Fluorothymidine Uptake Research Articles

MO-E-AUD C-03: Use of FLT-PET Imaging to Assess Tumor and Normal Tissue Response to Radiotherapy

Purpose: In radiation therapy, use of 2‐Fluoro‐2‐deoxy‐D‐glucose (FDG) PET imaging to assess treatment response in tumors and normal tissues is suboptimal because of the confounding effects of radiation‐induced inflammation. We have investigated the feasibility of 3′‐Deoxy‐3′‐fluorothymidine (FLT) PET imaging, as a surrogate of cell proliferation, to assess treatment response in tumors and as well as normal tissues.Method and Materials: Patients receiving radiation therapy were imaged twice with FLT‐PET/CT: prior to therapy, and after receiving approximately 10–20 Gy of radiationdose. A 90 minute dynamic FLT‐PET image acquisition was initiated after the injection of FLT. The average standardized uptake values (SUV) between 60–90 minutes were used in the analysis. The CT data between the imaging sessions was co‐registered and the corresponding PET data compared and analyzed. Both, the tumor region as well as the surrounding normal tissues within the radiation field were analyzed for their response. Results: The FLT‐PET SUVs decreased approximately 20–50% within the first two weeks of radiation therapy, which provides an optimal timing window for treatment response assessment. Of the normal tissues,radiation effects were most notable in bone marrow, because of the high normal FLT uptake. Radiationdoses in excess of 10 Gy lead to complete bone marrow ablation, as assessed with FLT‐PET imaging. For doses below 10 Gy, an exponential cell‐kill relation correlates well with the observed decrease in FLT bone marrow uptake. Radiation effects were observed in other normal tissues as well. However, due to the low baseline FLT uptake, the changes were not readily detectable. Conclusion: FLT‐PET imaging was demonstrated as a powerful tool for early assessment of proliferative response to radiation therapy.Treatment assessment is possible as early as one week after the initiation of therapy. In addition to tumor response, FLT‐PET imaging also provides means to assess normal tissue damage.

Predicting Treatment Response of Malignant Gliomas to Bevacizumab and Irinotecan by Imaging Proliferation With [18F] Fluorothymidine Positron Emission Tomography: A Pilot Study

Evaluation of treatment effects in malignant brain tumors is challenging because of the lack of reliable response predictors of tumor response. This study examines the predictive value of positron emission tomography (PET) using [18F] fluorothymidine (FLT), an imaging biomarker of cell proliferation, in patients with recurrent malignant gliomas treated with bevacizumab in combination with irinotecan. Patients with recurrent malignant gliomas treated with biweekly cycles of bevacizumab and irinotecan were prospectively studied with FLT-PET at baseline, after 1 to 2 weeks, and after 6 weeks from start of treatment. A more than 25% reduction in tumor FLT uptake as measured by standardized uptake value was defined as a metabolic response. FLT responses were compared with response as shown by magnetic resonance imaging (MRI) and patient survival. Twenty-one patients were included, and 19 were assessable for metabolic response evaluation with FLT-PET. There were nine responders (47%) and 10 nonresponders (53%). Metabolic responders survived three times as long as nonresponders (10.8 v 3.4 months; P = .003), and tended to have a prolonged progression-free survival (P = .061). Both early and later FLT-PET responses were more significant predictors of overall survival (1 to 2 weeks, P = .006; 6 weeks, P = .002), compared with the MRI responses (P = .060 for both 6-week and best responses). FLT-PET as an imaging biomarker seems to be predictive of overall survival in bevacizumab and irinotecan treatment of recurrent gliomas. Whether FLT-PET performed as early as 1 to 2 week after starting treatment is as predictive as the study indicates at 6 weeks warrants further investigation.

Application of the multicellular tumour spheroid model to screen PET tracers for analysis of early response of chemotherapy in breast cancer

IntroductionPositron emission tomography (PET) is suggested for early monitoring of treatment response, assuming that effective anticancer treatment induces metabolic changes that precede morphology alterations and changes in growth. The aim of this study was to introduce multicellular tumour spheroids (MTS) to study the effect of anticancer drugs and suggest an appropriate PET tracer for further studies.MethodsMTS of the breast cancer cell line MCF7 were exposed to doxorubicin, paclitaxel, docetaxel, tamoxifen or imatinib for 7 days for growth pattern studies and for 3 or 5 days for PET tracer studies. The effect on growth was computed using the semi-automated size determination method (SASDM). The effect on the uptake of PET tracers [18F]3'-deoxy-3'-fluorothymidine (FLT), [1-11C]acetate (ACE), [11C]choline (CHO), [11C]methionine (MET), and 2-[18F]fluoro-2-deoxyglucose (FDG) was calculated in form of uptake/viable volume of the MTS at the end of the drug exposures, and finally the uptake was related to effects on growth rate.ResultsThe drugs paclitaxel, docetaxel and doxorubicin gave severe growth inhibition, which correlated well with inhibition of the FLT uptake. FLT had, compared with ACE, CHO, MET and FDG, higher sensitivity in monitoring the therapy effects.ConclusionSASDM provides an effective, user-friendly, time-saving and accurate method to record the growth pattern of the MTS, and also to calculate the effect of the drug on PET tracer uptake. This study demonstrate the use of MTS and SASDM in combination with PET tracers as a promising approach to probe and select PET tracer for treatment monitoring of anticancer drugs and that can hopefully be applied for optimisation in breast cancer treatment.

Monitoring Response to Radiotherapy in Human Squamous Cell Cancer Bearing Nude Mice: Comparison of 2′-deoxy-2′-[18F]fluoro-d-glucose (FDG) and 3′-[18F]fluoro-3′-deoxythymidine (FLT)

ObjectiveThe uptake of 3′-[18F]fluoro-3′-deoxythymidine (FLT), a proliferation marker, was measured before and during fractionated radiotherapy to evaluate the potential of FLT-positron emission tomography (PET) imaging as an indicator of tumor response compared to 2′-deoxy-2′-[18F]fluoro-d-glucose (FDG).Materials and MethodsNude mice bearing established human head and neck xenografts (HNX-OE; nu/nu mice) were locally irradiated (three fractions/week; 22 Gy) using a 150-kVp unit. Multiple FDG- and FLT-PET scans were acquired during treatment. Tumor volume was determined regularly, and tissue was analyzed for biomarkers involved in tracer uptake.ResultsBoth groups revealed a significant decline in tumor volume (P < 0.01) compared to untreated tumors. For FDG as well as for FLT, a significant decline in retention was observed at day 4. For FLT, most significant decline in retention was observed at day 12; whereas, for FDG, this was already noted at day 4. Maximum decline in tumor-to-nontumor ratios (T/NT) for FDG and FLT was 42 ± 18% and 49 ± 16% (mean ± SD), respectively. FLT uptake was higher then that of FDG. For FLT, statistical significant correlations were found for both tumor volume at baseline and at day 29 with T/NT and ΔT/NT. All tumors demonstrated expression of glucose transporter-1, thymidine kinase-1, and hexokinase II. No differences were found for amount of tumor cells and necrosis at the end of treatment.ConclusionThis new experimental in vivo model supports the promise of using FLT-PET, as with FDG-PET, to monitor response to external radiotherapy. This warrants further clinical studies to compare these two tracers especially in cancers treated with radiotherapy.

PET imaging with [18F]3′-deoxy-3′-fluorothymidine for prediction of response to neoadjuvant treatment in patients with rectal cancer

Positron emission tomography (PET) using 18F-labelled 3'-deoxy-3'-fluorothymidine (FLT) was assessed for therapy monitoring in patients with rectal cancer undergoing neoadjuvant chemoradiotherapy. Ten patients with locally advanced rectal cancer were included and underwent long-course preoperative chemoradiotherapy (total dose 45 Gy, 1.8 Gy/day, concomitant 250 mg/m2 5-fluorouracil) followed by surgery. FLT-PET was performed prior to chemoradiotherapy, 2 weeks after initiation of chemoradiotherapy and preoperatively (3-4 weeks post chemoradiotherapy). FLT uptake was correlated with histopathological tumour regression and changes in T stage. Mean tumour FLT uptake was 4.2+/-1.0 SUV before therapy and decreased significantly to 2.9+/-0.6 SUV 14 days after initiation of chemoradiotherapy (-28.6%+/-10.7%, p=0.005). The preoperative scan showed a further decrease to 1.9+/-0.4 SUV (-54.7%+/-7.6%, p=0.005). However, the degree of change in FLT uptake 2 weeks after initiation and after completion of neoadjuvant therapy did not correlate with histopathological tumour regression. FLT-PET did not seem to be a promising method for assessment of tumour response in the studied chemoradiotherapy regimen in patients with rectal cancer.

Molecular Imaging of Proliferation in Malignant Lymphoma

We have determined the ability of positron emission tomography (PET) with the thymidine analogue 3'-deoxy-3'-[(18)F]fluorothymidine (FLT) to detect manifestation sites of malignant lymphoma, to assess proliferative activity, and to differentiate aggressive from indolent tumors. In this prospective study, FLT-PET was done additionally to routine staging procedures in 34 patients with malignant lymphoma. Sixty minutes after i.v. injection of approximately 330 MBq FLT, emission and transmission scanning was done. Tracer uptake in lymphoma was evaluated semiquantitatively by calculation of standardized uptake values (SUV) and correlated to tumor grading and proliferation fraction as determined by Ki-67 immunohistochemistry. FLT-PET detected a total of 490 lesions compared with 420 lesions revealed by routine staging. In 11 patients with indolent lymphoma, mean FLT-SUV in biopsied lesions was 2.3 (range, 1.2-4.5). In 21 patients with aggressive lymphoma, a significantly higher FLT uptake was observed (mean FLT-SUV, 5.9; range, 3.2-9.2; P < 0.0001) and a cutoff value of SUV = 3 accurately discriminated between indolent and aggressive lymphoma. Linear regression analysis indicated significant correlation of FLT uptake in biopsied lesions and proliferation fraction (r = 0.84; P < 0.0001). In this clinical study, FLT-PET was suitable for imaging malignant lymphoma and noninvasive assessment of tumor grading. Due to specific imaging of proliferation, FLT may be a superior PET tracer for detection of malignant lymphoma in organs with high physiologic fluorodeoxyglucose uptake and early detection of progression to a more aggressive histology or potential transformation.

3′-Deoxy-3′-[F-18]Fluorothymidine Positron Emission Tomography in Patients with Recurrent Glioblastoma Multiforme: Comparison with Gd-DTPA Enhanced Magnetic Resonance Imaging

The accumulation of 3'-deoxy-3'-[F-18]fluorothymidine (FLT) on positron emission tomography (PET) images in patients with glioblastoma multiforme was evaluated and correlated with gadopentetate dimeglumine (Gd-DTPA) enhancement in magnetic resonance images (MRIs). FLT studies in 10 patients with recurrent glioblastoma multiforme were retrospectively investigated. Dynamic emission data were acquired for 60 minutes immediately after injection of FLT. The standardized uptake value (SUV) for tumor and reference tissue (contralateral hemisphere and ipsilateral cerebellum) was calculated. The volumes of the metabolically active part of the tumor (V (PET)) and that of the Gd-DTPA enhancing part of the tumor (V (MR)) were calculated. FLT uptake in tumors peaked before 5 minutes and sometimes as early as 0.5 minutes, and reached a constant level at approximately 10 minutes after injection. The reference tissue time-activity curves had an early peak and reached a constant low background level. All tumors had increased FLT uptake and showed Gd-DTPA enhancement. The SUV in tumor was significantly higher than that in the reference tissue (P<0.0001). A significant correlation between V (PET) and V (MR) was found (P<0.0001) although there was a difference in the areas of Gd-DTPA enhancement and FLT uptake. These preliminary results indicate that FLT-PET may be useful for the detection of recurrent glioblastoma multiforme. Our data in a relatively small patient population do not support a clear-cut relationship between FLT accumulation and Gd-DTPA enhancement. Further pathologic correlation will determine if it can be used for detecting recurrent tumoral disease.

Use of 3′-deoxy-3′-[18F]fluorothymidine PET to monitor early responses to radiation therapy in murine SCCVII tumors

3'-Deoxy-3'-[(18)F]fluorothymidine (FLT) is a promising new radiopharmaceutical for imaging cell proliferation. We evaluated whether FLT PET can be used to monitor early responses to radiation treatment. C3H/HeN mice bearing murine squamous cell carcinomas were randomized to irradiation with 0, 10, or 20 Gy. Twenty-four hours later, the mice were sacrificed for histopathological and biological assessment such as cell cycle analysis, Hoechst staining, and clonogenic cell survival assay. PET scans were performed on other mice after injection of [(18)F]FLT or [(18)F]fluorodeoxyglucose (FDG) before and after radiation treatment, and tumor growth was assessed over 9 days. Histopathological examination detected no morphological changes 24 h after radiation treatment, but cell cycle analysis showed that irradiated tumors had a decreased fraction of cells in S phase and an increased fraction in G2-M phase, compared with nonirradiated tumors. Irradiated tumors also had a higher incidence of apoptotic features and reduced clonogenic cell survival. Tumor growth was significantly delayed in irradiated mice (p<0.001) compared with control mice. PET images showed increased tumoral uptake of both FLT and FDG before radiation treatment. Following irradiation, FLT uptake differed significantly (p=0.020) from that in control mice. In contrast, FDG uptake after irradiation did not differ significantly from that in control mice. Our finding that tumor uptake of FLT was reduced at 24 h after radiation treatment suggests that FLT PET may be a promising imaging modality for monitoring the early effects of radiation therapy.

Acquisition of resistance to antitumor alkylating agent ACNU: a possible target of positron emission tomography monitoring

Early detection of tumor response to chemotherapy is of great importance for appropriate treatment of tumors. In this study, characteristics of two positron emission tomography (PET) tracers, [ 18F]2-fluoro-2-deoxy- d-glucose (FDG) and[ 18F]3′-fluoro-3′-deoxy-thymidine (FLT), in the early detection of tumor cell response as well as tolerance development to chemotherapy was compared using rat C6 glioma cells and 1-(4-amino-2-methyl-5-pyrimidinyl)-methyl-3-(2-chloroethyl)-3-nitrosoureahydrochloride (ACNU). ACNU is an alkylating agent known to induce drug resistance through expression of O 6-methylguanine-deoxyribonucleic acid methyl transferase ( O 6-MGMT). We established an ACNU-resistant C6 glioma cell line (C6/ACNU) and investigated the effect of ACNU on the uptake of FLT and FDG. In C6 cells, DNA synthesis presented as [ 3H]thymidine ([ 3H]Thd) incorporation into DNA was quickly suppressed by ACNU. In C6/ACNU cells, the suppression was recovered promptly, indicating that DNA alkylation occurs initially but highly expressed O 6-MGMT repairs DNA, leading to the recovery of DNA synthesis. The patterns of FLT uptake in C6 and C6/ACNU were difficult to distinguish in the very early stage of the treatment, though it was reported that FLT uptake well correlated with proliferation in certain conditions. FDG uptake showed different patterns between the resistant and control cells, with significantly decreased uptake in C6 cells and unchanged uptake in C6/ACNU cells at 18–24 h after the treatment. Though difficult to be directly translated into clinical situation, the present study will provide a base to develop an appropriate protocol to assess tumor response to treatment by PET and to design effective treatment plans.

Usefulness of 3′-[F-18]Fluoro-3′-deoxythymidine with Positron Emission Tomography in Predicting Breast Cancer Response to Therapy

The usefulness of 2-deoxy-2-[F-18]fluoro-D-glucose (FDG)-positron emission tomography (PET) in monitoring breast cancer response to chemotherapy has previously been reported. Elevated uptake of FDG by treated tumors can persist however, particularly in the early period after treatment is initiated. 3'-[F-18]Fluoro-3'-deoxythymidine (FLT) has been developed as a marker for cellular proliferation and, in principle, could be a more accurate predictor of the long-term effect of chemotherapy on tumor viability. We examined side-by-side FDG and FLT imaging for monitoring and predicting tumor response to chemotherapy. Fourteen patients with newly diagnosed primary or metastatic breast cancer, who were about to commence a new pharmacologic treatment regimen, were prospectively studied. Dynamic 3-D PET imaging of uptake into a field of view centered over tumor began immediately after administration of FDG or FLT (150 MBq). After 45 minutes of dynamic acquisition, a clinically standard whole-body PET scan was acquired. Patients were scanned with both tracers on two separate days within one week of each other (1) before beginning treatment, (2) two weeks following the end of the first cycle of the new regimen, and (3) following the final cycle of that regimen, or one year after the initial PET scans, whichever came first. (Median and mean times of early scans were 5.0 and 6.6 weeks after treatment initiation; median and mean times for late scans were 26.0 and 30.6 weeks after treatment initiation.) Scan data were analyzed on both tumor-by-tumor and patient-by-patient bases, and compared to each patient's clinical course. Mean change in FLT uptake in primary and metastatic tumors after the first course of chemotherapy showed a significant correlation with late (av. interval 5.8 months) changes in CA27.29 tumor marker levels (r = 0.79, P = 0.001). When comparing changes in tracer uptake after one chemotherapy course versus late changes in tumor size as measured by CT scans, FLT was again a good predictor of eventual tumor response (r = 0.74, P = 0.01). Tumor uptake of FLT was near-maximal by 10 minutes after injection. The time frame five to 10 minutes postinjection of FLT produced standardized uptake value (SUV) values highly correlated with SUV values obtained after 45-minute uptake (r = 0.83, P < 0.0001), and changes in these early SUVs after the first course of chemotherapy correlated with late changes in CA27.29 (r = 0.93, P = 0.003). A 10-minute FLT-PET scan acquired two weeks after the end of the first course of chemotherapy is useful for predicting longer-term efficacy of chemotherapy regimens for women with breast cancer.

18F]3′-deoxy-3′-fluorothymidine PET for the diagnosis and grading of brain tumors

The aim of this study was to evaluate the feasibility of using [(18)F] 3'-deoxy-3'-fluorothymidine (FLT) positron emission tomography (PET) for the diagnosis and grading of brain tumors. The patient population comprised 26 patients (15 males, 11 females) with brain tumors (n=18) or nontumorous lesions (n=8). 2-[(18)F]fluoro-2-deoxy-D: -glucose (FDG) and FLT PET images were obtained using a dedicated PET scanner 1 h after the injection of 370 MBq of FDG or FLT. Uptake of FDG and FLT by the lesions was visually and semiquantitatively assessed in comparison with normal brain tissue. Of 26 brain lesions, four showed increased FDG uptake compared with normal gray matter (grade 5). These four lesions showed intensely increased FLT uptake and were all high-grade tumors. Twenty-two lesions with similar (grade 4) or decreased (grades 1-3) FDG uptake compared with normal gray matter showed variable pathology. Among the 18 brain tumors, FLT PET showed increased uptake in all 12 high-grade tumors but FDG uptake was variable. In 22 brain lesions with similar or decreased uptake compared with normal gray matter on FDG PET, the sensitivity and specificity of FLT PET for the diagnosis of brain tumor were 79% (11/14) and 63% (5/8), respectively. The uptake ratios of 14 brain tumors on FLT PET were significantly higher than the lesion to gray matter ratios (p=0.012) and lesion to white matter ratios (p=0.036) of FDG uptake and differed significantly between high (5.1+/-2.6) and low (2.1+/-1.1) grade tumors (p=0.029). In nine gliomas, FLT uptake was significantly correlated with the Ki-67 proliferation index (rho=0.817, p=0.007). These findings indicate that FLT PET is useful for evaluating tumor grade and cellular proliferation in brain tumors. It displayed high sensitivity and good contrast in evaluating brain lesions that showed similar or decreased uptake compared with normal gray matter on FDG PET. FLT PET, however, did not appear to be sufficiently useful for differentiating tumors from nontumorous lesions.

Clinical relevance of imaging proliferative activity in lung nodules

Recently, the thymidine analogue 3'-deoxy-3'[18F]fluorothymidine (FLT) has been introduced for imaging proliferation with positron emission tomography (PET). In this prospective study, we examined the accuracy of FLT for differentiation of benign from malignant lung lesions and for tumour staging. A total of 47 patients with newly diagnosed pulmonary nodules on chest CT suspicious for malignancy were examined with FLT-PET in addition to routine staging procedures. A total of 43 patients also underwent 2-[18F]fluoro-2-deoxy-D-glucose (FDG) PET imaging. Within 2 weeks, patients underwent resective surgery or core biopsy of the pulmonary lesion. Histopathology revealed malignant lung tumours in 32 patients (20 non-small cell lung cancer, 1 small cell lung cancer, 1 pulmonary carcinoid, 1 non-Hodgkin's lymphoma, nine metastases from extrapulmonary tumours) and benign lesions in 15 patients. Increased FLT uptake was exclusively related to malignant tumours. FLT-PET was false negative in two patients with non-small cell lung cancer, in the patient with a pulmonary carcinoid and in three patients with lung metastases. The sensitivity of FLT-PET for detection of lung cancer was 90%, the specificity 100% and the accuracy 94%. Fifteen out of 21 patients with lung cancer had mediastinal lymph node metastases. FLT-PET was true positive in 7/15 patients, resulting in a sensitivity of 53% for N-staging (specificity 100%, accuracy 67%). Clinical TNM stage was correctly identified in 67% (20/30) patients, compared to 85% (23/27) with FDG-PET. FLT-PET has a high specificity for the detection of malignant lung tumours. Compared with FDG, FLT-PET is less accurate for N-staging in patients with lung cancer and for detection of lung metastases. FLT-PET therefore cannot be recommended for staging of lung cancer.

Metabolism of 3′-deoxy-3′-[F-18]fluorothymidine in proliferating A549 cells: Validations for positron emission tomography

3′-Deoxy-3′-[F-18]fluorothymidine (FLT) is under clinical evaluation as a metabolic probe for imaging cell proliferation in vivo using positron emission tomography (PET). As part of our validation effort, we followed the short-term metabolism of FLT in exponentially growing tumor cells to demonstrate the enzyme activities within the DNA salvage pathway that influence retention of radioactivity. In A549 cells, thymidine kinase-1 ( TK1) activity produced FLTMP, which dominated the labeled nucleotide pool. Subsequent nucleotide phosphorylations by thymidylate kinase ( TMPK) and nucleotide diphosphate kinase ( NDPK) led to FLTTP. After 1h, the cellular metabolic pool contained ∼30% FLTTP. A putative deoxynucleotidase ( dNT), which degrades FLTMP to FLT, provided the primary mechanism for tracer efflux from cells. In contrast, FLTTP was resistant to degradation and highly retained. The uptake and retention characteristics of FLT were also compared to those of thymidine, FMAU (2′- arabino-fluoro-TdR) and FIAU (2′- arabino-fluoro-5-iodo-2′-dexoyuridine). Despite the fact that FLT lacks the 3′-hydroxy necessary for its incorporation into DNA it out performed both FMAU and FIAU in terms of uptake and retention.

Effect of p53 activation on cell growth, thymidine kinase-1 activity, and 3′-deoxy-3′fluorothymidine uptake

The use of thymidine (TdR) and thymidine analogs such as 3′-deoxy-3′-fluorothymidine (FLT) as positron emission tomography (PET)–based tracers of tumor proliferation rate is based on the hypothesis that measurement of uptake of these nucleosides, a function primarily of thymidine kinase-1 (TK 1) activity, provides an accurate measure of cell proliferation in tumors. Tumor growth is influenced by many factors including the oxygen concentration within tumors and whether tumor cells have been exposed to cytotoxic therapies. The p53 gene plays an important role in regulating growth under both of these conditions. The goal of this study was to investigate the influence of p53 activation on cell growth, TK 1 activity, and FLT uptake. To accomplish this, TK 1 activity, S phase fraction, and the uptake of FLT were determined in plateau-phase and exponentially growing cultures of an isogenic pair of human tumor cell lines in which p53 expression was normal or inactivated by human papilloma virus type 16 E6 expression. Ionizing radiation exposure was used to stimulate p53 activity and to induce alterations in cell cycle progression. We found that exposure of cells to ionizing radiation induced dose-dependent changes in cell cycle progression in both cell lines. The relationship between S phase percentage, TK 1 activity, and FLT uptake were essentially unchanged in the p53-normal cell line. In contrast, TK 1 activity and FLT uptake remained high in the p53-deficient variant even when S phase percentage was low due to a p53-dependent G2 arrest. We conclude that a functional p53 response is required to maintain the normal relationship between TK1 activity and S phase percentage following radiation exposure.

18F] 3-deoxy-3′-fluorothymidine positron emission tomography: alternative or diagnostic adjunct to 2-[18f]-fluoro-2-deoxy-d-glucose positron emission tomography in the workup of suspicious central focal lesions?

Background2-[18F]-fluoro-2-deoxy-d-glucose (FDG) positron emission tomography has been established as a standard diagnostic imaging method in the preoperative workup of suspicious pulmonary focal lesions, showing a sensitivity of more than 90% and a specificity of about 80%. Determination of malignant pulmonary lesions with FDG positron emission tomography depends on the assessment of glucose metabolism. However, false-positive findings can occur in inflammatory processes, such as sarcoidosis or pneumonia. The thymidine analogue 3-deoxy-3[18F]-fluorothymidine (FLT) is a new positron emission tomography tracer that more specifically targets proliferative activity of malignant lesions. The objective of this study was to determine whether FLT positron emission tomography, in comparison with FDG positron emission tomography, provides additional information in the preoperative workup of central pulmonary focal lesions. MethodsIn this prospective study FLT and FDG positron emission tomography examinations were performed as a part of the preoperative workup in 20 patients with histologically confirmed bronchial carcinoma, 7 patients with benign lesions, and 1 patient with an atypical carcinoid. Results were compared with final pathologic findings. ResultsFor staging of the primary tumor, FLT positron emission tomography revealed a sensitivity of 86% and a specificity of 100% compared with a sensitivity of 95% and a specificity of 73% for FDG positron emission tomography. For N staging, the sensitivity of FLT positron emission tomography was 57% and the specificity was 100%, and for FDG positron emission tomography, the sensitivity was 86% and the specificity was 100%, respectively. ConclusionsOur preliminary findings indicate specific FLT uptake in malignant lesions. The number of false-positive findings in FDG positron emission tomography might be reduced with FLT positron emission tomography. Therefore positron emission tomography imaging with FLT represents a useful supplement to FDG in assessing the malignancy of central pulmonary focal lesions.

Basis of FLT as a cell proliferation marker: comparative uptake studies with [ 3H]thymidine and [ 3H]arabinothymidine, and cell-analysis in 22 asynchronously growing tumor cell lines

The usefulness of radiolabeled 3′-fluoro-3′-deoxythymidine (FLT), a thymidine derivative with affinity to cytoplasmic thymidine kinase 1 (TK 1), as a tumor proliferation marker was evaluated using [ 3H]FLT and 22 cultured tumor cell lines. Asynchronously growing tumor cells were used for studies to mimic in vivo status of tumors. FLT uptake in each cell line was compared with [ 3H]thymidine ([ 3H]Thd) uptake and %S-phase fraction, both known as acceptable markers of proliferation. Uptake of the mitochondrial TK 2 specific substrate [ 3H]arabinothymidine ([ 3H]AraT) was studied as a reference. Metabolic fate of FLT in tumor cells was also analyzed to elucidate the retention mechanism of FLT. [ 3H]FLT uptake was mildly correlated with the %S-phase fraction (r=0.76, p<0.0001) and correlated better with [ 3H]Thd uptake (r=0.88, p<0.0001). In contrast, the TK 2 specific substrate, [ 3H]AraT, was not significantly correlated with the %S-phase fraction (r=0.19, p=0.39), although it showed some correlation with the [ 3H]Thd uptake (r=0.47, p<0.05). Over 90% of radioactivity of [ 3H]Thd was found in the DNA fraction after 60 minutes incubation. In contrast, most of the radioactivity of [ 3H]FLT was found in the acid-soluble fraction (95%). [ 3H]FLT incorporation into the DNA fraction was negligible (0.2%). The [ 3H]AraT was mainly distributed in the acid-soluble fraction (70%) and the DNA fraction (20%). From our results, we concluded that FLT uptake in tumor cells reflects tumor cell proliferation. However, much more convincing validation is needed to clarify the difference between FLT and true substrates for DNA synthesis, like thymidine.

Kinetics of 3′-Deoxy-3′-[F-18]Fluorothymidine Uptake and Retention in Dogs

We have developed 3′-deoxy-3′-[F-18] fluorothymidine ([F-18]FLT) as an agent to image cellular proliferation with PET. Recent work has demonstrated that [F-18]FLT is stable to degradation and produces high contrast images of proliferating tissues and tumors. To increase our understanding for the use of this agent we have explored the kinetics of [F-18]FLT clearance from the blood and uptake into tissues in normal and tumor bearing dogs. The results indicate that [F-18]FLT is readily modeled in canines with a three-compartment model, with parameter k 3 representing phosphorylation by thymidine kinase. During the first 60 minutes, little loss was measured from the phosphorylated compartment, therefore parameter k 4 could not be differentiated from zero. The extraction of marrow from normal dogs was consistent with this model and demonstrated retention of phosphorylated [F-18]FLT. It is concluded that [F-18]FLT produces images of the DNA synthetic pathway by phosphorylation via thymidine kinase. This pathway can be readily modeled using a three-compartment model. (Mol Imag Biol 2002;4:83–89)

Detection of G1 cell cycle checkpoint integrity in tumor cells by radiolabeled FLT

Purpose: Cell cycle checkpoint integrity plays an important role in both tumor progression and tumor response to therapy. We are developing methods to characterize checkpoint integrity in vivo. The objective of this study was to determine whether the thymidine analog [F-18] 3’-deoxy-3’-fluorothymidine (FLT), an agent being investigated for PET imaging of tumor cell proliferation, could detect differences in cell cycle checkpoint integrity.Materials and methods: We used two isogenic human lung adenocarcinoma cell lines derived from A549 cells. A549-E6 was transfected with a vector containing HPV type 16 E6 to abrogate the function of the tumor suppressor gene, p53, thereby eliminating the ability of radiation to inhibit movement of these cells through the cell cycle. A549-LXSN was transfected with a control vector. There were two types of experiments. In the first, plateau-phase cells were exposed to graded doses of radiation (2-10 Gy) and then released from plateau-phase by subculture. Twenty-four hours later, cells were cultured in the presence of [H-3]-FLT for 1 h and then harvested to measure FLT uptake. In the second experiment, exponentially growing cells were exposed to graded doses of radiation and then, at different times following irradiation (4-72 h), the cells were cultured in the presence of [H-3]-FLT for 1 h before being harvested to measure FLT uptake. For all experiments, the % S phase and TK1 activity were also determined.Results: In the plateau-phase experiments, there was a dose-dependent reduction in FLT uptake in A549-LXSN that corresponded to the reduction in %S phase cells and TK1 activity as increasing numbers of cells arrested in G1. In contrast, A549-E6 showed no decrease in FLT uptake, %S, or TK1 activity under these conditions as these cells failed to arrest in G1 following irradiation. For cells exposed under exponential growth conditions, A549-LXSN showed little change in FLT uptake following a 2-Gy exposure and only a small transient reduction in uptake following a 10-Gy exposure. For A549-E6, there was a small transient increase in FLT uptake following a 2-Gy exposure and a much larger increase following a 10-Gy exposure. As expected, FLT uptake was lowest in cells that arrested in G1. Surprisingly, FLT uptake was highest in cells that arrested in G2.Conclusion: FLT uptake following radiation exposure distinguishes cells with an intact G1 from those where the G1 checkpoint is compromised. Purpose: Cell cycle checkpoint integrity plays an important role in both tumor progression and tumor response to therapy. We are developing methods to characterize checkpoint integrity in vivo. The objective of this study was to determine whether the thymidine analog [F-18] 3’-deoxy-3’-fluorothymidine (FLT), an agent being investigated for PET imaging of tumor cell proliferation, could detect differences in cell cycle checkpoint integrity. Materials and methods: We used two isogenic human lung adenocarcinoma cell lines derived from A549 cells. A549-E6 was transfected with a vector containing HPV type 16 E6 to abrogate the function of the tumor suppressor gene, p53, thereby eliminating the ability of radiation to inhibit movement of these cells through the cell cycle. A549-LXSN was transfected with a control vector. There were two types of experiments. In the first, plateau-phase cells were exposed to graded doses of radiation (2-10 Gy) and then released from plateau-phase by subculture. Twenty-four hours later, cells were cultured in the presence of [H-3]-FLT for 1 h and then harvested to measure FLT uptake. In the second experiment, exponentially growing cells were exposed to graded doses of radiation and then, at different times following irradiation (4-72 h), the cells were cultured in the presence of [H-3]-FLT for 1 h before being harvested to measure FLT uptake. For all experiments, the % S phase and TK1 activity were also determined. Results: In the plateau-phase experiments, there was a dose-dependent reduction in FLT uptake in A549-LXSN that corresponded to the reduction in %S phase cells and TK1 activity as increasing numbers of cells arrested in G1. In contrast, A549-E6 showed no decrease in FLT uptake, %S, or TK1 activity under these conditions as these cells failed to arrest in G1 following irradiation. For cells exposed under exponential growth conditions, A549-LXSN showed little change in FLT uptake following a 2-Gy exposure and only a small transient reduction in uptake following a 10-Gy exposure. For A549-E6, there was a small transient increase in FLT uptake following a 2-Gy exposure and a much larger increase following a 10-Gy exposure. As expected, FLT uptake was lowest in cells that arrested in G1. Surprisingly, FLT uptake was highest in cells that arrested in G2. Conclusion: FLT uptake following radiation exposure distinguishes cells with an intact G1 from those where the G1 checkpoint is compromised.