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Clinical Trials |
The Taussig Cancer Center [R. v. S., J. C., D. M., R. G., T. B.], Departments of Gastroenterology [R. v. S., K. P.], Biostatistics and Epidemiology [P. E., L. R.], and Colorectal Surgery [J. C.], Cleveland Clinic Foundation, Cleveland, Ohio 44195; Colleges of Medicine and Public Health [G. S., K. C., B. D., L. K., B. H. K.] and Pharmacy [G. S., W. L. H., K. C.], The Ohio State University, Columbus, Ohio 43210; and National Cancer Institute, Bethesda, Maryland [E. H., G. K.]
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ABSTRACT |
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INTRODUCTION |
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Considerable epidemiological evidence suggests that nonsteroidal anti-inflammatory drugs can reduce the incidence of colorectal cancer (5, 6, 7) . In addition, the nonsteroidal anti-inflammatory drug sulindac has been shown to cause regression of colonic and rectal polyps in patients with FAP (8) . However, sulindac and other currently approved nonsteroidal anti-inflammatory drugs are associated with irritation of the upper gastrointestinal tract. This toxicity is attributed to the inhibition of COX1, whereas it is postulated that the cancer chemopreventive activity of these agents is attributable to the inhibition of the inducible enzyme COX2 (9) . Although relatively specific COX2 inhibitors are under investigation, there is evidence that the chemopreventive activity of nonsteroidal anti-inflammatory drugs is not solely attributable to COX inhibition. In Min/+ mice, which are heterozygous for a germ-line mutation in the murine APC gene, sulindac can decrease the incidence of intes- tinal adenomas. However, results are mixed with respect to the correlation of this activity with an alteration in prostaglandin E2 or leukotriene B4 levels in intestinal tissue (10 , 11) . Among the two major metabolites of sulindac is exisulind (also known as sulindac sulfone or FGN-1). This compound does not inhibit either COX1 or COX2 in preclinical systems (12 , 13) . Yet, it exerts chemopreventive activity in chemically induced rodent colon and breast cancer (12 , 14) , suggesting the existence of a novel mechanism of action unrelated to the inhibition of COX1 or COX2. On the basis of these preclinical observations, we performed a Phase I trial of exisulind in patients with FAP. The objectives of our study were to assess the safety and tolerability of exisulind, to determine the maximum safe dose of this agent in patients with FAP who had previously undergone subtotal colectomy with ileorectal anastamosis, and to examine the effects of treatment with exisulind on the natural history of FAP in this patient population. This report details the safety and pharmacokinetic results of this study as well as the effects of exisulind therapy on polyp numbers, histology, cellular proliferation, and apoptosis.
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PATIENTS AND METHODS |
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18 years of age, to have at
least five rectal polyps at the time of study entry, and to give
informed consent to participation. Fertile women were required to have
a negative pregnancy test and to be nonlactating, and all fertile
participants were required to use adequate contraception during the
course of the study. Patients were not allowed to take nonsteroidal
anti-inflammatory drugs for 2 weeks prior to study entry, and the use
of such drugs during the course of the study was prohibited. Patients
with known hypersensitivities to nonsteroidal anti-inflammatory drugs
were excluded from participation, as were patients who had been
documented to be resistant to a course of therapy with sulindac.
Potential participants underwent pre-study upper endoscopy, and
subjects with active peptic ulcer disease were excluded from further
participation. Also excluded from participation were patients who had
gastrointestinal problems that were felt by the investigators to be
likely to interfere with absorption of the study drug or assessment of
toxicity, patients in whom a proctectomy was planned prior to
completion of participation in the study, and patients who had used
another investigational medication within 1 month of study entry. FAP
patients who had hepatic or renal dysfunction that was significant in
the opinion of the investigators or that was associated with an
elevation of the aspartate aminotransferase or alanine aminotransferase
to >40 units/l were excluded, as were patients with a hemoglobin value
of <10 g/dl, a platelet count of <100 x
109/liter, or with any laboratory abnormality of
grade 2 or worse, based upon the Common Toxicity Criteria (version 1.0)
of the National Cancer Institute. Patients with a prior history of
malignancy, with the exception of nonmelanoma skin cancer, were
excluded.
Conduct of the Clinical Trial
Prior to study entry, informed consent to participate was
obtained, and potential participants underwent eligibility
screen- ing and baseline studies, including a medical and
surgical history, physical examination, flexible sigmoidoscopy,
esophagogastroduodenoscopy, serum pregnancy test (if appropriate),
serum chemistries, urinalysis, bleeding time, and complete blood count
with differential. Eligible patients were entered in cohorts of six
into successively higher dose levels according to an empirically
designed dose-escalation scheme. The dose of exisulind was never
escalated in an individual patient to a dose higher than the starting
dose. The planned dose levels were 200 and 400 mg p.o. bid and higher,
but dose escalation was halted after dose-limiting toxicity was noted
at the 400-mg p.o. bid dose level. An additional, intermediate dose
level of 300 mg p.o. bid was then added; six patients were treated at
this dose level. A minimum of 4 months of treatment was necessary for a
patient to be considered evaluable, unless dose-limiting toxicity was
experienced. Escalation to the next higher dose level occurred only
after all patients treated at all lower dose levels had been treated
for a minimum of 2 weeks and fewer than three of the six subjects in
each dose level had experienced grade 2 or worse toxicity according to
the Common Toxicity Criteria. The occurrence of grade 2 or worse
toxicity in three or more patients in a given dose level was considered
unacceptably toxic, defining the maximum safe dose as the next lower
dose level.
During the initial week of therapy, participants were contacted daily on weekdays by a study nurse to assess adverse events, record concomitant medications, and assess drug compliance. These contacts were repeated weekly for the first 8 weeks and monthly thereafter. Overnight fasting blood draws for a serum biochemistry profile, complete blood count with differential, and bleeding time were performed at weeks 1 and 2 and monthly thereafter. Urinalyses were performed at weeks 1 and 4 and then monthly. All participants underwent physical examination, Simplate bleeding time determination, and sigmoidoscopy after 1, 4, and 6 months of treatment.
On the first day of treatment, patients were administered a single dose of exisulind after an overnight fast, and detailed pharmacokinetic studies were performed. Thereafter, patients were dosed twice daily for 6 months. On the final day of treatment, the morning dose was taken after an overnight fast, and detailed pharmacokinetics were again performed; the second dose of exisulind was withheld on the final day of treatment to allow prolonged sampling for pharmacokinetic studies. For the detailed pharmacokinetic studies performed after the initial dose of exisulind, blood samples were drawn at time 0 (pre-dose) and again 0.25, 0.50, 1.0, 1.5, 2.0, 3.0, 4.0, 6.0, 8.0, 10.0, 12.0, 22.0, and 24.0 h postdose. A 24-h urine collection for pharmacokinetic studies and creatinine clearance was collected during this time. Patients were served a breakfast similar to one that they eat normally after the 1-h sample was drawn. A similar schedule was followed for the detailed pharmacokinetic studies performed after the final dose of exisulind, except that additional blood samples were drawn 36.0 and 48.0 h after the final dose. In addition, "trough" plasma samples and 24-h urine collections were assayed for exisulind prior to the morning dose after 1, 2, and 4 weeks of therapy and monthly thereafter.
Upper endoscopy was performed prior to study entry and after the completion of 6 months of therapy. All participants underwent physical examination and sigmoidoscopy prior to the initiation of therapy and after 1, 4, and 6 months of treatment. At each sigmoidoscopy, the polyps in each rectal segment were counted. At the time of each sigmoidoscopy, four biopsies were taken from normal-appearing mucosa from each rectal segment. In addition, multiple polyps were biopsied at the time of each sigmoidoscopic procedure. All biopsies of polyp and mucosal tissue were fixed in neutral-buffered formalin and examined histologically after H&E staining. In addition, all biopsy specimens were analyzed with respect to proliferation index, as determined by Ki-67 expression, and apoptotic index, as determined by cell morphology and TUNEL assay.
Analytic Methodology for Assaying Exisulind
The concentration of exisulind in plasma and urine was determined
by high-pressure liquid chromatography using a methodology described
previously (15)
. The analyte exisulind and the added
internal standard, indomethacin, were extracted using methylene
chloride. After separation and evaporation of the methylene chloride,
the residue was dissolved in mobile phase and analyzed by reversed
phase high-pressure liquid chromatography. A 5-µm, 15 x 0.46-cm
Octyl column was used to provide excellent chromatographic separation
with high sensitivity for the analyte. The analyte and internal
standard were measured by UV absorbance detection at 329 nm. The
standard curve used the peak height ratio of the analyte to that of the
internal standard. Samples were analyzed in duplicate, and duplicate
values were averaged. If the difference of the duplicate values
relative to their mean was >10%, the sample was reanalyzed. The limit
of detection in plasma was 1 ng/ml, and the standard curve was linear
up to 10,000 ng/ml. Validation studies were performed to determine the
accuracy and reproducibility of the assay. In addition, plasma samples
containing known concentrations of exisulind were shipped on dry ice
from the Ohio State University to the Cleveland Clinic and back, stored
for 3 months, and subsequently assayed to determine the stability of
exisulind handled under the same conditions as the clinical specimens.
In other experiments, samples were assayed after being maintained at
refrigerator temperature (2°C) for 2–6 days or at room temperature
(
25°C) for 1 day to examine potential sources of error in the
clinical specimens.
The assay was validated in plasma prior to the initiation of the study. The within-day coefficients of variation were 2.3, 1.9, and 1.7% at concentrations of 0.081, 12.0, and 24.1 µM, respectively, with corresponding accuracy values of 113, 102, and 101% (n = 8). The corresponding between-day coefficients of variation were 7.0, 3.8, and 2.0%. The quality control standard indicated <3% variation when assayed after 3 months as described above.
Pharmacokinetic and Pharmacodynamic Analyses
The plasma concentration-time profile after the first dose was
analyzed using the WinNonlin pharmacokinetic modeling software package
(16)
. The noncompartmental analysis was performed on each
individual profile after the first dose. This analysis was selected
because the multiple peaks that were observed in several patients,
apparently because of enterohepatic recirculation, precluded
compartmental analysis. In addition, average plasma concentration
profiles, which did not reflect the multiple peaks observed in the
individual profiles, were analyzed using a two-compartment model with
first-order absorption and a lag time before absorption began. The
weighting function was Y-2. The mean and SD of
each mean was calculated using a standard spreadsheet software program
(17)
. The influences of gender and body size were examined
using the Multiple General Linear Hypothesis-General Linear Model
section of the SYSTAT statistical analysis program (18)
.
The relationship between toxicity, dose, and pharmacokinetic parameters
was examined using the LogXact statistical analysis program for exact
logistic regression (19)
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Proliferation Index (Ki-67 Expression)
Ki-67 Staining.
The 4-µm sections on slides were deparaffinized, and antigen
retrieval was obtained by microwaving the slides in 0.01 M
citrate buffer (pH 6) for 10 min. Endogenous peroxidase was blocked
with 3% hydrogen peroxide for 15 min. Specific binding was blocked by
incubating the slides with 1% nonfat dried milk for 30 min. The slides
were incubated with a 1:100 dilution of MIB1 primary antibody
(Immunotech, Marseille, France) for 1 h in a humidified chamber at
room temperature. Normal mouse immunoglobulin G (Vector Laboratories,
Burlington, CA) was used as a secondary antibody in a 1:100 dilution
for 30 min. Immunoperoxidase detection was achieved with the
avidin-biotin conjugate method (Vector Laboratories), followed by the
diaminobenzidine (Vector Laboratories) enhancement reaction. Counter
staining was performed with Harris hematoxylin. A negative control was
prepared on each slide by omission of the primary antibody.
Imaging.
The percentage of Ki-67-positive nuclear area per total nuclear area
was determined on one slide of normal tissue and three slides, if
available, of dysplastic tissue from each patient. In the dysplastic
tissue, Ki-67 levels were determined in 10 normal-appearing glands
and 10 dysplastic glands. In normal tissue, Ki-67 levels were
determined in up to 10 normal-appearing glands. The Roche image
analysis software (Roche, Elon College, NC) was used for quantification
of Ki-67 levels, which were obtained by plotting the Ki-67-positive
nuclear area against the total nuclear area and expressing the positive
Ki-67 area as a percentage of the total. Stained slides were viewed
under a bright-field Zeiss Axioskop microscope with a x20 objective.
The level of negative and positive staining was determined by grayscale
threshold.
Apoptotic Index (TUNEL Assay)
The apoptotic indices of adenomas and normal-appearing mucosa were
determined in slides adjacent to the slides in which proliferative
indices were determined. Apoptotic cells in adenomas and
normal-appearing mucosa were identified by TUNEL labeling using an
adaptation of the methodology described by Gavrieli et al.
(20)
. The staining protocol used with the adenomas and
normal mucosa was found to give results similar to a method previously
reported for the detection for apoptotic cells (21
, 22)
.
Formalin-fixed tissue sections were cut on to Probe-On slides (Fisher),
deparaffinized, and rehydrated in graded alcohol. Sections were treated
for 30 min with 0.5% pepsin (1:2500 strength; Sigma Chemical Co., St.
Louis, MO) in 0.1 N HCl to digest protein, and
then washed in H2O (4 x 2 min), treated
with 2% H2O2 in PBS to
quench endogenous peroxidase, and washed (2 x 5 min) in PBS.
Samples were then covered with freshly prepared reaction buffer (126 ml
equilibrium buffer, 8.6 ml CoCl2 solution (0.037
g of CoCl2 in 10 ml of
H2O), 5.7 ml dUTP (dUTP solution, 6 parts
stock:100 parts H2O; Boehringer-Mannheim,
Indianapolis IN), 59 ml of PBS (Sigma), and 1.4 ml of TdT enzyme for a
final concentration of 0.1 enzyme unit/ml (TdT from Life Technologies,
Inc., Gaithersburg, MD). Slides were placed in a humid box inside a
37°C oven for 60 min and then transferred to stop wash (300
mM NaCl and 30 mM sodium
citrate) in 37°C agitating water bath for 30 min. After washing with
PBS (3 x 5 min), the sections were covered with 1:20 diluted
anti-digoxigenin peroxidase (Boehringer-Mannheim) for 30 min. Samples
were washed with PBS (3 x 5 min) and then placed in an ambient
temperature diaminobenzidine solution of one tablet Sigma
DABfast/H2O2 in 150 ml of
PBS (Sigma) for 9 min to develop the reaction product, washed under
running tap water, counterstained with hematoxylin, rinsed in tap
water, dehydrated through graded alcohols, xylene, and mounted with
Permount. For dysplastic tissue, the section was divided into
equal-size quadrants, and 250 epithelial cells/quadrant were counted
(or all of the cells, if <250 were present). For normal-appearing
mucosa, 10 crypts (20 crypt columns) were counted, with all epithelial
cells from the crypt base to five nuclei past the top of the column
crypt being tallied. Cells were scored as apoptotic only if they
displayed morphological features of apoptosis (i.e.,
condensed or segmented nuclei), regardless of whether they labeled
positive with the TUNEL method. The apoptotic index was calculated by
dividing the total number of apoptotic cells by the number of
epithelial cells counted and expressing the quotient as a percentage.
Adenoma specimens were confirmed for dysplasia by histology.
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RESULTS |
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1 month of
treatment when surgery was necessitated by the diagnosis of high-grade
dysplasia of the ampulla of Vater, based upon evaluation of the
pretreatment upper endoscopy specimens. The remaining 18 participants
completed the prescribed 6 months of therapy. Six additional subjects
were screened but were found to be ineligible. Table 1
summarizes the baseline characteristics
of the 18 fully evaluable participants.
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Safety and Toxicity.
Table 2
summarizes the most
frequently reported adverse events. Exisulind was generally well
tolerated, and only one symptomatic adverse event considered to be
"severe" (elevated alanine aminotransferase) was reported. Four of
six patients in each dose level experienced headaches. Gastrointestinal
toxicities were reported by all patients treated at the 300- and 400-mg
bid dose levels and by two patients treated at the 200-mg bid dose
level. These complaints consisted of nausea or vomiting, diarrhea,
changes in the frequency or consistency of bowel movements, abdominal
pain, or dyspepsia. These complaints were generally mild in degree and
did not require cessation of the study drug. Dose-limiting hepatic
toxicity was noted at a dose of exisulind of 400 mg p.o. bid (see Table 2
). When dose-limiting hepatic toxicity was noted at the 400-mg bid
dose level, all patients at that dose level were dose reduced. Dose
re-escalation was attempted in patients not suffering toxicity. Fig. 1
summarizes the relationship between the
liver function tests and the temporal course, severity, and the
administered dose for the six evaluable patients treated at the 400-mg
bid dose level. As can be seen, liver function abnormalities reversed
rapidly after treatment with exisulind was withheld or the dose
reduced. Reinstitution of therapy at a lower dose was well tolerated,
and all evaluable patients were able to complete 6 months of therapy
with exisulind.
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. Because of dose modifications for
toxicity, only two of the six patients starting therapy at 400 mg p.o.
bid were receiving the intended dose after 6 months of therapy; the
curve for the 6-month time point reflects the mean of these two
patients. The overall plasma concentration-time profiles of exisulind
at 6 months are higher than those after the first dose, indicating some
accumulation of the drug. Because exisulind has a half-life that is <8
h in most patients, 1.5 maintenance doses would accumulate at steady
state with a dosing interval of 12 h. The log-linear phases
(generally 10 h and beyond) appeared parallel, consistent with a
lack of change in the elimination of exisulind during the course of the
study. There was a systematic trend toward a decrease in clearance and
volume of distribution after 6 months (Table 3)
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summarizes the pharmacokinetic parameters for each dose level,
using a model-independent analysis of the plasma concentration-time
profiles. The Tmax values (time of
peak concentration) were 2 h after drug administration and were
similar for the three dosage levels studied. The
Cmax values (maximum plasma
concentration) increased proportionally with dose for the 200- and
400-mg doses, but the Cmax value for
the 300-mg dose was higher than expected. The
t1/2 was 6–7 h for the two lower
dosages but was somewhat shorter for the 400-mg dose, although
considerable interpatient variability was noted. The AUC also showed
considerable interpatient variability. As with the
Cmax values, the AUC values seemed to
display a nonlinear relationship to dose (Fig. 2)
. The apparent volume
of distribution (Vß and
Vss) was slightly less than 1 l/kg,
consistent with distribution throughout total body water. The total
body clearance (CLs) was 8–10 l/h.
This relatively small value compared with blood flow to the organs of
elimination does not suggest a significant first pass elimination of
exisulind. There was no dose dependence in the
CLs,
Vß, and Vss
values after the first dose, and all of these parameters showed
considerable variability among patients. This variability was not
significantly reduced by normalizing the parameter values to body
weight, lean body weight, or to body surface area (data not shown).
A two-compartment model analysis of the average plasma
concentration-time profiles was performed at baseline and after 6
months (Table 4)
. The parameters that are
common to those in Table 3
(the noncompartmental analysis) have similar
values. The absorption half-life (t1/2,
abs) was 0.7 h. The lag time, the period after dose
administration during which there is no absorption, was 20–40 min.
These values are consistent with a lag time attributable to gastric
emptying and capsule disintegration, followed by rapid absorption.
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results for the 400-mg dose level are not shown because the actual
administered dose varied from patient to patient and over time as a
result of dose modifications for toxicity at that dose level. The
trough plasma concentrations observed over the 6-month course of the
study were relatively constant over time for each dose level.
Substantial interpatient variability in these values was present,
however. Barely detectable levels were found on several occasions for
patient 1001, consistent with the 60% compliance documented by capsule
counts. Overall, the observed trough concentrations were consistent
with those expected based upon the first-dose pharmacokinetics,
i.e., the expected average concentration would be dose rate
divided by total body clearance.
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Pharmacodynamics.
Adjusting for dose, the occurrence of hepatic toxicity was unrelated to
any of the pharmacokinetic parameters summarized in Table 3
(P 0.28 for all parameters).
Polyp Numbers and Morphology.
Fig. 3
summarizes the mean number of
polyps observed over time, accounting for polyps that were excised.
Data from one patient in the 400-mg p.o. bid dose group were incomplete
and are not included. Polyp counts were not statistically different
between any of the dose groups at baseline or after 1 month of therapy.
However, polyp numbers rose significantly after 4 and 6 months of
therapy at the 200-mg p.o. bid dose level (P < 0.001).
Polyp numbers were not significantly different from at baseline after 4
and 6 months of therapy at the 300- and 400-mg p.o. bid dose levels
(also referred to as the 400/200-mg dose level because all patients
were dose reduced during the course of therapy).
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. We have used the term "halo
polyps" to describe these lesions. Because these polyps were an
unanticipated finding, they were not counted prospectively during the
course of the trial. Halo polyps were occasionally seen prior to
therapy with exisulind but were more common after therapy. These polyps
have been found to represent adenomas when biopsied.
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Apoptotic Index of Intestinal Crypt Cells in Normal-appearing
Mucosa and Adenomatous Polyps.
Fig. 5
summarizes the apoptotic index, as
determined by the TUNEL assay, of the intestinal crypt cells in
normal-appearing mucosa and in adenomatous polyps. Both prior to and
after therapy, the apoptotic index was greater in polyps than in
normal-appearing mucosa (P < 0.001 at both baseline
and month 6, Wilcoxon Signed Rank Test). Overall, the apoptotic index
was not significantly different after 6 months of therapy than at
baseline in either normal-appearing mucosa or in adenomas, although
there appeared to be a trend for the apoptotic index of the polyps to
increase after therapy (P = 0.09, Wilcoxon-Signed Rank
Test). This appeared particularly true at the 300-mg bid dose level,
but the number of patients treated at each dose level was too small to
allow any conclusion to be drawn with respect to dose and response, and
there was no statistically significant dose effect (P =
0.14, Jonckheere-Terpstra Test).
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. The degree of this
mucinous differentiation was subjectively graded as "none,"
"mild," "moderate," or "extensive." Overall, mucinous
differentiation was noted in the polyps of only 4 of 15 patients with
evaluable polyps at baseline, and the degree of this differentiation
was termed "mild" in all cases. After 6 months of therapy, some
degree of mucinous differentiation was observed in 16 of 17 patients
from whom polyps were available. The degree of this change was termed
"mild" in 4 cases, "moderate" in 7 cases, and "extensive"
in 5 patients.
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DISCUSSION |
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23
knockout mice are a murine model of FAP in which a truncating mutation
of the APC gene results in gastrointestinal polyp formation
(23)
. In APC23 mice in whom COX-2
has also been knocked-out, the rate of adenoma formation is greatly
attenuated, further suggesting that COX-2 is important to the process
of adenoma formation (24)
. In other colon cancer model systems, however, inhibition of COX-2 has not been found to be the mechanism of action of NSAIDs and related drugs. The COX-2 inhibitor NS-398 induced apoptosis in both the HT29 colon carcinoma cell line, in which COX-2 is expressed constitutively, and in the S/KS cell line, in which COX-2 protein is undetectable (25) . Chiu et al. (11) did not find COX-2 to be overexpressed in the normal-appearing mucosa of Min/+ mice and dietary supplementation with arachidonic acid did not increase tumor formation. The observation that the sulfone metabolite of sulindac, exisulind, does not inhibit COX-2 (12 , 13) but inhibits azoxymethane-induced colon carcinogenesis (14) has been interpreted as consistent with the hypothesis that exisulind, and perhaps NSAIDs, have a chemopreventive mechanism independent of COX-2 inhibition. In vitro studies have indicated that exisulind could induce apoptosis independent of COX-2 inhibition, and that this effect was not reversed by the prostaglandin analogue dimethyl-prostaglandin E2 (13) . The current trial was undertaken on the basis of the hypothesis that exisulind treatment would induce apoptosis in rectal polyps of patients with FAP and result in a reduction in polyp numbers.
This trial demonstrates that the dose-limiting toxicity of exisulind in patients with FAP is reversible hepatic dysfunction. The MTD of exisulind that can be given to this patient population is 300 mg p.o. bid, or a total daily dose of 600 mg/day. Over the 6-month treatment course, no long-term toxicities were noted. Patients completing the current study have been entered on a longer term extension trial to further evaluate safety and efficacy. Preliminary data from this trial suggest that long-term therapy at the MTD can be administered with acceptable safety, although the study remains ongoing. Clinical trials in patients with sporadic polyps or other lower risk populations might be considered after long-term safety has been demonstrated in patients with FAP.
The pharmacokinetics of exisulind in patients with FAP who have undergone colectomy are nonlinear over the dosage range of 200–400 mg p.o. bid. The pharmacokinetics are not time dependent over the 6-month course of the study, and no gender-associated differences are apparent. The volume of distribution of exisulind is consistent with distribution throughout total body water, with clearance studies suggesting low extraction. Pharmacodynamic studies show no clear relationship between pharmacokinetic parameters and hepatic toxicity, with dose being the most important predictor of toxicity. Pharmacokinetic studies indicate that exisulind is absorbed rapidly and, likely, completely. Although large interpatient differences are noted, in general, the half-life is in the range of 6–9 h in FAP patients.
These pharmacological findings should be related to those reported for normal volunteers with intact colons and to the concentrations of sulindac sulfone used in preclinical studies. At the 200- and 300-mg single doses of exisulind, major pharmacokinetic parameters of exisulind are similar in these patients of both sexes with FAP who have undergone colectomy and ileorectal anastomosis and those reported previously for normal healthy male volunteers (Ref. 15 ; Cmax 200 = 4.91 mg/l for FAP patients versus 6.67 in normal healthy male volunteers; AUC200 = 27.96 mg x h/l in FAP patients versus 32.25 in normal healthy male volunteers; Cmax300 = 11.08 mg/l in FAP patients versus 7.85 in normal healthy male volunteers; AUC300 = 49.21 mg x h/l in FAP patients versus 40.49 in normal healthy male volunteers). At the 400-mg dose level, however, the Cmax in patients with FAP was lower than in normal male volunteers (9.85 versus 13.14 mg/l), as was the AUC (48.7 versus 85.04 mg x h/l). Future studies in patients with FAP who have undergone subtotal colectomy should investigate a daily dose of exisulind of 600 mg/day. The increased reabsorption of exisulind and the resultant prolonged mean residence time in patients with intact colons may necessitate a reduction in the total daily dose in patients with intact colons (15) .
At the maximum safe dose of 300 mg p.o. bid, steady-state plasma concentrations in the range of 1–5 mg/l (2.7–13.4 µM) and peak plasma concentrations of 8.5–11.1 mg/l (23–30 µM) were achieved. These concentrations are lower than the IC50 of exisulind for various cell lines (90–200 µM; Refs. 26 and 27 ) or the concentration of exisulind reported to induce apoptosis in HT-29 colon or MCF-7 breast carcinoma cells (240 µM; Refs. 26 and 27 ). The achieved plasma concentrations of exisulind are, however, in the range of the concentration that has been reported to increase the expression of APC mRNA in malignant colonic epithelial cells (10 µM; Ref. 28 ) and to inhibit 7,12-dimethylbenz(a)anthracene-induced mammary lesions in organ culture studies (10 µM; Ref. 27 ).
At no dose did we observe a reduction in overall polyp numbers, although the numbers of polyps remained stable during the 6-month treatment period for patients treated at the MTD of 300 mg p.o. bid. This might represent a chance selection of patients with relatively indolent disease at this dose level or might represent a perturbation of the generally progressive natural history of the disease. Treatment with exisulind did not affect cellular proliferation in either normal-appearing mucosa or in polyps, consistent with preclinical observations. No statistically significant change in apoptotic rate was noted, but a nonsignificant trend toward an increased apoptotic index after 6 months of therapy as compared with baseline was noted. The most interesting observations were those related to the gross and microscopic morphology of polyps in patients treated with exisulind. There was an increase in the number of polyps with a "halo" appearance and an increase in mucinous differentiation of polyps after therapy. Polyps showing these changes, however, remained adenomatous.
These observations suggest that exisulind may have biological effects in patients with FAP, but that these effects are insufficient to produce a reduction in polyp numbers. These findings are similar to those reported for sulindac sulfone in preclinical studies in FAP model systems. Exisulind has proven inactive in two studies in the Min/+ mouse model of FAP (29 , 30) , despite showing activity in the azoxymethane rat colon carcinogenesis system. Thus, although reversible hepatic toxicity prevented the achievement of plasma concentrations of exisulind in the range found to be biologically active in the majority of the preclinical investigations of this agent, further explication of the biological effects of exisulind and its congeners may result in the development of clinically useful chemopreventive and therapeutic approaches.
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FOOTNOTES |
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1 Supported by National Cancer Institute Contract
NO1-55119 
2 To whom requests for reprints should be
addressed, at The Cleveland Clinic Foundation, Desk T-40, 9500 Euclid
Avenue, Cleveland, OH 44195. Phone: (216) 444-6480; Fax:
(216) 444-9464; E-mail: buddg{at}ccf.org
3 The abbreviations used are: FAP, familial
adenomatous polyposis; COX, cyclooxygenase; bid, twice a day; TUNEL,
terminal deoxynucleotidyl transferase-mediated nick end labeling; AUC,
area under the plasma concentration-time curve, extrapolated to time
infinity; MTD, maximum tolerated dose.
Received 10/26/98; revised 6/30/99; accepted 8/17/99.
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TopABSTRACTINTRODUCTIONPATIENTS AND METHODSRESULTSDISCUSSIONREFERENCES |
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716 knockout mice by inhibition of cyclooxygenase 2 (COX-2). Cell, 87: 803-809, 1996.[CrossRef][Medline]
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) and after 6 months (
);
bars, SE. 200 mg: each point represents
the mean of six patients; 300 mg: each point represents
the mean of seven patients after the first dose and six patients after
6 months; 400 mg: each point represents the mean of two
patients, and SEs are not shown.
