3'-deoxy-3'-[18F]fluorothymidine as a new marker for monitoring tumor response to antiproliferative therapy in vivo with positron emission tomography

3'-Deoxy-3'-[(18)F]fluorothymidine ([(18)F]FLT) has been proposed as a new marker for imaging tumor proliferation by positron emission tomography (PET). The uptake of [(18)F]FLT is regulated by cytosolic S-phase-specific thymidine kinase 1 (TK1). In this article, we have investigated the use of [(18)F]FLT to monitor the response of tumors to antiproliferative treatment in vivo. C3H/Hej mice bearing the radiation-induced fibrosarcoma 1 tumor were treated with 5-fluorouracil (5-FU; 165 mg/kg i.p.). Changes in tumor volume and biodistribution of [(18)F]FLT and 2-[(18)F]fluoro-2-deoxy-D-glucose ([(18)F]FDG) were measured in three groups of mice (n = 8-12/group): (a) untreated controls; (b) 24 h after 5-FU; and (c) 48 h after 5-FU. In addition, dynamic [(18)F]FLT-PET imaging was performed on a small animal scanner for 60 min. The metabolism of [(18)F]FLT in tumor, plasma, liver, and urine was determined chromatographically. Proliferation was determined by staining histological sections for proliferating cell nuclear antigen (PCNA). Tumor levels of TK1 protein and cofactor (ATP) were determined by Western blotting and bioluminescence, respectively. Tumor [(18)F]FLT uptake decreased after 5-FU treatment (47.8 +/- 7.0 and 27.1 +/- 3.7% for groups b and c, respectively, compared with group a; P < 0.001). The drug-induced reduction in tumor [(18)F]FLT uptake was significantly more pronounced than that of [(18)F]FDG. The PET image data confirmed lower tumor [(18)F]FLT retention in group c compared with group a, despite a trend toward higher radiotracer delivery for group c. Other than phosphorylation in tumors, [(18)F]FLT was found to be metabolically stable in vivo. The decrease in tumor [(18)F]FLT uptake correlated with the PCNA-labeling index (r = 0.71, P = 0.031) and tumor volume changes after 5-FU treatment (r = 0.58, P = 0.001). In this model system, the decrease in [(18)F]FLT uptake could be explained by changes in catalytic activity but not translation of TK1 protein. Compared with group a, TK1 levels were lower in group b (78.2 +/- 5.2%) but higher in group c (141.3 +/- 9.1%, P < 0.001). In contrast, a stepwise decrease in ATP levels was observed from group a to b to c (P < 0.001). In conclusion, we have demonstrated the ability to measure tumor response to antiproliferative treatment with [(18)F]FLT and PET. In our model system, the radiotracer uptake was correlated with PCNA-labeling index. The decrease in [(18)F]FLT uptake after 5-FU was more pronounced than that of [(18)F]FDG. [(18)F]FLT is, therefore, a promising marker for monitoring antiproliferative drug activity in oncology that warrants additional testing.     

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.     

Early detection of tumor response to chemotherapy by 3'-deoxy-3'-[18F]fluorothymidine positron emission tomography: the effect of cisplatin on a fibrosarcoma tumor model in vivo

We have assessed the potential of [18F]fluorothymidine positron emission tomography ([18F]FLT-PET) to measure early cytostasis and cytotoxicity induced by cisplatin treatment of radiation-induced fibrosarcoma 1 (RIF-1) tumor-bearing mice. Cisplatin-mediated arrest of tumor cell growth and induction of tumor shrinkage at 24 and 48 hours, respectively, were detectable by [18F]FLT-PET. At 24 and 48 hours, the normalized uptake at 60 minutes (tumor/liver radioactivity ratio at 60 minutes after radiotracer injection; NUV60) for [18F]FLT was 0.76 +/- 0.08 (P = 0.03) and 0.51 +/- 0.08 (P = 0.03), respectively, compared with controls (1.02 +/- 0.12). The decrease in [18F]FLT uptake at 24 hours was associated with a decrease in cell proliferation assessed immunohistochemically (a decrease in proliferating cell nuclear antigen labeling index, LI(PCNA), from 14.0 +/- 2.0% to 6.2 +/- 1.0%; P = 0.001), despite the lack of a change in tumor size. There were G1-S and G2-M phase arrests after cisplatin treatment, as determined by cell cycle analysis. For the quantitative measurement of tumor cell proliferation, [18F]FLT-PET was found to be superior to [18F]fluorodeoxyglucose-PET (NUV60 versus LIPCNA: r = 0.89, P = 0.001 and r = 0.55, P = 0.06, respectively). At the biochemical level, we found that the changes in [18F]FLT and [18F]fluorodeoxyglucose uptake were due to changes in levels of thymidine kinase 1 protein, hexokinase, and ATP. This work supports the further development of [18F]FLT-PET as a generic pharmacodynamic readout for early quantitative imaging of drug-induced changes in cell proliferation in vivo.     

Changes in tumor metabolism as readout for Mammalian target of rapamycin kinase inhibition by rapamycin in glioblastoma.

PURPOSE: Inhibition of the protein kinase mammalian target of rapamycin (mTOR) is being evaluated for treatment of a variety of malignancies. However, the effects of mTOR inhibitors are cytostatic and standard size criteria do not reliably identify responding tumors. The aim of this study was to evaluate whether response to mTOR inhibition could be assessed by positron emission tomography (PET) imaging of tumor metabolism. Experiment Design: Glucose, thymidine, and amino acid utilization of human glioma cell lines with varying degrees of sensitivity to mTOR inhibition were assessed by measuring in vitro uptake of [18F]fluorodeoxyglucose ([18F]FDG), [18F]fluorothymidine ([18F]FLT), and [3H]l-tyrosine before and after treatment with the mTOR inhibitor rapamycin. The tumor metabolic activity in vivo was monitored by small-animal PET of tumor-bearing mice. The mechanisms underlying changes in metabolic activity were analyzed by measuring expression and functional activity of enzymes and transporters involved in the uptake of the studied imaging probes. RESULTS: In sensitive cell lines, rapamycin decreased [18F]FDG and [18F]FLT uptake by up to 65% within 24 hours after the start of therapy. This was associated with inhibition of hexokinase and thymidine kinase 1. In contrast, [3H]l-tyrosine uptake was unaffected by rapamycin. The effects of rapamycin on glucose and thymidine metabolism could be imaged noninvasively by PET. In sensitive tumors, [18F]FDG and [18F]FLT uptake decreased within 48 hours by 56 +/- 6% and 52 +/- 8%, respectively, whereas there was no change in rapamycin-resistant tumors. CONCLUSIONS: These encouraging preclinical data warrant clinical trials evaluating [18F]FDG and [18F]FLT-PET for monitoring treatment with mTOR inhibitors in patients.     

Imaging bone and soft tissue tumors with the proliferation marker [18F]fluorodeoxythymidine.

PURPOSE: We have determined the ability of positron emission tomography (PET) with the thymidine analogue 3'-deoxy-3'[18F]fluorothymidine (FLT) to detect manifestation sites of bone and soft tissue tumors, to assess tumor grading, and to differentiate malignant from benign tumors. MATERIALS AND METHODS: In this prospective bicenter trial, FLT-PET was done in 22 patients with established or suspected soft or bone tissue lesions. Routine diagnostic procedures included incisional biopsy, magnetic resonance imaging, and/or contrast-enhanced spiral computed tomography in all patients and [18F]fluorodeoxyglucose (FDG)-PET in 15 patients. Forty-five to 60 minutes after i.v. injection of 350 to 425 MBq FLT, emission and transmission scanning was done. Tracer uptake in the tumor was evaluated semiquantitatively by calculation of mean and maximum standardized uptake values (FLT-SUV) and compared with respective values of FDG. Results were correlated to histopathology and tumor grading. RESULTS: FLT-PET detected all malignant bone or soft tissue tumors (17 of 17). Mean FLT-SUV in benign lesions was 0.7 (range, 0.3-1.3), and 1.3 in low-grade sarcoma (grade 1; range, 1.0-1.6), 4.1 (range, 2.2-6.0; P = 0.002) and 6.1 (range, 2.5-8.3; P = 0.001) in grade 2 and grade 3 tumors, respectively. FLT but not FDG uptake correlated significantly with tumor grading (r = 0.71 versus r = 0.01), and a cutoff value of 2.0 for FLT-SUV discriminated between low- and high-grade tumors. CONCLUSION: In this clinical study, the proliferation marker FLT was suitable for imaging malignant bone or soft tissue tumors. FLT but not FDG uptake correlated significantly with the tumor grade, suggesting FLT as superior PET tracer for noninvasive grading of sarcomas.     

Imaging pharmacodynamics of the alpha-folate receptor-targeted thymidylate synthase inhibitor BGC 945

The assessment of tissue-specific pharmacodynamics is desirable in the development of tumor-targeted therapies. Plasma deoxyuridine (dUrd) levels, a measure of systemic thymidylate synthase (TS) inhibition, has limited application for studying the pharmacodynamics of novel TS inhibitors targeted to the high affinity alpha-folate receptor (FR). Here, we have evaluated the utility of [(18)F]fluorothymidine positron emission tomography ([(18)F]FLT-PET) for imaging the tissue pharmacodynamics of BGC 945, an FR-targeted antifolate TS inhibitor; the nontargeted antifolate BGC 9331 was used for comparison. TS inhibition by both drugs induced a concentration-dependent increase in [(3)H]thymidine uptake in FR-positive human epidermoid KB cells. Membrane-associated equilibrative nucleoside transporter type 1 levels increased from 55,720 +/- 6,101 to 118,700 +/- 5,193 and 130,800 +/- 10,800 per cell at 100 mug/mL of BGC 9331 and BGC 945, respectively, suggesting this as a potential mechanism of increased nucleoside uptake. In keeping with these in vitro findings, tumor [(18)F]FLT accumulation in KB xenografts increased by >/=2-fold after drug treatment with maximal levels at 1 to 4 hours and 4 to 24 hours after BGC 9331 and BGC 945 treatment, respectively. Of interest to FR targeting, BGC 9331, but not BGC 945, induced accumulation of [(18)F]FLT uptake in intestine, a proliferative and TS-responsive tissue. For both drugs, quantitative changes in tumor [(18)F]FLT uptake were associated with increased tumor dUrd levels. In conclusion, we have validated the utility of [(18)F]FLT-PET to image TS inhibition induced by antifolates and shown the tumor-specific activity of BGC 945. This imaging biomarker readout will be useful in the early clinical development of BGC 945.     

FLT-PET may not be a reliable indicator of therapeutic response in p53-null malignancy

FDG (18F-deoxy-glucose) is the current gold standard for PET imaging. FLT (3'-deoxy-3'-(18F-fluorothymidine), a PET imaging marker of proliferation, has been proposed as an alternative to FDG for the assessment of therapeutic response. We examined the therapeutic predictive value of FLT-PET and FDG-PET using CALU-6, a human, p53-null, non-small cell lung cancer cell line with comparison of combined targeted therapy, TRAIL and sorafenib, versus combined conventional chemotherapy, docetaxel and cisplatin. CALU-6 tumor-bearing nu/nu mice (n=46) were evaluated in 3 therapeutic trials measuring FLT and FDG prediction of tumor response at 72 h following initiation of daily combination therapy with targeted agents, TRAIL (200 μg i.v.) and sorafenib (30 mg/kg i.p.) and compared to conventional chemotherapeutics cisplatin (3 mg/kg i.p.) and docetaxel (7.5 mg/kg i.p.). PET imaging response was compared to morphological and histological indicators of therapeutic response, including decreased vascularity (in vivo AngioSense imaging and anti-CD31 staining), slowed tumor growth (caliper measurements), decreased cellular proliferation (Ki-67 staining) and increased apoptosis (TUNEL staining). Decreases in tumor accumulation of FLT (FLTMAX -30%, p=0.03) at 72 h post treatment were observed in response to TRAIL and sorafenib combination therapy resulting in smaller, less vascular, more apoptotic tumors. No similar reduction in tumor accumulation of FLT (FLTMAX -2%, p=0.67) was observed 72 h following initiation of cisplatin and docetaxel combination therapy, despite histological and morphological evidence of drug response. In contrast, tumor imaging with FDG did demonstrate a decrease in accumulation in both treatment groups, -21% (p=0.30) in response to cisplatin/docetaxel and -8% (p=0.59) in response to TRAIL/sorafenib, but did not reach statistical significance. FLT, but not FDG, is predictive of therapeutic response to the targeted regimen TRAIL/sorafenib. However, FLT-PET may not predict therapeutic response to DNA damaging agents in p53-null tumors, likely due to loss of cell cycle control of thymidine kinase 1 (TK1). Thus, tumor imaging response by FLT may be limited in human tumors without functional p53.     

Early response assessment using 3'-deoxy-3'-[18F]fluorothymidine-positron emission tomography in high-grade non-Hodgkin's lymphoma.

PURPOSE: To evaluate 3'-deoxy-3'-[(18)F]fluorothymidine-positron emission tomography (FLT-PET) for early monitoring response of high-grade non-Hodgkin's lymphoma to treatment with cyclophosphamide-adriamycin-vincristine-prednisone chemotherapy with or without rituximab immunotherapy (R-CHOP/CHOP). EXPERIMENTAL DESIGN: Twenty-two patients with histologically proven high-grade non-Hodgkin's lymphoma scheduled to undergo first line treatment with R-CHOP/CHOP were included. All patients received baseline imaging before therapy with FLT-PET. For noninvasive assessment of treatment response, FLT-PET was repeated at following time points: group 1 (n = 6), 1 and 6 weeks after R-CHOP/CHOP; group 2 (n = 16), 2 days after rituximab and 2 days after CHOP application. Emission images were acquired 45 min after injection of 300 to 370 MBq of FLT. FLT uptake was quantified by region-of-interest technique on a lesion basis. Maximum standardized uptake values (SUV) for FLT were calculated using circular region of interest (diameter, 1.5 cm). RESULTS: In all patients, morphologically proven lesions showed initially high FLT uptake (mean SUV, 8.1 +/- 3.9). In group 1, mean FLT SUV decreased 7 days after R-CHOP/CHOP by 77% (P < 0.001), the reduction in FLT SUV from baseline was 85% after 40 days (P = 0.003). In group 2, FLT uptake in patients without dexamethasone pretreatment revealed no significant reduction after rituximab (P = 0.3) but significantly decreased 2 days after CHOP to 32% compared with the baseline value (P = 0.004). CONCLUSIONS: Administration of R-CHOP/CHOP is associated with an early decrease in lymphoma FLT uptake. Interestingly, there was no reduction of FLT uptake after rituximab alone, indicating no early antiproliferative effect of immunotherapy. FLT-PET seems to be promising for early evaluation of drug effects in lymphoma.     

3-deoxy-3-[(18)F]fluorothymidine-positron emission tomography for noninvasive assessment of proliferation in pulmonary nodules

We investigated whether uptake of the thymidine analogue 3-deoxy-3-[(18)F]fluorothymidine ([(18)F]FLT) reflects proliferation in solitary pulmonary nodules (SPNs). Thirty patients with SPNs were prospectively examined with positron emission tomography. Standardized uptake values were calculated for quantification of FLT uptake. Histopathology revealed 22 malignant and 8 benign lesions. Proliferation was evaluated by Ki-67 immunostaining and showed a mean proliferation fraction of 30.9% (range, 1-65%) in malignant SPNs and <5% in benign lesions. Linear regression analysis indicated a significant correlation between FLT-standardized uptake values and proliferative activity (P < 0.0001; r = 0.87). FLT uptake was specific for malignant lesions and may be used for differential diagnosis of SPNs, assessment of proliferation, and estimation of prognosis.     

Quantification of cellular proliferation in tumor and normal tissues of patients with breast cancer by [18F]fluorothymidine-positron emission tomography imaging: evaluation of analytical methods

There is an unmet need to develop imaging methods for the early and objective assessment of breast tumors to therapy. 3'-Deoxy-3'-[18F]fluorothymidine ([18F]FLT)-positron emission tomography represents a new approach to imaging thymidine kinase activity, and hence, cellular proliferation. We compared graphical, spectral, and semiquantitative analytic methodologies for quantifying [18F]FLT kinetics in tumor and normal tissue of patients with locally advanced and metastatic breast cancer. The resultant kinetic parameters were correlated with the Ki-67 labeling index from tumor biopsies. [18F]FLT accumulation was detected in primary tumor, nodal disease, and lung metastasis. In large tumors, there was substantial heterogeneity in regional radiotracer uptake, reflecting heterogeneity in cellular proliferation; radiotracer uptake in primary tumors also differed from that of metastases. [18F]FLT was metabolized in patients to a single metabolite [18F]FLT-glucuronide. Unmetabolized [18F]FLT accounted for 71.54 +/- 1.50% of plasma radioactivity by 90 minutes. The rate constant for the metabolite-corrected net irreversible uptake of [18F]FLT (Ki) ranged from 0.6 to 10.4 x 10(-4) and from 0 to 0.6 x 10(-4) mL plasma cleared/s/mL tissue in tumor (29 regions, 15 patients) and normal tissues, respectively. Tumor Ki and fractional retention of radiotracer determined by spectral analysis correlated with Ki-67 labeling index (r = 0.92, P < 0.0001 and r = 0.92, P < 0.0001, respectively). These correlations were superior to those determined by semiquantitative methods. We conclude that [18F]FLT-positron emission tomography is a promising clinical tool for imaging cellular proliferation in breast cancer, and is most predictive when analyzed by graphical and spectral methods.     

Fluorine-18-alpha-methyltyrosine positron emission tomography for diagnosis and staging of lung cancer: a clinicopathologic study.

PURPOSE: L-[3-(18)F]-alpha-methyltyrosine ([(18)F]FMT) is an amino acid tracer for positron emission tomography (PET). We evaluated the diagnostic usefulness of [(18)F]FMT PET in non-small-cell lung cancer (NSCLC) patients. Tumor uptake of [(18)F]FMT was compared with that of 2-[(18)F]-fluoro-2-deoxy-D-glucose ([(18)F]FDG) and correlated with L-type amino acid transporter 1 (LAT1) expression. EXPERIMENTAL DESIGN: Fifty NSCLC patients were enrolled in this study, and a pair of PET study with [(18)F]FMT and [(18)F]FDG was done. LAT1 expression and Ki-67 labeling index of the resected tumors were analyzed by immunohistochemical staining. RESULTS: For the primary tumor detection, [(18)F]FMT PET exhibited a sensitivity of 90% whereas the sensitivity for [(18)F]FDG PET was 94%. For lymph node staging, the sensitivity and specificity of [(18)F]FMT PET were 57.8% and 100%, and those of [(18)F]FDG PET were 65.7% and 91%, respectively. The expression of LAT1 in squamous cell carcinoma and large cell carcinoma was significantly higher than that in adenocarcinoma. [(18)F]FMT uptake was also higher in squamous cell carcinoma and large cell carcinoma than in adenocarcinoma. Uptake of [(18)F]FMT in the tumor is closely correlated with LAT1 expression (rho = 0.890). CONCLUSION: [(18)F]FMT PET had no false-positives in the detection of primary tumor and lymph node metastasis and could improve the diagnostic performance in NSCLC. Uptake of [(18)F]FMT correlated with the expression of LAT1 that showed a significant association with cellular proliferation.     

Redistribution of nucleoside transporters to the cell membrane provides a novel approach for imaging thymidylate synthase inhibition by positron emission tomography

Thymidylate synthase (EC 2.1.1.45) is a key enzyme for the de novo synthesis of DNA and as such a target for anticancer drug development. There is a need to develop noninvasive methods for assessing thymidylate synthase inhibition in tumors. The aim of this study was to assess the potential of 3'-deoxy-3'-[(18)F]fluorothymidine ([(18)F]FLT) positron emission tomography (PET) for early measurement of thymidylate synthase inhibition and to elucidate the cellular mechanisms involved. Radiation-induced fibrosarcoma-1 tumor-bearing mice were injected with a single i.p. dose of the thymidylate synthase inhibitor 5-fluorouracil (5-FU; 165 mg/kg) and imaged by [(18)F]FLT-PET at 1 to 2 hours after treatment. Deoxyuridine, thymidine kinase 1 (cytoplasmic thymidine kinase; EC2.7.1.21), and ATP levels in excised tumors were measured. Cellular assays for membrane transport were also done. There was a 1.8-fold increase in the 60-minute [(18)F]FLT tumor/heart radioactivity ratio in drug-treated mice compared with vehicle controls (P = 0.0016). Plasma and tumor deoxyuridine levels increased significantly but thymidine kinase and ATP levels were unchanged. Whole-cell assays implicated a (low level) functional role for the type-1 equilibrative nucleoside transporter (ENT). There was an increase in type-1 ENT-binding sites per cell from 49,110 in untreated cells to 73,142 (P = 0.03) in cells treated with 10 microg/mL 5-FU for 2 hours, without a change in transporter affinity (P = 0.41). We conclude that [(18)F]FLT-PET can be used to measure thymidylate synthase inhibition as early as 1 to 2 hours after treatment with 5-FU by a mechanism involving redistribution of nucleoside transporters to the plasma membrane.     

In vivo validation of 3'deoxy-3'-[(18)F]fluorothymidine ([(18)F]FLT) as a proliferation imaging tracer in humans: correlation of [(18)F]FLT uptake by positron emission tomography with Ki-67 immunohistochemistry and flow cytometry in human lung tumors.

PURPOSE: Tumor proliferation has prognostic value in resected early stage non-small cell lung cancer (NSCLC) and can, therefore, predict which NSCLCs are at high risk for recurrence after resection and would benefit from additional therapy. It may also predict which tumor will respond to cell cycle-targeted chemotherapy and help assess the tumor response, besides helping to differentiate benign from malignant lung lesions. We evaluated whether the uptake of the new positron emission tomography (PET) tracer 3'deoxy-3'-[18F]fluorothymidine (FLT) in a series of suspected NSCLCs correlated with tumor proliferation assessed by Ki-67 immunohistochemistry and flow cytometry. EXPERIMENTAL DESIGN: Ten patients with 11 biopsy-proven or clinically suspected NSCLC underwent 2-h dynamic PET imaging after i.v. injection of 0.07 mCi/kg FLT. Tumor FLT uptake was quantitated with the maximum pixel standardized uptake value (maxSUV), the partial volume corrected maxSUV (PV-corr-maxSUV), the average SUV over a small region-of-interest (aveSUV) and with Patlak analysis of FLT flux (aveFLTflux). The lesion diameter from computed tomography was used to correct the maxSUV for PV effects using recovery coefficients determined for the General Electric Advance PET scanner. Two of the 11 lesions were benign inflammatory lesions and 9 were NSCLCs. Immunohistochemistry for Ki-67 (proliferation index marker) was performed on all 11 tissue specimens (10 resections, 1 NSCLC percutaneous biopsy), and the S-phase fraction (SPF) from flow cytometry could be determined for 10. The specimens were reviewed for histology and cellular differentiation (poor, moderate, well). Lesions ranged from 1.6 to 7.7 cm. RESULTS: Excellent correlations were found between SUV measures of FLT uptake and Ki-67 scores [percentage of positive cells; maxSUV versus Ki-67: Rho = 0.78, P = 0.0043 (n = 11); PV-corr-maxSUV versus Ki-67: Rho = 0.83, P = 0.0028 (n = 10); aveSUV versus Ki-67: Rho = 0.84, P = 0.0011 (n = 11)]. Correlation between Ki-67 proliferation scores and Patlak measures of FLT uptake were also strong: aveFLTflux versus Ki-67: Rho = 0.94, P < 0.0001 (n = 11). The correlation between the SPF and all indices of FLT uptake was weaker and reached statistical significance for only two uptake indices [maxSUV versus SPF: Rho = 0.69, P = 0.03 (n = 10); PV-corr-maxSUV versus SPF: Rho = 0.36, P = 0.35 (n = 9); aveSUV versus SPF: Rho = 0.67, P = 0.03 (n = 10); aveFLTflux versus SPF: Rho = 0.46, P = 0.18 (n = 10)]. CONCLUSION: FLT PET may be used to noninvasively assess proliferation rates of lung masses in vivo. Therefore, FLT PET may play a significant role in the evaluation of indeterminate pulmonary lesions, in the prognostic assessment of resectable NSCLC, and possibly in the evaluation of NSCLC response to chemotherapy.     

A systematic review on [18F]FLT-PET uptake as a measure of treatment response in cancer patients

Imaging biomarkers have a potential to depict the hallmarks of cancers that characterise cancer cells as compared to normal cells. One pertinent example is 3′-deoxy-3′-¹⁸F-fluorothymidine positron emission tomography ([¹⁸F]FLT-PET), which allows non-invasive in vivo assessment of tumour proliferation. Most importantly, [¹⁸F]FLT does not seem to be accumulating in inflammatory processes, as seen in [¹⁸F]-fludeoxyglucose, the most commonly used PET tracer for assessment of cell metabolism. [¹⁸F]FLT could therefore provide additional information about the tumour biology before, during and after treatment. This systematic review focuses on the use of [¹⁸F]FLT-PET tumour uptake values as a measure of tumour response to therapeutic interventions. The clinical studies which evaluated the role of [¹⁸F]FLT-PET as a measure of tumour response to treatment are summarised and the evidence linking [¹⁸F]FLT-PET tumour uptake values with clinical outcome is evaluated.     

18F]FDG and [18F]FLT uptake in human breast cancer cells in relation to the effects of chemotherapy: an in vitro study

Increased 2'-deoxy-2'-[18F]fluoro-D-glucose (FDG) uptake is the most commonly used marker for positron emission tomography in oncology. However, a proliferation tracer such as 3'-deoxy-3'-[18F]fluorothymidine (FLT) might be more specific for cancer. 3'-deoxy-3'-[18F]fluorothymidine uptake is dependent on thymidine kinase 1 (TK) activity, but the effects of chemotherapeutic agents are unknown. The aim of this study was to characterise FDG and FLT uptake mechanisms in vitro before and after exposure to chemotherapeutic agents. The effects of 5-fluorouracil (5-FU), doxorubicin and paclitaxel on FDG and FLT uptake were measured in MDA MB231 human breast cancer cells in relation to cell cycle distribution, expression and enzyme activity of TK-1. At IC50 concentrations, 5-FU resulted in accumulation in the G1 phase, but doxorubicin and paclitaxel induced a G2/M accumulation. Compared with untreated cells, 5-FU and doxorubicin increased TK-1 levels by >300. At 72 h, 5-FU decreased FDG uptake by 50% and FLT uptake by 54%, whereas doxorubicin increased FDG and FLT uptake by 71 and 173%, respectively. Paclitaxel increased FDG uptake with >100% after 48 h, whereas FLT uptake hardly changed. In conclusion, various chemotherapeutic agents, commonly used in the treatment of breast cancer, have different effects on the time course of uptake of both FDG and FLT in vitro. This might have implications for interpretation of clinical findings.     

False-positive findings on [18F]FDG-PET caused by non-neoplastic cellular elements after neoadjuvant chemoradiotherapy for non-small cell lung cancer

We report two patients with non-small cell lung cancer who had a pathologically complete response after neoadjuvant chemoradiotherapy, although they had positive [(18)F]fluoro-deoxyglucose positron emission tomography ([(18)F]FDG-PET) scans. They underwent concurrent chemoradiotherapy, which resulted in a partial response determined by computed tomography (CT). While [(18)F]FDG-PET after chemoradiotherapy was positive, pathological examination showed that the tumors were fibrotic lesions with infiltration of lymphocytes and macrophages, with the appearance of metaplastic epithelial cells. The reasons for the false-positive results on [(18)F]FDG-PET were considered to be the high uptake of FDG in non-neoplastic inflammatory cellular elements, i.e. macrophages, lymphocytes and metaplastic epithelial cells, and squamous metaplasia induced by chemoradiotherapy. Although several studies demonstrated that [(18)F]FDG-PET could predict the response of neoadjuvant treatment of non-small cell lung cancer, one should bear in mind that false-positive results could be observed in pathological complete response of non-small cell lung cancer after neoadjuvant chemoradiotherapy.     

Quantifying the activity of adenoviral E1A CR2 deletion mutants using renilla luciferase bioluminescence and 3'-deoxy-3'-[18F]fluorothymidine positron emission tomography imaging

The adenoviral E1A CR2 mutant dl922-947 has potent activity in ovarian cancer. We have used Renilla luciferase bioluminescence imaging to monitor viral E1A expression and replication and [18F]fluorothymidine positron emission tomography ([18F]FLT-PET) to quantify the activity of dl922-947 in vivo. We created dlCR2 Ren, with the same E1A CR2 deletion as dl922-947 and the luciferase gene from Renilla reniformis downstream of E1. Light emitted from s.c. and i.p. IGROV1 ovarian carcinoma xenografts was measured following treatment with dlCR2 Ren. Mice bearing s.c. IGROV1 xenografts were injected with 2.96 to 3.7 MBq of [18F]FLT 48 and 168 hours following i.t. injection of dl922-947 or control virus Ad LM-X. The presence of Renilla luciferase in dlCR2 Ren did not reduce in vitro nor in vivo potency compared with dl922-947. Light emission correlated closely with E1A expression in vitro and peaked 48 hours after dlCR2 Ren injection in both s.c. and i.p. IGROV1 xenografts. It diminished by 168 hours in s.c. tumors but persisted for at least 2 weeks in i.p. models. Normalized tumor [18F]FLT uptake at 60 minutes (NUV60), fractional retention, and area under radioactivity curve all decreased marginally 48 hours after dl922-947 treatment and significantly at 168 hours compared with controls. There was a close linear correlation between NUV60 and both tumor proliferation (Ki67 labeling index) and thymidine kinase 1 expression. Renilla luciferase bioluminescence and [18F]FLT-PET imaging are capable of quantifying the activity and effectiveness of E1A CR2-deleted adenoviral mutants in ovarian cancer.     

Functional diagnosis of residual lymphomas after radiochemotherapy with positron emission tomography comparing FDG- and FLT-PET

Positron emission tomography (PET) using 2-deoxy-2-[(18)F]fluoro-D-glucose (FDG) is used as a functional imaging technique for the staging and follow-up of lymphomas. However, additional information about the tumor proliferation rate using 3'-deoxy-3'-[(18)F]fluorothymidine (FLT) may be useful for the assessment of prognosis. We enrolled 48 patients with Hodgkin's (n = 15) and non-Hodgkin's lymphoma (n = 33) with residual masses >2 cm examined by tracer studies with FDG and FLT. The results were related to median overall and progression-free survival. In 15 out of 48 patients analysed using FDG, positive results were found. Using FLT, 10 out of 48 patients were positive. 33 patients were FDG negative. Eight patients were positive both using FDG and FLT. Overall survival for patients with a negative PET scan was significantly higher than for patients with positive PET, irrespective of the tracer used. FLT alone was able to discriminate between patients with long or short overall survival. However, there was no statistical significance comparing FDG/FLT negative versus FDG negative alone. Although FDG detected more lesions than did FLT, the additional biological characterization of tumor tissue with respect to proliferation by FLT might be useful by providing complementary information for the identification of recurrence. However, the present data show no advantage of combined FDG/FLT studies over FDG alone with respect to the prediction of survival.     

3'-[18F]fluoro-3'-deoxythymidine ([18F]-FLT) as positron emission tomography tracer for imaging proliferation in a murine B-Cell lymphoma model and in the human disease

Here we describe the evaluation of 3'-[(18)F]fluoro-3'-deoxythymidine [[(18)F]-FLT] as a tracer for positron emission tomography (PET) in a murine model of B-cell lymphoma and in human malignant lymphoma. The human B-cell line DoHH2 expressed high levels of active thymidine kinase 1 (TK-1) as the key enzyme of [(18)F]-FLT metabolism. Immunostaining confirmed high levels of TK-1 in DoHH2 derived xenograft tumors in SCID/SCID mice. In vitro studies demonstrated a time-dependent uptake of [(18)F]-FLT, an efficient phosphorylation to the respective monophosphate and the incorporation of [(18)F]-FLT into the perchloric acid insoluble fraction in DoHH2 cells, indicating the incorporation of this tracer into the DNA. After incubation with [(18)F]FLT for 240 min, 12.5% +/- 1.0% of radioactivity applied to the medium was intracellularly trapped in DoHH2 cells. Specific accumulation of [(18)F]-FLT in the malignant cell clone was confirmed in biodistribution studies in SCID/SCID mice bearing DoHH2-derived tumors. The percentage of injected dose of [(18)F]-FLT per gram of tumor tissue correlated with the tumor-proliferation index as evaluated in BrdUrd-labeling experiments. In a pilot study of 11 patients with both indolent and aggressive lymphoma, [(18)F]-FLT was suitable and comparable to [(18)F]-FDG in the ability to detect malignant lesions by PET scan. Furthermore, we found a close correlation (r = 0.95, P < 0.005) of the [(18)F]-FLT standardized uptake values with the Ki67-labeling index of tissue biopsies (n = 10) in these patients. These results suggest that [(18)F]-FLT represents a novel tracer for PET that enables imaging of proliferation in human lymphoma in vivo.