Fluorothymidine Uptake Research Articles

Early reduction in tumour [18F]fluorothymidine (FLT) uptake in patients with non-small cell lung cancer (NSCLC) treated with radiotherapy alone

PurposeChanges in tumour 3′-deoxy-3′-[18F]fluorothymidine (FLT) uptake during concurrent chemo-radiotherapy in patients with non-small cell lung cancer (NSCLC) have been reported, at variable time points, in two pilot positron emission tomography (PET) studies. The aim of this study was to assess whether FLT changes occur early in response to radiotherapy (RT) without concurrent chemotherapy and whether such changes exceed test-retest variability.MethodsSixteen patients with NSCLC, scheduled to have radical RT, underwent FLT PET once/twice at baseline to assess reproducibility and/or after 5–11 RT fractions to evaluate response. Primary and nodal malignant lesions were manually delineated on CT and volume, mean and maximum standardized uptake values (SUVmean and SUVmax) estimated. Analysis included descriptive statistics and parameter fitting to a mixed-effects model accounting for patients having different numbers of evaluable lesions.ResultsIn all, 35 FLT PET scans from 7 patients with a total of 18 lesions and 12 patients with a total of 30 lesions were evaluated for reproducibility and response, respectively. SUVmean reproducibility in primary tumours (SD 8.9 %) was better than SUVmax reproducibility (SD 12.6 %). In nodes, SUVmean and SUVmax reproducibilities (SD 18.0 and 17.2 %) were comparable but worse than for primary tumours. After 5–11 RT fractions, primary tumour SUVmean decreased significantly by 25 % (p = 0.0001) in the absence of significant volumetric change, whereas metastatic nodes decreased in volume by 31 % (p = 0.020) with a larger SUVmean decrease of 40 % (p < 0.0001). Similar changes were found for SUVmax.ConclusionAcross this group of NSCLC patients, RT induced an early, significant decrease in lesion FLT uptake exceeding test-retest variability. This effect is variable between patients, appears distinct between primary and metastatic nodal lesions, and in primary tumours is lower than previously reported for concurrent chemo-RT at a similar time point. These results confirm the potential for FLT PET to report early on radiation response and to enhance the clinical development of novel drug-radiation combinations by providing an interpretable, early pharmacodynamic end point.Electronic supplementary materialThe online version of this article (doi:10.1007/s00259-013-2632-3) contains supplementary material, which is available to authorized users.

Imaging of Treatment Response to the Combination of Carboplatin and Paclitaxel in Human Ovarian Cancer Xenograft Tumors in Mice Using FDG and FLT PET

IntroductionA combination of carboplatin and paclitaxel is often used as first line chemotherapy for treatment of ovarian cancer. Therefore the use of imaging biomarkers early after initiation of treatment to determine treatment sensitivity would be valuable in order to identify responders from non-responders. In this study we describe the non-invasive PET imaging of glucose uptake and cell proliferation using 2-deoxy-2-[18F]fluoro-D-glucose (FDG) and 3’-deoxy-3’-[18F]fluorothymidine (FLT) for early assessment of treatment response in a pre-clinical mouse model of human ovarian cancer treated with carboplatin and paclitaxel.Methods In vivo uptake of FLT and FDG in human ovarian cancer xenografts in mice (A2780) was determined before treatment with carboplatin and paclitaxel (CaP) and repeatedday 1, 4 and 8 after treatment start. Tracer uptake was quantified using small animal PET/CT. Tracer uptake was compared with gene expression of Ki67, TK1, GLUT1, HK1 and HK2.ResultsTumors in the CaP group was significantly smaller than in the control group (p=0.03) on day 8. On day 4 FDG SUVmax ratio was significantly lower in the CaP group compared to the control group (105±4% vs 138±9%; p=0.002) and on day 8 the FDG SUVmax ratio was lower in the CaP compared to the control group (125±13% vs 167±13%; p=0.05). On day 1 the uptake of FLT SUVmax ratio was 89±9% in the CaP group and 109±6% in the control group; however the difference was not statistically significant (p=0.08). ConclusionsOur data suggest that both FDG and FLT PET may be used for the assessment of anti-tumor effects of a combination of carboplatin and paclitaxel in the treatment of ovarian cancer. FLT provides an early and transient signal and FDG a later and more prolonged response. This underscores the importance of optimal timing between treatment and FLT or FDG imaging since treatment response may otherwise be overlooked.

Preclinical FLT-PET and FDG-PET imaging of tumor response to the multi-targeted Aurora B kinase inhibitor, TAK-901

The Aurora kinases play a key role in mitosis and have recently been identified as attractive targets for therapeutic intervention in cancer. The aim of this study was therefore to investigate the utility of 3'-[(18)F]fluoro-3'-deoxythymidine (FLT) and 2-deoxy-2-[(18)F]fluoro-D-glucose (FDG) for assessment of tumor response to the multi-targeted Aurora B kinase inhibitor, TAK-901. Balb/c nude mice bearing HCT116 colorectal xenografts were treated with up to 30mg/kg TAK 901 or vehicle intravenously twice daily for two days on a weekly cycle. Tumor growth was monitored by calliper measurements and PET imaging was performed at baseline, day 4, 8, 11 and 15. Tumors were harvested at time points corresponding to days of PET imaging for analysis of ex vivo markers of cell proliferation and metabolism together with markers of Aurora B kinase inhibition including phospho-histone H3 (pHH3) and senescence associated β-galactosidase. Tumor growth was inhibited by 60% on day 12 of 30mg/kg TAK-901 therapy. FLT uptake was significantly reduced by day 4 of treatment and this corresponded with reduction in bromodeoxyuridine and pHH3 staining by immunohistochemistry. All biomarkers rebounded towards baseline levels by the commencement of the next treatment cycle, consistent with release of Aurora B kinase suppression. TAK-901 therapy had no impact on glucose metabolism as assessed by FDG uptake and GLUT1 staining by immunohistochemistry. FLT-PET, but not FDG-PET, is a robust non-invasive imaging biomarker of early HCT116 tumor response to the on-target effects of the multi-targeted Aurora B kinase inhibitor, TAK-901. This is the first report to demonstrate the impact of the multi-targeted Aurora B kinase inhibitor, TAK-901 on tumor FLT uptake. The findings provide a strong rationale for the evaluation of FLT-PET as an early biomarker of tumor response in the early phase clinical development of this compound.

Abstract B147: Tumor response imaging with [F-18] fluorothymidine (FLT).

Abstract Background: Current cancer treatments have different mechanisms and variable responses in most histologic groups. The ability to determine treatment response at the molecular level by measuring tumor thymidine kinase 1 activity is being evaluated with FLT PET in groups of patients with different tumor histology. Methods: Under FDA IND approved protocols, patients treated on standard clinical and clinical trial protocols for glioblastoma, carcinoma brain metastases, breast cancer, and AML underwent quantitative PET imaging with FLT at baseline, mid-therapy, and post therapy. Dynamic acquisitions of the sites of known tumor were acquired for 60 minutes, followed by a whole-body static image survey. All images were reconstructed with CT attenuation correction. Regions of interest for the tumor, liver, and surrounding tissues were analyzed for uptake at each data time. Regional tissue FLT uptake was described as the tissue standard uptake variable (SUV), and FLT transport (K1) and flux using a compartmental model analysis. The tumor K1 values were generated to quantify FLT delivery to tumor. Comparisons were made between FLT-PET obtained at sequential times in individual patients and with clinical response. Results: At this time, 19 patients with primary brain tumors, 3 patients with brain metastases, 9 breast cancer patients, and 7 AML patients have been enrolled; at least one post-therapy image has been completed in all but 2 patients. The results have been analyzed semi-quantitatively as SUV and quantitatively by compartmental modeling to determine K1 and flux values for tumor baseline and post therapy comparisons. In most tumors, uptake by either SUV or flux declined in response to treatment but trends in tumor SUV values were not consistent with FLT flux values. In several cases, the FLT K1 and flux values were divergent, emphasizing the requirement to account for FLT delivery changes in observed tumor activity in response to therapy. This effect was most prominent in brain tumors and AML patients. Tumor blood flow/delivery is likely an independent response parameter that can be estimated from analysis of dynamic FLT PET. The poster will show evaluation of the predictive ability of baseline FLT studies as well as the role of pre/post comparisons. Conclusions: FLT PET imaging shows increased tumor uptake across several histologic types. This uptake decreases significantly with therapy, however the flow/delivery parameters and flux values from imaging are important uptake parameters to consider individually to understand changes in response to therapy. Supported by NIH/NCI P01 CA042045-23 and S10 RR017229. Citation Information: Mol Cancer Ther 2013;12(11 Suppl):B147. Citation Format: Janet F. Eary, Jeanne M. Link, Mark Muzi, Finbarr O'Sullivan, Jason K. Rockhill, James R. Fink, Hannah M. Linden, Kenneth A. Krohn. Tumor response imaging with [F-18] fluorothymidine (FLT). [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr B147.

Increased intratumoral fluorothymidine uptake levels following multikinase inhibitor sorafenib treatment in a human renal cell carcinoma xenograft model

An early identification of the tumor response to sorafenib treatment is indispensable for selecting optimal personalized treatment strategies. However, at present, no reliable predictors are clinically available. 18F-fluorothymidine (18F-FLT) positron emission tomography (PET) is used to assess tumor proliferation, since the FLT uptake level reflects thymidine kinase-1 (TK-1) activity. Thus, the present study determined whether FLT was able to evaluate the early tumor response to sorafenib treatment in a human renal cell carcinoma (RCC; A498) xenograft in comparison with the tumor proliferation marker, Ki-67. Mice bearing A498 tumors were assigned to the control and sorafenib-treated groups and the tumor volume was measured every day. [Methyl-3H(N)]-3′-fluoro-3′-deoxythymidine (3H-FLT) was injected 2 h prior to the sacrifice of the mice on days three and seven following the treatment. 3H-FLT autoradiography (ARG) and Ki-67 immunohistochemistry (IHC) were performed using adjacent tumor sections. In the visual assessment, the intratumoral 3H-FLT uptake level diffusely increased following the treatment, while no significant changes were observed in Ki-67 IHC. The intratumoral 3H-FLT uptake levels significantly increased by 2.7- and 2.6-fold on days three and seven following the treatment, while the tumor volume and Ki-67 index did not significantly change. Thus, an increased FLT uptake level was demonstrated following the treatment, which may indicate the suppression of thymidylate synthase (TS) and the compensatory upregulation of TK-1 activity by sorafenib.

Methodological Considerations in Quantification of 3'-Deoxy-3'-[18F]Fluorothymidine Uptake Measured with Positron Emission Tomography in Patients with Non-Small Cell Lung Cancer

To investigate the effect of image-derived input functions (IDIF), input function corrections and volume of interest (VOI) size in quantification of [(18)F]FLT uptake in non-small cell lung cancer (NSCLC) patients. Twenty-three NSCLC patients were scanned on a HR+ scanner. IDIFs were defined over the aorta and left ventricle. Activity concentration and metabolite fraction were measured in venous blood samples. Venous blood samples at 30, 40 and 60min after injection were used to calibrate the IDIF time-activity curves. Adaptive thresholds were used for VOI definition. Full kinetic analysis and simplified measures were performed. Non-linear regression analysis showed better fits for the irreversible model compared to the reversible model in the majority. Calibrated and metabolite corrected plus plasma-to-blood ratio corrected input function resulted in high correlations between SUV and Patlak K i (Pearson correlation coefficients 0.86-0.96, p value < 0.001). No significant differences in correlation between SUV and Patlak K i were observed with variation of IDIF structure or VOI size. Plasma-to-blood ratio correction, metabolite correction and calibration improved the correlation between SUV and Patlak K i significantly, indicating the need for these corrections when K i is used to validate semi-quantitative measures, such as SUV.

NCI 8609: Interim fluoro-3’-deoxythymidine (FLT) PET imaging findings from the phase I trial of PARP inhibitor veliparib (V) and carboplatin (C) in advanced breast cancer.

1023 Background: We are currently conducting a phase I trial of PARP inhibitor, V on an intermittent (7 or 14 day) or continuous (21 day) schedule in combination with C in patients (pts) with advanced breast cancer. We are using FLT PET/CT sequentially to assess DNA damage induced by varying dose schedules of PARP inhibitor, where uptake of FLT depends on the proliferation rate of the tumor. Methods: Eligible pts received C-AUC 5 Q 3weeks (except dose level 1-AUC 6) plus escalating doses of V, BID on 7, 14, or 21-day schedules based on a standard 3+3 dose escalation design. We performed FLT PET/CT at baseline, cycle 1 day 7 and 14 and after cycle 3. Lesions were track-matched with the FDG PET/CT and semi-quantitatively assessed using 2D ROI placement in a matched, blinded fashion. Results: 38 pts have been accrued to 7 dose levels and FLT-PET imaging was successfully obtained in all pts with the proliferative whole body mapping revealing expected bone-marrow, liver and RESuptake. FLT-PET uptake showed a significant (p &lt; 0.001) decrease between baseline and day 7 (N = 25) with an overall trend to rebound nearly to baseline at day 14 for pts that did not show a significant decrease in FLT uptake reduction after cycle 3. The 14-day (n = 15) dosing schedule resulted in more pronounced day 14 reduction in FLT uptake when compared to those on the 7-day (n = 7) schedule. A FLT rebound to baseline level appeared to be associated with limited therapy response. There were no reported toxicities from FLT imaging. Conclusions: FLT-PET was consistently obtained with excellent whole body quality. All lesions revealed a FLT (proliferation) uptake that was different from the FDG (metabolism) uptake. FLT uptake indicated an initial reduction of proliferation at day 7, followed by a rebound at day 21 in all patients on the 7 or 14 day schema. The trial protocol was therefore amended to include a 21 day schema which is currently still ongoing. FLT appears to be a promising in-vivo imaging marker that may serve as a guiding tool to optimize dosing schema in addition to assessing/ predicting overall response. Study support- U01 CA076576 /Wright Center of Innovation ODSA TECH09-028. Clinical trial information: NCT01251874.

Etoposide increases equilibrative nucleoside transporter 1 activity and fluorothymidine uptake: Screening of 60 cytotoxic agents

Equilibrative nucleoside transporter 1 (ENT1) is a major regulator for the uptake of [(18)F]fluorothymidine ([(18)F]FLT), a promising positron emission tomography tracer for treatment monitoring. Various antimetabolites such as 5-fluorouracil often increase ENT1 activity and [(18)F]FLT uptake (flare) in spite of cell death. However, it has not yet been defined which agents induce [(18)F]FLT flare and what is the role of [(18)F]FLT flare in cell viability. Sixty cytotoxic agents from the LOPAC1280 library were screened for ENT1 activity in HeLa cells which predominantly express ENT1, and topoisomerase inhibitors (i.e., aurintricarboxylic acid, idarubicin, camptothecin and etoposide) were identified as potent inducers of ENT1 activity. The changes in ENT1 activity were closely correlated with [(3)H]FLT uptake (Spearman's correlation coefficient, r=0.66; P<0.01). Etoposide significantly increased ENT1 activity and [(3)H]FLT uptake accompanying cell cycle arrest at S/G2/M phase and the increase in TK1 expression and activity in both ENT1-low expressing HT29 and ENT1-high expressing MDA-MB-231 cells. The inhibition of ENT1 activity by dipyridamol or S-(p-nitrobenzyl)-6-thioinosine repressed the etoposide-induced cell death in HeLa cells, whereas it induced no changes in the other cell lines. In conclusion, etoposide is identified as a potent inducer for ENT1 activity and [(3)H]FLT uptake. The role of ENT1 activity by etoposide was cell-type dependent, which requests caution for the application of ENT1-mediated [(18)F]FLT flare for treatment monitoring.

Levels of human equilibrative nucleoside transporter-1 are higher in proliferating regions of A549 tumor cells grown as tumor xenografts in vivo

3'-Fluoro-3'-deoxythymidine (FLT) has been proposed for positron emission tomography (PET)-based identification of tumor chemosensitivity that is mediated by the human equilibrative nucleoside transporter-1 (ENT1). ENT1 facilitates transport of FLT into cells and elevated levels of FLT are associated with both larger FLT-PET signals and increased response to nucleoside-based chemotherapies. FLT-PET is also used as a measure of tumor proliferation. The present study examined the extent to which ENT1 levels vary in a proliferation-dependent manner in tumor cells in vivo. The human adenocarcinoma cell line A549 was used to establish tumor xenografts in nude mice. FLT uptake was measured in vivo using PET, and further examined ex vivo using autoradiography. FLT uptake patterns were compared to immunohistochemical (IHC) analysis of ENT1 and the proliferation markers Ki67 and BrdU. Regional differences in FLT uptake matched differences in IHC proliferation markers. All cells stained for ENT1, but the staining intensity was twice as high for Ki67(+) cells than for Ki67(-) cells. Under in vivo conditions, proliferating regions of tumors show increased FLT uptake and higher ENT1 levels than nonproliferating tumor regions.

Predictive value of 3′-deoxy-3′-[18F]fluorothymidine positron emission tomography/computed tomography for outcome of carbon ion radiotherapy in patients with head and neck mucosal malignant melanoma

The purpose of this prospective study was to assess the prognostic value of 3'-deoxy-3'-[(18)F]fluorothymidine (FLT) positron emission tomography/computed tomography (PET/CT) for the outcome of carbon ion radiotherapy (CIRT) in patients with mucosal malignant melanoma (MMM) of the head and neck. Thirteen patients (69 ± 13 years) with histologically proven MMM tumor were enrolled. CIRT was performed with a total dose of 57.6-64.0 gray equivalents per 16 fractions over a period of 4 weeks. FLT-PET/CT was performed before and again 1 month after CIRT. Tumor FLT uptake was quantitatively assessed using the maximum standardized uptake value (SUV(max)). FLT-PET parameters [pre-CIRT SUV(max), post-CIRT SUV(max), and the reduction rate (RR)] and clinical parameters [age, gender, tumor site, tumor status, gross tumor volume (GTV), and regional lymph node involvement] were evaluated in relation to survival estimates. The follow-up period was 16.1 ± 5.9 months for 9 deceased patients, and 36.7 ± 7.9 months for 4 survivors. Pre-CIRT SUV(max) of ≥4.3, age of ≥80 years old, sinonasal cavity tumor site, and GTV of ≥39 mL were found to be statistically significant prognostic factors for better overall survival. Pre-CIRT SUV(max) of ≥5.0, age of ≥80 years old, sinonasal cavity tumor site, and the absence of regional lymph node involvement were statistically significant prognostic factors for better metastasis-free survival. RR of ≥35 % and GTV of <73 mL were predictive of better local control. The present study indicated for the first time that in patients with the head and neck MMM, FLT-PET/CT imaging was useful for predicting the therapeutic outcome of CIRT. Our results will contribute to the establishment of an effective staging system for MMM based on prognostic factors, depending on treatment choice.

Positron Emission Tomography with [18F]-3′-Deoxy-3′fluorothymidine (FLT) as a Predictor of Outcome in Patients with Locally Advanced Resectable Rectal Cancer: a Pilot Study

This pilot study was performed to evaluate whether tumor uptake of (18)F-labeled 3'-deoxy-3'fluorothymidine (FLT), a proliferative radiotracer, at baseline and early during therapy, is predictive of outcome in locally advanced rectal cancer. Fourteen patients underwent positron emission tomography (PET) with 2-deoxy-2-[(18)F]fluoro-D-glucose (FDG) and FLT before therapy and PET with FLT approximately 2weeks after initiating neoadjuvant chemoradiotherapy. FLT and FDG uptake were evaluated qualitatively and by maximum standardized uptake value (SUV(max)). Tumor FLT and FDG uptake were correlated with disease-free survival (DFS). Thirteen patients underwent surgery after therapy, one died before surgery with progressive disease. FDG-PET/computed tomography detected regional lymph node metastases in five and FLT-PET was positive in one. High pretherapy FDG uptake (SUV(max) ≥ 14.3), low during-therapy FLT uptake (SUV(max) < 2.2), and high percentage change in FLT uptake (≥60%) were predictive of improved DFS (p < 0.05 for all three values). Pretherapy FDG uptake, during-therapy FLT uptake, and percentage change in FLT uptake were equally predictive of DFS.

The effects of 5-fluoruracil treatment on 3′-fluoro-3′-deoxythymidine (FLT) transport and metabolism in proliferating and non-proliferating cultures of human tumor cells

3'-Fluoro-3'-deoxythymidine (FLT) positron emission tomography (PET) has been proposed for imaging thymidylate synthase (TS) inhibition. Agents that target TS and shut down de novo synthesis of thymidine monophosphate increase the uptake and retention of FLT in vitro and in vivo because of a compensating increase in the salvage pathway. Increases in both thymidine kinase-1 (TK1) and the equilibrative nucleoside transporter hENT1 have been reported to underlie this effect. We examined whether the effects of one TS inhibitor, 5-fluorouracil (5FU), on FLT uptake require proliferating cells and whether the effects are limited to increasing TK1 activity. The effects of 5FU on FLT transport and metabolism, TK1 activity, and cell cycle progression were evaluated in the human tumor cell line, A549, maintained as either a proliferating or non-proliferating culture. There were dose-dependent increases in FLT uptake that peaked after a 10 μM 5FU exposure and then declined to baseline levels or below at higher doses in both proliferating and non-proliferating cultures. The dose-dependence for FLT uptake was mirrored by changes in TK1 activity. S phase fraction did not correlate with FLT uptake in proliferating cultures. Chemical inhibition of hENT1 reduced overall levels of FLT uptake but did not affect the low dose increase in FLT uptake. 5FU only affects FLT uptake in proliferating A549 cells and increases in FLT uptake are directly related to increased TK1 activity. Our studies did not support a role for hENT1 in the increased uptake of FLT after exposure to 5FU. Our studies with A549 cells support the suggestion that FLT-PET could provide a measure of TS inhibition in vivo.

Validity of Simplified 3′-Deoxy-3′-[18F]Fluorothymidine Uptake Measures for Monitoring Response to Chemotherapy in Locally Advanced Breast Cancer

PurposePositron emission tomography using 3′-deoxy-3′-[18F]fluorothymidine ([18F]FLT) has been suggested as a means for monitoring response to chemotherapy. The aim of this study was to evaluate the validity of simplified uptake measures for assessing response to chemotherapy using [18F]FLT in locally advanced breast cancer (LABC).ProceduresFifteen LABC patients underwent dynamic [18F]FLT scans both prior to and after the first cycle of chemotherapy with fluorouracil, epirubicin or doxorubicin, and cyclophosphamide. The net uptake rate constant of [18F]FLT, K i, determined by non-linear regression (NLR) of an irreversible two-tissue compartment model was used as the gold standard. In addition to Patlak graphical analysis, standardised uptake values (SUV) and tumour-to-whole blood ratio (TBR) were used for analysing [18F]FLT data. Correlations and relationships between simplified uptake measures and NLR before and after chemotherapy were assessed using regression analysis.ResultsNo significant differences in both pre- and post-chemotherapy relationships between any of the simplified uptake measures and NLR were found. However, changes in SUV between baseline and post-therapy scans showed a significant negative bias and slope less than one, while TBR did not.ConclusionsIn LABC, TBR instead of SUV may be preferred for monitoring response to chemotherapy with [18F]FLT.

18F]FLT–PET Imaging Does Not Always “Light Up” Proliferating Tumor Cells

[(18)F]FLT (3'-Fluoro-3' deoxythymidine)-PET imaging was proposed as a tool for measuring in vivo tumor cell proliferation. The aim of this article was to validate the use of [(18)F]FLT-PET imaging for measuring xenograft proliferation and subsequent monitoring of targeted therapy. In exponentially growing xenografts, factors that could impact the outcome of [(18)F]FLT-PET imaging, such as nucleoside transporters, thymidine kinase 1, the relative contribution of DNA salvage pathway, and the ratio of FLT to thymidine, were evaluated. The [(18)F]FLT tracer avidity was compared with other proliferation markers. In a panel of proliferating xenografts, [(18)F]FLT or [(3)H]thymidine tracer avidity failed to reflect the tumor growth rate across different tumor types, despite the high expressions of Ki67 and TK1. When FLT was injected at the same dose level as used in the preclinical [(18)F]FLT-PET imaging, the plasma exposure ratio of FLT to thymidine was approximately 1:200. Thymidine levels in different tumor types seemed to be variable and exhibited an inverse relationship with the FLT tracer avidity. In contrast, high-dose administration of bromdeoxyuridine (BrdUrd; 50 mg/kg) yielded a plasma exposure of more than 4-fold higher than thymidine and leads to a strong correlation between the BrdUrd uptake and the tumor proliferation rate. In FLT tracer-avid models, [(18)F]FLT-PET imaging as a surrogate biomarker predicted the therapeutic response of CDK4/6 inhibitor PD-0332991. Tumor thymidine level is one of the factors that impact the correlation between [(18)F]FLT uptake and tumor cell proliferation. With careful validation, [(18)F]FLT-PET imaging can be used to monitor antiproliferative therapies in tracer-avid malignancies.

Comparison of 3′-deoxy-3′-[18F]fluorothymidine positron emission tomography (FLT PET) and FDG PET/CT for the detection and characterization of pancreatic tumours

Despite recent advances in clinical imaging modalities, differentiation of pancreatic masses remains difficult. Here, we tested the diagnostic accuracy of molecular-based imaging including 3'-deoxy-3'-[(18)F]fluorothymidine (FLT) positron emission tomography (PET) and [(18)F]fluorodeoxyglucose (FDG) PET/CT in patients with suspected pancreatic masses scheduled to undergo surgery. A total of 46 patients with pancreatic tumours suspicious for malignancy and scheduled for resective surgery were recruited prospectively. In 41 patients, FLT PET and FDG PET/CT scans were performed. A diagnostic CT performed on a routine basis was available in 31 patients. FLT PET and FDG PET/CT emission images were acquired according to standard protocols. Tracer uptake in the tumour [FDG and FLT standardized uptake value (SUV)] was quantified by the region of interest (ROI) technique. For FDG PET/CT analysis, correct ROI placement was ensured via side-by-side reading of corresponding CT images. Of 41 patients, 33 had malignancy, whereas 8 patients had benign disease. Visual analysis of FDG and FLT PET resulted in sensitivity values of 91% (30/33) and 70% (23/33), respectively. Corresponding specificities were 50% (4/8) for FDG PET and 75% (6/8) for FLT PET. In the subgroup of patients with contrast-enhanced CT (n = 31), sensitivities were 96% (PET/CT), 88% (CT alone), 92% (FDG PET) and 72% (FLT PET), respectively. Mean FLT uptake in all malignant tumours was 3.0 (range SUV(max) 1.1-6.5; mean FDG SUV(max) 7.9, range 3.3-17.8; p < 0.001). For differentiation of pancreatic tumours, FDG PET and FDG PET/CT showed a higher sensitivity but lower specificity than FLT PET. Interestingly, visual analysis of FLT PET led to two false-positive findings by misinterpreting physiological bowel uptake as pathological FLT uptake in the pancreas. Due to the limited number of patients, the clinical value of adding FLT PET to the diagnostic workup of pancreatic tumours remains to be determined.

Pharmacodynamic Evaluation of Irinotecan Therapy by FDG and FLT PET/CT Imaging in a Colorectal Cancer Xenograft Model

Longitudinal changes of 3'-[(18) F]fluoro-3'-deoxythymidine (FLT) and 2-deoxy-2-[(18) F]fluoro-D-glucose (FDG) in response to irinotecan therapy in an animal model of colorectal cancer were compared. SCID/CB-17 mice with HCT116 tumors were treated with 50mg/kg irinotecan by intraperitoneal injection weekly for 3weeks. FLT and FDG-positron emission tomography (PET) were performed at baseline, the day after each treatment, and 5days after the first treatment. Proliferation and apoptosis were evaluated by immunohistochemistry (IHC) after day 15 of imaging. Irinotecan treatment resulted in a suppression of tumor growth. Tumor FLT uptake was decreased the day after each treatment but to a lesser extent 5days after the first treatment. FDG uptake increased the day after each treatment with a continuous increase throughout the experiment. IHC analysis of phospho-H3 and Ki67 confirmed FLT-PET results, indicating a decrease in proliferation the day after the final irinotecan treatment. Increased apoptosis monitored by caspase-3 was observed after day 15 with irinotecan treatment. FLT-PET may be a better method than FDG-PET for assessing treatment response to irinotecan. Changes in imaging occur before changes in tumor volume.

Biological basis of [11C]choline-positron emission tomography in patients with breast cancer

The biological significance of [¹¹C]choline (CHO) uptake in human tumours is unclear and probably linked to choline kinase-α (CHKα) expression and cell proliferation. We directly compared CHO with [¹⁸F]fluorothymidine (FLT), an imaging biomarker of proliferation, by positron emission tomography (PET) in patients with breast cancer to investigate whether cell proliferation is an important determinant of CHO uptake. Furthermore, we evaluated CHKα and the Ki67-labelling index (LIKi67) in tumour biopsies. Sequential CHO-PET and FLT-PET within the same imaging session were performed in 21 patients with oestrogen receptor (ER)-positive breast cancer (28 lesions). Average and maximum CHO standardized uptake values (SUV) at 60 min: SUV60,av, and SUV60,max, and the rate constant of net irreversible uptake, Ki, were compared with FLT uptake at 60 min: SUV60,av and SUV60,max. Biopsies were stained for CHKα and LIKi67 in eight cases. Tumours were equally visible on CHO-PET and FLT-PET imaging. Tumour CHO-PET strongly correlated with FLT imaging variables (Pearson's r=0.83; P<0.0001 for CHO-SUV60,max vs. FLT-SUV60,max). A statistically significant association was found between CHO-PET variables and categorical scores of cytoplasmic CHKα intensity and between FLT-PET and LIKi67 (P<0.05, one-way analysis of variance). Choline metabolism and proliferation as assessed by PET were correlated in ER-positive breast cancer, indicating that high CHO uptake is a measure of cellular proliferation in this setting. CHO uptake was also found to be related to cytoplasmic CHKα expression.

The role of nucleoside/nucleotide transport and metabolism in the uptake and retention of 3′-fluoro-3′-deoxythymidine in human B-lymphoblast cells

Recent studies in the human adenocarcinoma cell line A549 have identified cell growth-dependent equilibrative nucleoside transporter-1 (hENT1) as a modifier of 3'-fluoro-3'-deoxythymidine (FLT) uptake and retention. In the present study, we used the ability to isolate human lymphoblastoid clones deficient in thymidine kinase 1 (TK1) to study how metabolism and nucleoside transport influence FLT uptake and retention. Transport and metabolism of FLT were measured in the human lymphoblastoid cell line TK6 and in eight clones isolated from TK6. Four clones were TK1-proficient, while four were TK1-deficient. Both influx and efflux of FLT were measured under conditions where concentrative and equilibrative transport could be distinguished. Sodium-dependent concentrative FLT transport dominated over equilibrative transport mechanisms and while inhibition of hENT1 reduced FLT uptake, there were no correlations between clonal variations in hENT1 levels and FLT uptake. There was an absolute requirement of TK1 for concentration of FLT in TK6 cells. FLT uptake reached a peak after 60 min of incubation with FLT after which intracellular levels of FLT and FLT metabolites declined. Efflux was rapid and was associated with reductions in FLT and each of its metabolites. Both FLT and FLT-monophosphate were found in the efflux buffer. Initial rates of FLT uptake were a function of both concentrative and equilibrative transporters. TK1 activity was an absolute requirement for the accumulation of FLT. Retention was dependent on nucleoside/nucleotide efflux and retrograde metabolism of FLT nucleotides.

Axitinib inhibition of [18F] fluorothymidine (FLT) uptake in patients (pts) with colorectal cancer (CRC): Implications for cytotoxic chemotherapy combinations.

3591 Background: Axitinib is an oral, potent, and selective inhibitor of VEGF receptors (VEGFR) 1, 2, and 3. Clinical trials of VEGFR inhibitors in combination with cytotoxic chemotherapies have yet to yield positive results. Inhibition of cancer cell proliferation by VEGFR inhibitors may be antagonizing cytotoxic chemotherapy. This serial FLT-PET imaging study was performed to determine whether continuous dosing of axitinib inhibits FLT uptake and whether a 3- or 7-day drug holiday could reverse the FLT uptake inhibition. Methods: Pts with CRC or other GI cancers with suitable lesions were enrolled. Axitinib 7 mg (cohort 1) or 10 mg (cohort 2) twice daily (BID) starting dose was administered continuously for 7 days, then interrupted for ≥7 days. Dynamic and whole-body FLT-PET/CT scans were obtained at baseline, at the end of 7 days of continuous axitinib BID dosing, and then 3 and 7 days after axitinib interruption. Kinetic parameters calculated included the Patlak influx constant (Kpatlak) and changes in standardized uptake values (SUVmax, SUVmean). For liver lesions, SUVmean was used. Results: A total of 19 pts were enrolled, 3 and 16 into Cohorts 1 and 2, respectively. 15 had CRC and 4 had other GI cancers; 14 were male, and median age was 56 yrs. Serial FLT-PET data were available for 18 pts: 6 with lung, 5 with liver, 3 with lymph node, 2 with abdominal wall, and 2 with pelvic lesions. Following an initial 7 days of continuous axitinib, tumor FLT uptake (mean DSUV) changed by –46.0% (95% CI: –31.8%, –60.2%) from baseline. Then, after 3 days of axitinib holiday, tumor FLT uptake increased from nadir by +54.5% (95% CI: +17.5%, +91.5%), which further increased by +96.6% (95% CI: +36.4%, +156.8%) after 7 days of axitinib holiday. Conclusions: Continuous axitinib therapy for 7 days markedly inhibited fluorothymidine uptake. Since inhibition of cancer cell proliferation can protect against the cytotoxic effects of chemotherapeutics, this could explain the failure of VEGFR inhibitor + chemotherapy regimens to improve chemotherapy. Inclusion of a short drug holiday in axitinib dosing schedule may allow resensitization of cancer cells.

ACRIN 6688 phase II study of fluorine-18 3′-deoxy-3′ fluorothymidine (FLT) in invasive breast cancer.

TPS125 Background: Neoadjuvant chemotherapy (NAC) prior to surgery provides enhanced options for locoregional management and has become an integral component of primary breast cancer management. Initial tumor response in patients receiving NAC is generally determined at therapy completion. This evaluation is determined by either the presence/absence of palpable tumor as a clinical response and/or presence/absence of invasive tumor cells in the breast and nodes as a pathological response. The ability to evaluate the effectiveness of neoadjuvant therapy early during treatment would be of significant importance. FDG PET imaging has been shown to be predictive of subsequent tumor response, but the tendency of FDG to accumulate in inflammatory tissues can complicate image interpretation. MRI changes have also been touted as predictors of response. Preliminary data suggest that early FLT PET is better able to predict response to therapy, as FLT uptake has been shown to correlate with cellular proliferation, and does not significantly accumulate in inflammatory tissue (Kenny et al, Eur J Nucl Med Mol Imaging 2007:1339-1347). Methods: In this phase II multi-institutional study, breast cancer patients with locally advanced disease with a tumor size ≥2cm (measured on imaging or estimated by physical exam) are eligible. Participants will receive standard of care NAC at their respective institutions. Participants will have 3 FLT imaging sessions to evaluate therapy response: at baseline, early-treatment (5-10 days after initiating treatment), and post-treatment. The primary objective is to correlate the percentage change in the standardized uptake value between baseline and early therapy FLT in the primary tumor with pathologic response. Correlatively, FLT PET parameters will be compared with proliferative indices from the initial biopsy and residual tumor surgical samples using Ki-67 staining, mitotic index, and residual cancer burden. Potential safety issues and the physiologic effects associated with FLT administration will also be evaluated. The analysis of these data may provide a better understanding of early treatment response and improve the clinical management of breast cancer in the future.