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-DEOXY-3 -[18F]FLUOROTHYMIDINE PET/CT FOR EVALUATING...
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USE OF 30-DEOXY-30-[18F]FLUOROTHYMIDINE PET/CT FOR EVALUATING
RESPONSE TO CYTOTOXIC CHEMOTHERAPY IN DOGS WITH
NON-HODGKIN’S LYMPHOMA
JESSICA LAWRENCE, MATTHEW VANDERHOEK, DAVID BARBEE, ROBERT JERAJ, DANIEL B. TUMAS, DAVID M. VAIL
Imaging and measurement of proliferation with computed tomography (CT) and positron emission tomography
(PET) provide a noninvasive method for improved staging and monitoring of response to cancer treatment. We
evaluated prospectively the proliferation marker 30-deoxy-30[18F] fluorothymidine (FLT) in the context of FLT-
PET/CT for detection of early response, confirmation of posttreatment response, and prediction of relapse in
dogs with non-Hodgkin’s lymphoma. Nine dogs with non-Hodgkin’s lymphoma who were scheduled to receive
five cycles of an investigational cytotoxic chemotherapy agent were included. All dogs received baseline FLT-
PET/CT imaging immediately before chemotherapy. Intent was to repeat imaging with FLT-PET/CT at
various time points: group 1 (n¼ 4), 5 days after initiation of chemotherapy and 3 weeks following the last
chemotherapy administration; group 2 (n¼ 5), before the fourth cycle of chemotherapy and 3 weeks following
the last administration. Two dogs in group 2 did not undergo repeat PET/CT. Body mass standardized uptake
values (SUV) for FLT were calculated for each dog. Eight dogs had initially increased FLT uptake (mean
SUVmax¼ 9.8 [2.6–22.3]). Mean SUV decreased significantly for the seven dogs that underwent follow-up
PET/CT following chemotherapy (mean SUVmax¼ 3.5 [1.1–7.9], Po0.016). Increased uptake preceded clin-
ical and cytological evidence of relapse in two dogs. Ki-67 immunohistochemistry confirmed decreased pro-
liferation corresponding to decreased SUV in three canine lymph node samples. FLT-PET/CT functional and
anatomical imaging shows promise for the evaluation of response to cytotoxic chemotherapy in dogs with non-
Hodgkin’s lymphoma and for predicting relapse before standard clinical and clinicopathologic confirmation.
Veterinary Radiology & Ultrasound, Vol. 50, No. 6, 2009, pp 660–668.
Key words: 30-deoxy-30[18F] fluorothymidine, canine, non-Hodgkin’s lymphoma, positron emission
tomography.
Introduction
CANINE NON-HODGKIN’S LYMPHOMA is a spontaneous,
rapidly proliferative neoplasia that is similar in bio-
logic behavior to high-grade non-Hodgkin’s lymphoma in
humans. Standard therapy for lymphoma involves multi-
agent chemotherapy administered systemically. Initial
treatment of lymphoma is often rewarding, however, re-
lapse is common, and only 25% of dogs survive 2 years
or longer with standard protocols.1 Relapse is most often
detected by reenlargement of lymph nodes or return of
clinical signs, at which point significant tumor volume has
developed. Early detection of relapse permits earlier inter-
vention, which may result in longer response duration and
improved outcome. Methods of detecting early response
and relapse are of interest in the management of canine
lymphoma, among other tumors. In addition, accurate
monitoring of therapy is extremely important as new cyto-
toxic or molecular agents are investigated.
There is a tremendous interest in molecular imaging
for the diagnosis, staging, and evaluation of response
for individuals with cancer. Modalities such as photon
emission computed tomography (SPECT), positron emis-
sion tomography (PET), magnetic resonance imaging,
and computed tomography (CT) allow precise anatomic
localization and offer a unique method of analyzing
tissue differences in response to therapy. Nuclear medicine
imaging methods such as PET use tracers to target
specific mechanisms that typically differ between normal
and cancer cells, often preceding changes in anatomic
structure identified with conventional imaging.2–10 Func-
tional imaging has garnered much attention due to its po-
tential ability to detect early response to therapy and
differentiation of tumor from inflammation, necrosis, or
fibrosis.2,11–14
Funding: Provided by Gilead Sciences Inc.Material presented in part: Veterinary Cancer Society 2007 (Ft. Lau-
derdale, FL) and American College of Veterinary Radiology 2008 (SanAntonio, TX).Address correspondence and reprint requests to David M. Vail, at the
above address. E-mail: [email protected] May 17, 2009; accepted for publication June 24, 2009.doi: 10.1111/j.1740-8261.2009.01612.x
From the School of Veterinary Medicine (Lawrence), Department ofMedical Physics (Vanderhoek, Barbee, Jeraj), Paul P Carbone Compre-hensive Cancer Center (Jeraj, Vail), University of Wisconsin-Madison,2015 Linden Drive, Madison, WI 53706, and the Department of DrugSafety Evaluation, Gilead Sciences Inc., 333 Lakeside Drive, Foster City,CA 94404 (Tumas).
660
PET using the glucose analogue 2-[18F]fluoro-2-deoxy-D-
glucose (FDG) is the most commonly utilized imaging
metabolic biomarker.15 FDG significantly enhances the
sensitivity of detection of lymphoma and residual uptake is
an accurate and independent predictor of progression-free
survival.16–22 Consequently, FDG-PET is now considered
a standard component to the routine workup of human
lymphoma patients.15,23,24 FDG-PET imaging has recently
been evaluated in canine lymphoma and mast cell tumor
and was useful in staging both diseases.25
FDG is not tumor specific, however, and can also ac-
cumulate in inflammatory tissues, which can confound on-
cologic imaging.26–28 Much attention has been directed
toward other tracers that can increase the specificity for
malignant lesions to complement the information currently
obtained with FDG functional studies. For example, pro-
liferative activity was more specific for malignant tumors
when compared with an increase of glucose metabolism.29
Measurement of tumor growth and DNA synthesis in vivo
may therefore be more appropriate for staging and assess-
ment of response to therapy for neoplasia.
The thymidine-analogue 30-deoxy-30[18F]fluorothymi-
dine (FLT) is a PET tracer that accumulates in prolifer-
ating tissues, including malignant tumors.30 FLT appears
to accurately reflect DNA synthesis, is taken up via passive
diffusion as well as facilitated transport and is subsequently
phosphorylated by thymidine kinase 1 (TK1) and trapped
intracellularly as [18F]FLT-monophosphate.13 FLT is not
incorporated into DNA, but rather acts as a chain termi-
nator due to the absence of 30-hydroxyl necessary for in-
tegration in the existing chain.12,31 TK1 activity is minimal
in quiescent cells whereas it may be three- to fourfold
higher in malignant cells.32,33 In addition, tumor cells may
harbor mutations in the carboxyl terminus of TK1, dis-
rupting normal degradation at mitosis, thereby leading to
deregulation of enzyme activity.34 Alterations in TK1 reg-
ulation and increased activity render FLT an attractive
molecular tracer for detection of malignant cells. In hu-
mans, physiologic FLT uptake occurs in bone marrow,
liver and gallbladder, and the urinary tract. In contrast to
FDG-PET, however, no uptake is evident in the brain,
skeletal muscles, or myocardium. In humans there is sig-
nificant correlation between tumor proliferation measured
using Ki-67 immunohistochemistry and FLT uptake in
lymphoma and other solid tumors.35–37 Also, FLT-PET
imaging has promise for early evaluation of remission sta-
tus in human non-Hodgkin’s lymphoma patients treated
with standard chemotherapy regimens.38 We theorized that
similar results could be obtained in dogs with non-Hodg-
kin’s lymphoma. In this study, we report a pilot study of
nine dogs with non-Hodgkin’s lymphoma that were eval-
uated with FLT-PET/CT imaging. Our hypotheses were
threefold: that FLT-PET/CT would be useful in dogs with
lymphoma for detection of early response to chemother-
apy, for confirmation of response following chemotherapy
completion, and for prediction of relapse.
Materials and Methods
Nine dogs were evaluated prospectively as part of
a clinical trial evaluating the efficacy of a novel nucleoside
analogue chemotherapy agent, GS-9219.39,40 Exclusion
criteria were dogs assessed as clinical substage b, dogs
weighing o10kg, concurrent cytotoxic therapy, and pres-
ence of concurrent illnesses that might affect drug tolerance
or short-term survival. Dogs were of various breeds, in-
cluding Golden Retriever (n¼ 2), Beagle (n¼ 1), Grey-
hound (n¼ 1), German Shepherd (n¼ 1), Bernese
Mountain Dog (n¼ 1), Bassett Hound (n¼ 1), Dachshund
(n¼ 1), and Hound Cross (n¼ 1). The median age was
8 years. Four dogs had naı̈ve lymphoma, four had relapsed
following previous treatment with a standard cyclophosph-
amide–doxorubicin–vincristine–prednisone (CHOP) con-
taining chemotherapy protocol and one dog had relapsed
following prednisone therapy. Three dogs were classified as
stage III non-Hodgkin’s lymphoma, four with stage IV
non-Hodgkin’s lymphoma, and two with stage V non-
Hodgkin’s lymphoma. Two dogs had T-cell lymphoma
while the remaining dogs had B-cell lymphoma following
immunochemical analysis.
A baseline FLT-PET/CT examination was performed
within 24h preceding the first administration of chemo-
therapy. FLT-PET/CT imaging was planned to be re-
peated at day 5 (n¼ 4; group 1) or before the fourth dose of
chemotherapy (n¼ 5; group 2). All dogs were scheduled to
undergo a third FLT-PET/CT scan 3 weeks following
completion of chemotherapy. Six of the nine dogs com-
pleted the protocol as outlined (Table 1).
Chemotherapy was administered intravenously at vari-
ous intervals, as determined by randomization at enroll-
ment, for a total of 15 weeks of planned therapy.40 Dogs
were evaluated with bloodwork, physical examination, and
lymph node aspirates at each visit. Clinical remission status
was documented for each dog using the most current cri-
teria applied to humans with lymphoma that predates
the inclusion of PET/CT assessment (international work-
shop to standardize response criteria for non-Hodgkin’s
lymphoma).41
FLT was synthesized as described42 and obtained from
the cyclotron and radiopharmaceutical laboratory at the
Department of Medical Physics at the University of Wis-
consin-Madison. Dogs were sedated and subsequently
anesthetized with propofol induction and isoflurane main-
tenance. Dogs were in sternal recumbency in the PET/CT
gantry and imaging was performed with a clinical GE
Discovery LS PET/CT scanner.� Noncontrast whole body
�General Electric, Waukesha, WI.
661CANINE LYMPHOMA MONITORING WITH FLT-PET/CTVol. 50, No. 6
Table1.ClinicalOutcomeandFLT-PET/C
TResponse
Dog
Diagnosis
Stage
Planned
PET/C
TSchedule
(Group)
Baseline
FLT-PET/C
TScan
Clinical
Response
toChem
o-Therapy
Day5
FLT-PET/C
TResponse:
Day5
Clinical
Response
toChem
o-Therapy
ThirdCycle
FLT-PET/C
TResponse:
ThirdCycle
Clinical
Response
Postchem
o-
therap
y
FLT-PET/C
TResponse
Post-FLT
Outcomefrom
Initial
PET/C
T
1B-cellLSA
StageIIIa
2Low
uptakein
lymphnodes
(LN)
CR
NA
CR
Nouptake
CR
Nouptake
Relap
seat9months
2B-cellLSA
StageIV
a2
Widespread
LN
uptake
CR
NA
CR
Nouptake
CR
Nouptake
Relap
seat9months
3B-cellLSA
StageIIIa
1Widespread
LN
uptake
PR
CR
CR
NA
CR
HighuptakeR
tonsilan
dR
retropharyngeal
LN
Relap
seat6months
4T-cellLSA
StageIIIa
2Widespread
uptake
PR
NA
NA
NA
NA
NA
Euthanized
at18days
dueto
lymphoma
5T-cellLSA
StageIV
a2
Widespread
LN
uptake
NA
NA
NA
NA
NA
NA
Acute
tetraparesis
2daysafter
treatm
ent.Died
3daysafter
treatm
ent;
widespread
lymphomapresent
atnecropsy
6B-cellLSA
StageIV
a1
Widespread
LN
uptake
CR
Decreased
uptake
CR
NA
CR
Highuptakeleft
mandibularand
retro-pharyngeal
LN
Outofremissionat
4.5
months
cytologically
(1month
post
final
PET/C
T)
7B-cellLSA
StageVa
1Widespread
LN
uptake
PR
Decreased
uptake
CR
NA
CR
Nouptake
Outofremissionat
4.75months
(3weekspost
final
PET/C
T)
8B-cellLSA
StageVa
2Widespread
LN
uptake
PR
NA
CR
Nouptake
CR
Nouptake
Outofremissionat
6.8
months
9B-cellLSA
StageIV
a1
Widespread
LN
uptake
SD
Widespread
uptake
NA
NA
NA
NA
Progressivelymphoma
at15days
CT,computedtomography;FLT,fluorothym
idine;
PET,positronem
issiontomography.
662 LAWRENCE ET AL. 2009
spiral CT scans were obtained, followed immediately by
FLT-PET imaging which was automatically coregistered to
the CT images. Acquisition of the whole body CT was
obtained in approximatley 50–60 s with a multislice helical
technique. As the table moved forward from CT to PET
acquisition, table height and cantilever point were con-
stant. Both scans were acquired using the same cranial and
caudal borders, field of view, and slice thickness, thus per-
mitting automatic coregistration of the images by the
hardware. Whole body images were acquired over six to
eight bed positions (205–273 slices) with 10-min acquisi-
tions per bed position 50–60min following injection of ap-
proximately 220MBq FLT (range, 185–260MBq). The
PET scanner contains 12,096 individual crystals arranged
into 18 rings of 672 crystals, and permits the simultaneous
acquisition of 35 transaxial PET emission images over an
axial field of view (FOV) with a 4.25mm slice thickness
(matrix size 128 � 128 � 35).43 The PET scanner permitted
evaluation of large breed dogs with imaging FOV of
50.0 cm and bore size of 60.0 cm. The total system sensi-
tivity for true events is 223kcps/(microCi/cc) with septa in
and 1200kcps/(microCi/cc) with septa out.43 The GE Dis-
covery LS PET scanner contains two rotating 68Ge rod
sources for transmission scanning at a speed of 20 rpm.
Each rod is 15.3mm long and 4.0mm in diameter over an
axial FOV of 15.3 cm, with a maximum activity of
370MBq per rod and a total activity 740MBq. Emission
data were corrected for attenuation, and subsequently re-
constructed using an iterative ordered-subset expectation
maximization (OSEM) algorithm. Image pixel counts were
calibrated to activity concentrations (Becquerel/ ml) and
decay corrected using the scan start time as a reference
point.
All PET/CT images were assessed by two individuals.
Lymph nodes were contoured on all CT images and cir-
cular regions of interest with a diameter of 0.5 cm were
placed within each lymph node in the area with highest
FLT uptake, as described previously.38,43 The maximum
body mass standardized uptake values (SUVmax) used in
analysis were calculated from each region of interest using
the formula
SUV ¼ Imaged activity concentration ðBq=gÞ � bodyweight ðgÞInjected activity ðBqÞ
For definition of regions of interest and data analysis,
AMIRA software was used.w Statistical analyses were per-
formed using a commercial statistical software package.zNonparametric two-tailed paired T tests were done to
compare maximum SUV values at various time-points.
Differences were considered significant in all analyses if a
P value o0.05 was obtained.
Ki-67 immunohistochemistry was performed using stan-
dard techniques on tumor tissues in three dogs in group 1
by a commercial laboratoryy using a rabbit Ki-67mono-
clonal antibody zpreviously validated in the dog. Popliteal
or prescapular lymph node samples were obtained from
anesthetized dogs several hours before the PET/CT scan.
Lymph node biopsies were performed the same day as the
PET/CT scans immediately before the initial cytotoxic
treatment and subsequently 5 days later. Lymph nodes
from normal beagle dogs removed before and following
treatment with GS-9219 were used as positive controls.39
Briefly, paraffin imbedded tissues were deparaffinized, air
dried out of alcohol, and antigens retrieved using heat-
induced steam. Slides underwent protein/enzymes blockade
followed by incubation with antibody, then developed and
counterstained with hematoxylin.
Results
Increased baseline FLT uptake was documented in the
lymph nodes of eight of nine dogs. One dog had low FLT
uptake before, during and following chemotherapy. Two
dogs in group 2 died shortly after the baseline scan was
performed, thus subsequent scans were not performed.
Mean SUVmax before chemotherapy for all dogs was 9.8
(range, 2.6–22.3) and 3.5 following chemotherapy (range,
1.1–7.9). These differences were statistically significant
(Po0.016) (Fig. 1).
FLT-PET/CT images indicated evidence of early re-
sponse to chemotherapy in both group 1 and group 2 dogs.
One dog in group 1 was considered to be in complete clinical
remission on the basis of physical examination and lymph
node aspirates at the time of the second FLT-PET/CT
scan (day 5), two dogs were considered to have achieved
partial remission, and the final dog had stable disease as
defined by International Workship response criteria. In
one of the dogs achieving partial remission at day 5, a
Fig. 1. Mean maximum body mass standardized uptake value (SUVmax)for seven dogs in which prechemotherapy and postchemotherapy flu-orothymidine (FLT)-positron emission tomography (PET)/computed to-mography (CT) scans were performed. Mean SUVmax before chemotherapyfor all dogs was significantly higher (9.8; range 2.6–22.3) compared withmean SUVmax following cytotoxic chemotherapy (3.5; 1.1–7.9; Po0.016).
wAMIRA 4.1.1, Visage Imaging Inc., Carlsbad, CA.zPrism 4.0b; GraphPad Software, San Diego, CA.
yIHCtech, Aurora, CO.zClone SP6, Lab Vision, Fremont, CA.
663CANINE LYMPHOMA MONITORING WITH FLT-PET/CTVol. 50, No. 6
firm left prescapular lymph node was noted on physical
examination that was cytologically consistent with lym-
phoma on fine needle aspirate. FLT-PET/CT confirmed
persistent increased FLT uptake within the left prescapular
lymph node, however, other lymph nodes displayed low
tracer uptake, corroborating the physical examination
findings. PET/CT scan indicated decreased FLT uptake
in the peripheral nodes compared with baseline, although
increased SUV remained present compared with surround-
ing tissue. The dog classified as having stable disease had
palpably smaller lymph nodes that did not as yet meet
International Workship criteria for a response; FLT-PET/
CT scan indicated decreased FLT uptake in peripheral
lymph nodes, although the lymph nodes remained enlarged
on physical exam and CT images. This dog developed
progressive lymphoma and was euthanized 2 weeks later.
One dog achieved complete remission in group 1 and had
low FLT uptake on FLT-PET/CT scans 3 weeks follow-
ing completion of chemotherapy (Fig. 2A and B). This
dog was in partial remission on day 5 based on enlarged
lymph nodes and presence of lymphoma on cytological
evaluation.
Three dogs in group 2 were considered to be in complete
clinical remission based on physical examination and
lymph node aspirates. FLT-PET/CT scans performed be-
fore fourth cycle of chemotherapy were characterized by
low FLT uptake compared with initial scans. The two re-
maining dogs in group 2 died before subsequent scans.
One dog was euthanized 3 days following the initial FLT-
PET/CT scan as a result of diffuse swelling and degener-
ation of the caudal spinal cord and widespread lymphoma.
The second dog was euthanized due to progressive lym-
phoma 3 weeks following the initial FLT-PET/CT scan.
The three dogs that were in complete remission from their
lymphoma at the first repeat scan remained in complete
remission following completion of chemotherapy. FLT-
PET/CT scans indicated minimal lymph node uptake in
these dogs, which corroborated examination and lymph
node aspirate findings.
FLT-PET/CT scans were successful in detecting early
lymphoma recrudescence in two dogs, both in group 1.
One dog was in complete remission based on physical ex-
amination, lymph node aspirate, and FLT-PET/CT scan
on day 5 following initiation of chemotherapy. This dog
was considered to remain in remission at recheck evalua-
tion 3 weeks following all chemotherapy cycles based on
physical examination. Mandibular lymph node aspirate
was consistent cytologically with a normal lymph node.
Fig. 2. (A) Left—fluorothymidine (FLT)-positron emission tomography (PET)/computed tomography (CT) image of a 3-year-old dog illustrating FLTuptake in the peripheral nodes, bone marrow, kidneys bladder, and spleen. (B) Right—FLT-PET/CT image of the same dog 3 weeks following the final dose ofchemotherapy. The lymph nodes were small on CT images with minimal FLT uptake on PET images. Note the persistent uptake in the bone marrow, kidneys,and bladder.
664 LAWRENCE ET AL. 2009
However, FLT-PET/CT scan indicated increased FLT up-
take within the right tonsil and right retropharyngeal
lymph node (Fig. 3A and B). Fine needle aspirate of the
right tonsil was performed following the scan and was cy-
tologically consistent with lymphoma.
The second dog in group 1 was in partial remission
based on physical examination characteristics at reevalu-
ation on day 5 following initiation of chemotherapy. Fine
needle aspirate of his left mandibular lymph node was
consistent cytologically with reactive lymphoid hyperpla-
sia. FLT-PET/CT findings were consistent with reduced
FLT uptake compared with baseline scan. The lymph
nodes became smaller over time and the dog was consid-
ered to have achieved complete remission within 3 weeks of
starting cytotoxic therapy. Three weeks following comple-
tion of chemotherapy, the dog was considered on physical
examination to be in complete remission. It was difficult to
obtain representative samples of lymph node to confirm
remission cytologically due to their small size. However, on
the PET/CT scan there was marked increase in FLT up-
take within the left mandibular lymph node. Aspirates of
the left mandibular lymph node were obtained under an-
esthesia but cytologically were not diagnostic of lym-
phoma. The dog was monitored and subsequently came
out of remission in the left mandibular lymph node within
30 days of the FLT-PET/CT scan.
Ki-67 IHC on lymph node tissue from normal beagle
dogs was characterized by typical proliferative activity in
the germinal centers of lymph node follicles before treat-
ment (Fig. 4A) and significant suppression of proliferation
5 days following cytotoxic chemotherapy (Fig. 4B). Similar
decrease in proliferative activity, as measured by Ki-67,
was also found in pet dogs with non-Hodgkin’s lymphoma
before and after cytotoxic chemotherapy (Fig. 4C and D)
at similar time periods. The decrease in Ki-67 labeling
fraction mimicked the reduction in FLT-PET/CT SUV at
similar time periods of analysis.
Discussion
Based on this trial, our hypotheses that FLT-PET/CT is
useful in dogs with lymphoma for detection of early re-
sponse and confirmation of response to chemotherapy,
were correct. Importantly, FLT-PET/CT was also able to
predict early lymphoma recrudescence in two patients be-
fore clinical detection. These features could result in real
time modification of chemotherapy protocols, timing and
intensity, ultimately translating into improvements in dis-
ease-free interval and overall survival in dogs with non-
Hodgkin’s lymphoma.
There are few published data regarding clinical use of
PET/CT scanning in staging or treatment monitoring of
Fig. 3. (A) Left—fluorothymidine (FLT)-positron emission tomography (PET)/computed tomography (CT) image of a 10-year-old dog with lymphoma,illustrating lack of FLT uptake in the peripheral lymph nodes 5 days following initiation of chemotherapy. There is increased FLT uptake surrounding theoriginal intravenous catheter site in the left saphenous region. (B) Right—FLT-PET/CT image of the same dog, illustrating increased FLT uptake within theright tonsil and right retropharyngeal lymph node. Note the lack of FLT uptake in the area in (A); cytologically, the tonsil was consistent with lymphoma.
665CANINE LYMPHOMA MONITORING WITH FLT-PET/CTVol. 50, No. 6
dogs with cancer. 18FDG-PET imaging was used in dogs
with lymphoma and with mast cell tumor.25 With regard to
the dogs with non-Hodgkin’s lymphoma in that report,18FDG-PET imaging was helpful for staging and assessing
response to therapy.25 In the dog with small cell lymphoma
in the prior report, persistent increased 18FDG uptake was
observed in one lymphocenter during chemotherapy, how-
ever, differentiation of residual disease from inflammation
or lymphoid hyperplasia was not possible.25 Clearly, it
would be of benefit to distinguish reactive lymphadenopathy
from malignant disease.
Our results suggest that FLT-PET is highly sensitive in
detecting lymphoma, as previously shown in humans and
rodents.36,38 Two dogs with relapsed lymphoma were iden-
tified on FLT-PET/CT before detection of recrudescence
on physical examination. Interestingly, the dog with lym-
phoma relapse within the right tonsil and right retropha-
ryngeal lymph node was cytologically free of lymphoma in
the right mandibular lymph node. This highlights the abil-
ity of lymphoma to relapse in a locoregional manner,
which may be missed on routine examination. Lymphoma
within peripheral lymph nodes was detected in all patients,
similar to human patients with lymphoma.38 In dogs, pe-
ripheral lymph node enlargement is the most common sign
of relapse, and this study indicates that FLT-PET/CT
imaging may provide earlier detection. Theoretically, ear-
lier detection would allow quicker reinstitution of cytotoxic
therapy, which may favorably affect remission duration
with rescue protocols. It may also aid in the detection of
dogs that are not in remission following induction chemo-
therapy, allowing earlier commencement to alternative,
more effective protocols.
It is interesting to note that one dog in our study did not
have appreciable FLT uptake within his lymph nodes on
any PET/CT scan. This dog still had decreased mean
lymph node SUV postchemotherapy compared with pre-
chemotherapy (mean SUV 11.0 vs. 3.1). This biopsy sam-
ple from this dog was not available for review. However,
the clinical presentation was characteristic of high-grade
malignant lymphoma, with rapidly increasing peripheral
lymphadenopathy and thrombocytopenia, presumably im-
mune mediated. The dog responded to chemotherapy
within 14 days of administration, suggesting a proliferative
neoplasia. The reason for the poor FLT uptake is unclear.
However, a similar experience has been found in a human
with anaplastic large cell lymphoma, in which FLT-PET
scan indicated low FLT uptake within the tumor despite
high tumor cell Ki-67 positivity.36 Review of this patient’s
tumor indicated marked fibrosis that accounted for430%
of the mass and it was concluded that the low cellularity
Fig. 4. Ki-67 Immunohistochemistry of lymphoid tissues and lymphoma. (A) Normal peripheral lymph node from a beagle dog 5 days after receivingplacebo drug vehicle. Note the significant Ki-67 immunoreactivity of cells in the highly proliferative germinal center (� 100) characterized by amber staining.(B) Normal peripheral lymph node from a beagle dog 5 days after receiving GS-9219 cytotoxic chemotherapy. Note the lack of Ki-67 immunoreactivity anddepletion of cells in the germinal center (� 100). Lymphoma tissue from the prescapular lymph node of a dog in study before initiation of GS-9219 cytotoxicchemotherapy (C) compared with the contralateral prescapular lymph node in the same dog biopsied 4 days following GS-9219 cytotoxic chemotherapy (D)(� 600). Note the significant decrease in Ki-67 immunoreactivity following therapy indicating an antiproliferative effect.
666 LAWRENCE ET AL. 2009
and desmoplastic reaction secondary to lymphoma may
have explained the low FLT uptake.36 Additionally, mu-
tations or alterations in TK1 activity in an individual can-
cer could result in decreased FLT trapping.
The antiproliferative effect of chemotherapy docu-
mented by FLT-PET/CT in our study correlates with Ki-
67 immunoreactivity, an immunohistochemical measure of
proliferation; a correlation also observed in humans with
high-grade non-Hodgkin’s lymphoma.35,36
The use of FLT-PET/CT is not limited to detection of
lymphoma and may be able to more accurately identify
lesion localization for solid tumors. Response assessment
via cellular proliferation has been evaluated in humans with
adenocarcinoma, high-grade sarcoma, small cell lung can-
cer, and breast carcinoma.44–46 In human patients with pri-
mary and metastatic breast cancer, changes in FLT uptake
after one course of chemotherapy were significantly corre-
lated to eventual changes in tumor marker levels.45 In vet-
erinary medicine, specific tumor markers are rarely used for
continued monitoring, thus subsequent PET/CT imaging
may offer a reliable method of tracking remission status.
It is unlikely that FLT-PET/CT will provide an advan-
tage over FDG-PET for staging purposes in lymphoma,
given that there is normally high FLT uptake in the bone
marrow of dogs. However, given the evaluation of larger
numbers of dogs, it is possible that a significant difference
in FLT uptake is detected between normal dogs and those
with bone marrow involvement. We evaluated a dog with
multiple myeloma via FLT-PET/CT imaging and found
the initial scan to show low FLT uptake in bone marrow.
Following a positive response to systemic chemotherapy,
the dog’s bone marrow displayed high FLT uptake, sug-
gesting a return to normal marrow function. We noted
elevated FLT uptake in the liver compared with fat (data
not shown), however, further study of a larger subset of
dogs will be required to definitively comment on the utility
of FLT-PET for detecting lymphoma infiltration within the
liver. FLT-PET/CT offers significant benefit over FDG-
PET in patients that have received chemotherapy or radi-
ation therapy, as it is less likely to accumulate in inflam-
matory cells that often occur following a substantial
amount of tumor cell death. It is likely that information
garnered from both FLT-PET and FDG-PET will be at
least in part complementary.
There are several limitations to our study. Only a
small number of dogs were evaluated, making it difficult to
evaluate differences between histologic subtypes of non-
Hodgkin’s lymphoma (B-cell vs. T-cell), stage at diagnosis,
or body size differences. The two dogs in this study with
T-cell non-Hodgkin’s lymphoma appeared to have extraor-
dinarily high FLT uptake compared with the dogs with B-
cell non-Hodgkin’s lymphoma, however, it was impossible
to draw conclusions with such small numbers. A consensus
does not currently exist as to the optimal method of cal-
culating SUV based on PET scans. Several different meth-
ods have been described, as well as the use of ratios to help
minimize variability among observers. Regardless, the
differences in FLT uptake here were significant when we
assessed individual lymph nodes or total lymph node vol-
ume, as mathematic models were consistent between scan-
ning points. The role of chemotherapy must be addressed
with respect to its affect on FLT uptake as well. The drug
used to treat the dogs that underwent FLT-PET/CT scan-
ning in this trial were treated with a nucleoside analogue,
which upregulates TK1 activities.47,48 In contrast, agents
such as cisplatin decrease FLT uptake.47 The role of che-
motherapy may not have a marked impact on overall re-
sponses identified on PET/CT, however, various agents are
likely to affect how quickly changes may be seen. Dogs
treated with an agent that increases TK1 activity may have
enhanced FLT uptake 24h postadministration of the drug,
which may not accurately reflect response of the tumor
cells to therapy. Optimal timing of sequential PET/CT
scanning will need to be determined with larger numbers of
dogs and are likely to depend on the underlying tumor
histology and the type of chemotherapy applied.
In conclusion, our data suggest that FLT-PET/CT in
dogs with lymphoma is useful for detection of disease
sites, determination of both early and late responses to
chemotherapy, and the recognition of disease recrudes-
cence. Further investigation should continue to evaluate
the utility of FLT-PET/CT in the response monitoring of
lymphoma as well as for lesion localization, staging and
detection of response, or recurrence for other forms of
canine cancer.
ACKNOWLEDGEMENTS
The authors are extremely grateful to MacKenzie Wessel, Drs. Ce-cilia Robat, Angela Kozicki, Timothy Stein, and Kristi Hall for theirassistance and dedication to the dogs in this trial. Special thanks areextended toward the owners of all dogs enrolled in the trial. This studywas supported by research funding from Gilead Sciences Inc. to D.M.V.D.B.T. is an employee of Gilead Sciences Inc.
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