primary afferent neurons. The presence of
aromatase and estrogen receptor immunoreactivity in neurons suggests that estrogens may be
produced locally and act to influence pain pathways via autocrine or paracrine signaling.4,5,6
Therefore, we hypothesized that AI-induced depletion of neuronal estrogen synthesis and
activity cause altered pain sensitivity. As an initial step to explore the mechanism by which AIs
may alter pain pathways, we are developing a rodent model to test the effect of aromatase
inhibition on nociception. To simulate the physiological conditions of AI therapy in postmenopausal women, we treated ovariectomized (OVX) rats with letrozole. The primary
outcome for these experiments was the AI induced change in nociceptive threshold for
mechanical and thermal stimuli.
Mean Response Threshold (g)
Methods: To generate a model to explore causative mechanisms underlying AIMSS, we have
utilized behavioral pain models in rodents. Using ovariectomized Sprague Dawley rats as a
model for the post-menopausal state, we have characterized acute nociceptive responses
following administration of letrozole, an aromatase inhibitor.
Figure2 A & B. Time course of mechanical hyperalgesia
Animals
Behavioral experiments were performed on female Sprague-Dawley rats (Harlan Laboratories,
USA). At approximately 150g, rats were bilaterally ovariectomized by Harlan, shipped, and
used following 2 weeks of recovery. Animals were housed in a controlled environment in the
Animal Laboratory Resource Center of the Indiana University School of Medicine. Food and
water were available ad libitum. Experimental protocols were approved by the Indiana
University School of Medicine, Institutional Animal Care and Use Committee.
Administration of letrozole
Letrozole was dissolved in hydroxypropyl-β-cyclodextrin (HPβCD) (10% in sterile PBS).
Letrozole was administered to treatment group rats (N=6) as a single intraperitoneal injection
(1.0 ml/kg) at a dose of 1.0 mg/kg. Control rats (N=5) received a single intraperitoneal
injection (1ml/kg) of vehicle (10% HPβCD).
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Behavioral Experiments
Baseline mechanical and thermal nociceptive thresholds were measured 2 days prior to drug
administration. The measurement taken 24 hours prior to drug was used as the baseline
response. Thresholds were re-assessed 30min following administration to detect acute
changes in response. Thresholds were measured again at 3, 5, 8, 12, and 21 days.
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A. Withdrawal latencies to a mechanical stimulus (von Frey hair) to the plantar surface of
the paw. Responses were measured at baseline and following I.P. administration of
letrozole (N=5animals/10 paws) or vehicle (N=5animals/10 paws). Each point represents
mean latency response ± SEM. Treatment group response was not significantly different at
baseline, while overall mean change in response was different between groups (p=0.0427).
At 8 days, the response compared to baseline was significantly lower in letrozole treated
animals (Tukey’s HSD post hoc analysis, p=0.00732**).
von Frey filaments
Mechanical hypernociception was evaluated using the von Frey filaments (Stoelting Co.).
Before each test, the animals were acclimated in the test cage for approximately 30 min. The
hind paw plantar surface was touched with one of a series of filaments with logarithmically
incremental stiffness (0.6–26g). A single trial consisted of three applications of a particular
filament, applied once every 3–4s. A response was defined as withdrawal of the stimulated
paw. The up–down method was used to record the threshold.
B. Responses are represented as percent change from baseline. Each point represents
mean ± SEM.
Hargreaves test
Paw withdrawal latency to radiant heat was assessed using a infra-red heat source (Ugo
Basile, Italy). Before each test, the animals were acclimated in the test cage for approximately
30 min. The I.R. source was placed underneath the mid-plantar surface of the hind paw. The
intensity of the heat source was chosen (30) to yield baseline latencies ranging from 8s to 10s
in non-OVX animals of equivalent weight, and a cut-off of 30s was used to avoid tissue
damage at the paw. A paw withdrawal in response to heat was detected by a photocell, which
switched off the I.R. source and timer. The paw withdrawal latency was taken to be the mean
of three trials from both hind paws, with at least 10s in-between.
CONCLUSIONS AND FUTURE DIRECTIONS
Letrozole reduced the nociceptive threshold of rats to a mechanical stimulus. However, it did
not alter the basal sensitivity to thermal stimulus.
These preliminary studies indicate that letrozole sensitizes rats to mechanical stimuli. To better
understand these results, our future studies will focus on the direct effects of letrozole on
neuron function. These experimental models should provide insights into the mechanisms of
letrozole induced musculoskeletal pain in breast cancer patients.
Data analysis
Group means and change from baseline were analyzed using repeated measures ANOVA.
Data analysis was performed using R, v2.7.1.
RESULTS
REFERENCES
Figure1 A & B. Time course of thermal hyperalgesia
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Mean paw-withdrawal
latency (sec)
Background: Aromatase inhibitor-induced musculoskeletal symptoms (AIMSS) are a limiting
toxicity developed during breast cancer therapy with aromatase inhibitors. Poorly understood,
these arthralgias are associated with improved response, while negatively impacting patient
quality of life and drug compliance. We hypothesize that aromatase inhibitor therapy alters
basal pain thresholds by inhibiting peripheral aromatase and reducing 17β-estradiol signaling
that modulates nociception.
RESULTS
METHODS
% Change in Threshold (g)
ABSTRACT
1. Henry, N.L. et al. Prospective characterization of musculoskeletal symptoms in early stage
breast cancer patients treated with aromatase inhibitors. Breast Cancer Res Treat. 111(2): 36572, 2007.
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Letrozole
6
Vehicle
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2
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Days from injection
20
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% Change in latency (sec)
1Department
2. Crew, K.D. et al. Prevalence of joint symptoms in postmenopausal women taking aromatase
inhibitors for early-stage breast cancer. J Clin Oncol. 25(25): 3877-83, 2007.
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20
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Letrozole
Vehicle
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-30
-5
0
5
10
15
20
Days from injection
A. Withdrawal latencies to a thermal nociceptive stimulus applied to the plantar surface of
the paw. Response were measured at baseline and following I.P. administration of
letrozole (N=6 animals/12 paws) or vehicle (N=5 animals/10 paws). Each point represents
mean latency response ± SEM. There were no significant differences between treatment
means nor significant changes from baseline in either treatment group.
B. Responses are represented as percent change from baseline. Each point represents
mean ± SEM.
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3. Cuzick, J., Sestak, I., Cella, D. & Fallowfield, L. Treatment-emergent endocrine symptoms
and the risk of breast cancer recurrence: a retrospective analysis of the ATAC trial. Lancet
Oncol. 9(12):1143-8, 2008.
4. Evrard HC, Harada N, Balthazart J. Localization of estrogen-synthase (aromatase) in the rat
spinal cord. Soc Neurosci Abstract. 27: 508.6, 2001.
5. Evrard, H.C. and Erskine, M.S. Spinal estrogen synthesis alters nociception-related
behaviors in male rat. Soc Neurosci Abstract. 746.16, 2005.
6. Evrard HC. Estrogen synthesis in the spinal dorsal horn: a new central mechanism for the
hormonal regulation of pain. Am J Physiol Regul Integr Comp Physiol. 291(2):R291-9, 2006.