Radiosensitizing effect of the novel Hsp90 inhibitor NVP-AUY922 in human tumour cell lines silenced for Hsp90α

Background

Hsp90 inhibitors can enhance the tumour sensitivity to ionising radiation (IR). However, Hsp90 inhibition leads to the up-regulation of anti-apoptotic Hsp90 and Hsp70, which might diminish the radiosensitizing effects of the inhibitors. Therefore, inhibition of the up-regulation of Hsp90 by siRNA might be a promising strategy to enhance drug-mediated radiosensitization.

Materials and methods

The expression of Hsp90α was silenced in A549 and GaMG tumour cell lines by siRNA treatment. Pre-silenced for Hsp90α cells were treated with NVP-AUY922, a novel Hsp90 inhibitor, for 24?h and then irradiated. Radiation response was determined by colony-forming ability. The expression of several marker proteins was analysed by Western blot. DNA damage and repair were assessed by histone γH2AX measurements.

Results

We found that transfection with siRNA against Hsp90α reduced Hsp90α at mRNA and protein levels. Pre-silencing of Hsp90α reduced NVP-AUY922-mediated up-regulation of Hsp90α but it did not increase drug-mediated radiosensitization in both tumour cell lines. As revealed by Western blot, pre-silencing of Hsp90α followed by NVP-AUY922 did not change the expression of Hsp90 client proteins (Akt, Raf-1, Cdk1 and Cdk4) compared with drug treatment alone, suggesting unchanged chaperone function in transfected cells.

Conclusion

Pre-silencing of Hsp90α followed by Hsp90 inhibition did not enhance the radiosensitizing effect of NVP-AUY922 in both tested tumour cell lines. Future work will be done on stable transfection with shRNA against Hsp90α or simultaneous silencing of both Hsp90 isoforms, Hsp90α and Hsp90β, in order to optimize tumour cell killing.     

Modeling spheroid growth, PET tracer uptake, and treatment effects of the Hsp90 inhibitor NVP-AUY922.

For a PET agent to be successful as a biomarker in early clinical trials of new anticancer agents, some conditions need to be fulfilled: the selected tracer should show a response that is related to the antitumoral effects, the quantitative value of this response should be interpretable to the antitumoral action, and the timing of the PET scan should be optimized to action of the drug. These conditions are not necessarily known at the start of a drug-development program and need to be explored. We proposed a translational imaging activity in which experiments in spheroids and later in xenografts are coupled to modeling of growth inhibition and to the related changes in the kinetics of PET tracers and other biomarkers. In addition, we demonstrated how this information can be used for planning clinical trials. METHODS: The first part of this concept is illustrated in a spheroid model with BT474 breast cancer cells treated with the heat shock protein 90 (Hsp90) inhibitor NVP-AUY922. The growth-inhibitory effect after a pulse treatment with the drug was measured with digital image analysis to determine effects on volume with high accuracy. The growth-inhibitory effect was described mathematically by a combined E(max) and time course model fitted to the data. The model was then used to simulate a once-per-week treatment; in these experiments the uptake of the PET tracers (18)F-FDG and 3'-deoxy-3'-(18)F-fluorothymidine ((18)F-FLT) was determined at different doses and different time points. RESULTS: A drug exposure of 2 h followed by washout of the drug from the culture medium generated growth inhibition that was maximal at the earliest time point of 1 d and decreased exponentially with time during 10-12 d. The uptake of (18)F-FDG per viable tumor volume was minimally affected by the treatment, whereas the (18)F-FLT uptake decreased in correlation with the growth inhibition. CONCLUSION: The study suggests a prolonged action of the Hsp90 inhibitor that supports a once-per-week schedule. (18)F-FLT is a suitable tracer for the monitoring of effect, and the (18)F-FLT PET study might be performed within 3 d after dosing.     

Comparison of conventional and novel PET tracers for imaging mesothelioma in nude mice with subcutaneous and intrapleural xenografts

IntroductionMalignant mesothelioma is a highly aggressive tumor originating in the pleura, peritoneum and pericardium, and the prognosis of patients undergoing current treatment remains poor. To develop new therapies, it is important to have a noninvasive imaging system for evaluating the efficacy of such prospective treatments. We have established clinically relevant mouse models and evaluated conventional and novel positron emission tomography (PET) tracers.MethodsEpithelioid and sarcomatoid mesothelioma cells were inoculated subcutaneously and intrapleurally into nude mice. Biodistribution and PET imaging studies were conducted by injecting [¹⁸F]fluoro-2-deoxy-d-glucose (FDG), 3′-[¹⁸F]fluoro-3′-doxythymidine (FLT) or 4′-methyl-[¹¹C]thiothymidine (S-dThd) into the mouse models. In vitro cellular uptake of [¹⁴C]FDG and [³H]FLT and thymidine kinase 1 (TK₁) activity in both cell lines were measured. Expression of glucose transporter 1 (GLUT-1) and Ki-67 in xenografted tumors was evaluated by immunohistochemical staining.ResultsIn epithelioid mesothelioma models, biodistribution experiments showed that tumor uptake of [¹¹C]S-dThd was significantly higher than that of [¹⁸F]FDG. On the other hand, in sarcomatoid models, [¹⁸F]FDG showed significantly higher accumulation than the other two tracers. These differential uptakes of the three tracers were confirmed by PET imaging. The cellular uptake of [¹⁴C]FDG and [³H]FLT and TK₁ activity in sarcomatoid cells were higher than those of epithelioid cells. GLUT-1 protein was strongly expressed in sarcomatoid but not in epithelioid tumor. We observed a high percentage of Ki-67-positive cells in both epithelioid and sarcomatoid tumors.ConclusionsWe established nude mouse models of epithelioid and sarcomatoid subtypes of mesothelioma. PET tracers applicable for the evaluation of epithelioid and sarcomatoid mesothelioma would vary: [¹⁸F]FLT and [¹¹C]S-dThd seemed suitable for the epithelioid subtype and [¹⁸F]FDG seemed suitable for the sarcomatoid subtype in our mouse models. Our results indicated that cellular uptake and TK₁ activity in vitro are not always consistent with tracer uptake of [¹⁸F]FLT and [¹¹C]S-dThd in vivo. These mouse models and PET imaging might be useful tools for evaluating new and effective treatments in mesothelioma.     

Comparison of semiquantitative fluorescence imaging and PET tracer uptake in mesothelioma models as a monitoring system for growth and therapeutic effects

IntroductionVarious techniques are available for in vivo imaging, and precise understanding of their characteristics is essential for effective use of the imaging results. We established human mesothelioma cell lines expressing red fluorescent protein (RFP) and examined their fluorescence intensity and uptake of positron emission tomography (PET) tracer analogs to compare their characteristics and assess their usefulness in the evaluation of therapeutics.MethodA human mesothelioma cell line was stably transfected to express RFP. Fluorescence, cell number and protein amount were measured during cell growth and treatment with cytotoxic reagents. In in vivo experiments, RFP-expressing cells were injected subcutaneously or into the pleural cavity of nude mice, and fluorescence images were taken with or without pemetrexed treatment. The uptake of [³H]3′-deoxy-3′-fluorothymidine ([³H]FLT) and [¹⁴C]2-fluoro-2-deoxy-d-glucose ([¹⁴C]FDG) under treatment with the above reagents in vitro and in vivo were examined.ResultsStrong correlation was observed between fluorescence intensity and total cell number with or without cytotoxic treatment. The uptake of [³H]FLT and [¹⁴C]FDG decreased rapidly after the initiation of treatment with actinomycin D or cycloheximide. When treated with pemetrexed, the uptake of [³H]FLT temporarily increased. The cells formed subcutaneous and orthotopic tumors, with fluorescence intensity correlating with tumor volume. The correlation was sustained under pemetrexed treatment. The uptake of [³H]FLT in vivo increased significantly early after pemetrexed treatment.ConclusionFluorescence imaging could be used to semiquantitatively monitor tumor size, whereas PET could be used to monitor tumor response to therapeutic treatments, and especially, FLT might be a good marker of the response to anti-folate chemotherapeutics.     

Changes in 18F-fluorothymidine and 18F-fluorodeoxyglucose positron emission tomography imaging in patients with head and neck cancer treated with chemoradiotherapy

Objective

This study compared change of ¹⁸F-fluorothymidine (FLT) uptake with that of ¹⁸F-fluorodeoxyglucose (FDG) in head and neck squamous cell cancer (HNSCC) patients during and after treatment and evaluated the utility for early monitoring of response to chemoradiotherapy.

Methods

Thirty patients with newly diagnosed HNSCCs treated with concurrent chemoradiotherapy underwent FLT and FDG PET in pre-treatment (PET1), mid-treatment (PET2) and post-treatment (PET3) stages. The PET images were evaluated quantitatively using maximum standardized uptake values (SUVs). Ratios between SUVs at PET2 and PET3 were also calculated.

Results

According to the SUVs, no significant differences were found with primary site location, cellular differentiation and T category in all PET scans. About a 78 % median decrease in FLT SUV was observed at the total dose (TD) of 30 Gy and no apparent change was observed thereafter. About a 40 % decrease in FDG SUV was observed at TD 30 Gy and significant decreases were then found at the 4- and 6-week time points after the therapy. FLT PET demonstrated no recurrence regions in patients with a PET3/PET2 ratio of <1.5. In comparison, FLT SUVs in PET3 with recurrence were increased more than three times. However, no significant difference was found between the values with recurrence and those with no recurrence in FDG PET.

Conclusion

FLT PET signal change preceded FDG PET change and the increase of FLT uptake after the therapy can imply recurrence or a residual tumor. FLT PET seems promising for early evaluation of chemoradiation effects in HNSCCs.     

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

Purpose

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′-[¹⁸F]fluorothymidine (FLT) positron emission tomography (PET) and [¹⁸F]fluorodeoxyglucose (FDG) PET/CT in patients with suspected pancreatic masses scheduled to undergo surgery.

Methods

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.

Results

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?Conclusion 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.     

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

Abstract The aim of this study was to evaluate the feasibility of using [¹⁸F] 3-deoxy-3-fluorothymidine (FLT) positron emission tomography (PET) for the diagnosis and grading of brain tumors.Methods The patient population comprised 26 patients (15 males, 11 females) with brain tumors (n=18) or nontumorous lesions (n=8). 2-[¹⁸F]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.Results 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).Conclusion 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.     

Correlation of <Superscript>18</Superscript>F-FLT and <Superscript>18</Superscript>F-FDG uptake on PET with Ki-67 immunohistochemistry in non-small cell lung cancer

Purpose The nucleoside analogue 3′-deoxy-3′-¹⁸F-fluorothymidine (FLT) has recently been introduced for imaging cell proliferation with positron emission tomography (PET). We prospectively evaluated whether FLT uptake reflects proliferative activity as indicated by the Ki-67 index in non-small cell lung cancer (NSCLC), in comparison with 2-deoxy-2-¹⁸F-fluoro-D-glucose (FDG). Methods A total of 18 patients with newly diagnosed NSCLC were examined with both FLT PET and FDG PET. PET imaging was performed at 60 min after each radiotracer injection. Tumour lesions were identified as areas of focally increased uptake, exceeding background uptake in the lungs. For semi-quantitative analysis, the maximum standardised uptake value (SUV) was calculated. Proliferative activity as indicated by the Ki-67 index was estimated in tissue specimens. Immunohistochemical findings were correlated with SUVs. Results The sensitivity of FLT and FDG PET for the detection of lung cancer was 72% and 89%, respectively. Four of the five false-negative FLT PET findings occurred in bronchiolo-alveolar carcinoma. The mean FLT SUV was significantly lower than the mean FDG SUV. A significant correlation was observed between FLT SUV and Ki-67 index (r = 0.77; p < 0.0002) and for FDG SUV (r = 0.81; p < 0.0001). Conclusion The results of this preliminary study suggest that, compared with FDG, FLT may be less sensitive for primary staging in patients with NSCLC. Although FLT uptake correlated significantly with proliferative activity in NSCLC, the correlation was not better than that for FDG uptake.     

[<Superscript>18</Superscript>F]FLT-PET in oncology: current status and opportunities

In recent years, [¹⁸F]-fluoro-3-deoxy-3-L-fluorothymidine ([¹⁸F]FLT) has been developed as a proliferation tracer. Imaging and measurement of proliferation with PET could provide us with a non-invasive staging tool and a tool to monitor the response to anticancer treatment. In this review, the basis of [¹⁸F]FLT as a proliferation tracer is discussed. Furthermore, an overview of the current status of [¹⁸F]FLT-PET research is given. The results of this research show that although [¹⁸F]FLT is a tracer that visualises cellular proliferation, it also has certain limitations. In comparison with the most widely used PET tracer, [¹⁸F]FDG, [¹⁸F]FLT uptake is lower in most cases. Furthermore, [¹⁸F]FLT uptake does not always reflect the tumour cell proliferation rate, for example during or shortly after certain chemotherapy regimens. The opportunities provided by, and the limitations of, [¹⁸F]FLT as a proliferation tracer are addressed in this review, and directions are given for further research, taking into account the strong and weak points of the new tracer.     

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

Purpose 3′-Deoxy-3′-[¹⁸F]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. Methods 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 [¹⁸F]FLT or [¹⁸F]fluorodeoxyglucose (FDG) before and after radiation treatment, and tumor growth was assessed over 9 days. Results 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. Conclusion 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.     

[<Superscript>18</Superscript>F]FLT PET for diagnosis and staging of thoracic tumours

The nucleoside analogue 3'-deoxy-3'-[¹⁸F]fluorothymidine (FLT) has been introduced for imaging of tumour cell proliferation by positron emission tomography (PET). This study evaluated the use of FLT in patients with thoracic tumours prior to treatment. Whole-body FLT PET was performed in 16 patients with 18 tumours [17 thoracic tumours (nine non-small cell lung cancers, five oesophageal carcinomas, two sarcomas, one Hodgkin's lymphoma) and one renal carcinoma] before treatment. Fluorine-18 fluorodeoxyglucose (FDG) PET was performed for comparison except in those patients with oesophageal carcinoma. For semi-quantitative analysis, the average and maximum standardised uptake values (avgSUV and maxSUV, respectively) (FLT, 114±20 min p.i.; FDG, 87±8 min p.i.; 50% isocontour region of interest) was calculated. All 17 thoracic tumours and 19/20 metastases revealed significant FLT accumulation, resulting in easy delineation from surrounding tissue. The additional small grade 1 renal carcinoma was not detected with either FLT or FDG. In most lung tumours (avgSUV 1.5–8.2) and metastases, FLT showed intense uptake. However, one of two spinal bone metastases was missed owing to the high physiological FLT uptake in the surrounding bone marrow. Oesophageal carcinoma primaries (avgSUV 2.7–10.0) and occasional metastases showed particularly favourable tumour/non-tumour contrast. Compared with FDG, tumour uptake of FLT was lower (avgSUV, P=0.0006; maxSUV, P=0.0001), with a significant linear correlation (avgSUV, r²=0.45; maxSUV, r²=0.49) between FLT and FDG. It is concluded that FLT PET accurately visualises thoracic tumour lesions. In the liver and the bone marrow, high physiological FLT uptake hampers detection of metastases. On the other hand, FLT may be favourable for imaging of brain metastases owing to the low physiological uptake.     

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

IntroductionThe 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′-[¹⁸F]fluoro-3′-deoxythymidine (FLT) and 2-deoxy-2-[¹⁸F]fluoro-D-glucose (FDG) for assessment of tumor response to the multi-targeted Aurora B kinase inhibitor, TAK-901.MethodsBalb/c nude mice bearing HCT116 colorectal xenografts were treated with up to 30 mg/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.ResultsTumor growth was inhibited by 60% on day 12 of 30 mg/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.ConclusionsFLT-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.Advances in knowledge and implications for patient careThis 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.     

Comparison of <Superscript>18</Superscript>F-FLT PET and <Superscript>18</Superscript>F-FDG PET for preoperative staging in non-small cell lung cancer

Purpose The nucleoside analog 3′-deoxy-3′-¹⁸F-fluorothymidine (FLT) has been introduced for imaging cell proliferation with positron emission tomography (PET). We prospectively compared the diagnostic efficacy of FLT PET with that of 2-deoxy-2-¹⁸F-fluoro-d-glucose (FDG) PET for the preoperative nodal and distant metastatic staging of non-small cell lung cancer (NSCLC). Methods A total of 34 patients with NSCLC underwent FLT PET and FDG PET. PET imaging was performed at 60 min after each radiotracer injection. The PET images were evaluated qualitatively for regions of focally increased metabolism. For visualized primary tumors, the maximum standardized uptake value (SUV) was calculated. Nodal stages were determined by using the American Joint Committee on Cancer staging system and surgical and histologic findings reference standards. Results For the depiction of primary tumor, sensitivity of FLT PET was 67%, compared with 94% for FDG PET (P = 0.005). Sensitivity, specificity, positive predictive value, negative predictive value, and accuracy for lymph node staging on a per-patient basis were 57, 93, 67, 89, and 85%, respectively, with FLT PET and 57, 78, 36, 91, and 74%, respectively, with FDG PET (P > 0.1 for all comparisons). Two of the three distant metastases were detected with FLT and FDG PET. Conclusion In NSCLC, FLT PET showed better (although not statistically significant) specificity, positive predictive value and accuracy for N staging on a per-patient basis than FDG PET. However, FDG PET was found to have higher sensitivity for depiction of primary tumor than FLT PET.     

Initial evaluation of <Superscript>18</Superscript>F-fluorothymidine (FLT) PET/CT scanning for primary pancreatic cancer

Purpose The aim of this study was to evaluate the potential of ¹⁸F-fluorothymidine (FLT) PET/CT for imaging pancreatic adenocarcinoma. Methods This was a pilot study of five patients (four males, one female) with newly diagnosed and previously untreated pancreatic adenocarcinoma. Patients underwent FLT PET/CT, ¹⁸F-fluorodeoxyglucose (FDG) PET/CT, and contrast-enhanced CT scanning before treatment. The presence of cancer was confirmed by histopathological analysis at the time of scanning in all five patients. The degree of FLT and FDG uptake at the primary tumor site was assessed using visual interpretation and semi-quantitative SUV analyses. Results The primary tumor size ranged from 2.5×2.8 cm to 3.5 × 7.0 cm. The SUV of FLT uptake within the primary tumor ranged from 2.1 to 3.1. Using visual interpretation, the primary cancer could be detected from background activity in two of five patients (40%) on FLT PET/CT. By comparison, FDG uptake was higher in each patient with a SUV range of 3.4 to 10.8, and the primary cancer could be detected from background in all five patients (100%). Conclusions In this pilot study of five patients with primary pancreatic adenocarcinoma, FLT PET/CT scanning showed poor lesion detectability and relatively low levels of radiotracer uptake in the primary tumor.     

Detection of gastric cancer using <Superscript>18</Superscript>F-FLT PET: comparison with <Superscript>18</Superscript>F-FDG PET

Purpose  We prospectively investigated the feasibility of 3′-deoxy-3′-¹⁸F-fluorothymidine (FLT) positron emission tomography (PET) for the detection of gastric cancer, in comparison with 2-deoxy-2-¹⁸F-fluoro-d-glucose (FDG) PET, and determined the degree of correlation between the two radiotracers and proliferative activity as indicated by Ki-67 index. Methods  A total of 21 patients with newly diagnosed advanced gastric cancer were examined with FLT PET and FDG PET. Tumour lesions were identified as areas of focally increased uptake, exceeding that of surrounding normal tissue. For semiquantitative analysis, the maximal standardized uptake value (SUV) was calculated. Results  For detection of advanced gastric cancer, the sensitivities of FLT PET and FDG PET were 95.2% and 95.0%, respectively. The mean (±SD) SUV for FLT (7.0 ± 3.3) was significantly lower than that for FDG (9.4 ± 6.3 p < 0.05). The mean FLT SUV and FDG SUV in nonintestinal tumours were higher than in intestinal tumours, although the difference was not statistically significant. The mean (±SD) FLT SUV in poorly differentiated tumours (8.5 ± 3.5) was significantly higher than that in well and moderately differentiated tumours (5.3 ± 2.1; p < 0.04). The mean FDG SUV in poorly differentiated tumours was higher than in well and moderately differentiated tumours, although the difference was not statistically significant. There was no significant correlation between Ki-67 index and either FLT SUV or FDG SUV. Conclusion  FLT PET showed as high a sensitivity as FDG PET for the detection of gastric cancer, although uptake of FLT in gastric cancer was significantly lower than that of FDG.     

Compliance with PET acquisition protocols for therapeutic monitoring of erlotinib therapy in an international trial for patients with non-small cell lung cancer

Purpose

The Response Evaluation Criteria in Solid Tumors (RECIST) are widely used but have recognized limitations. Molecular imaging assessments, including changes in ¹⁸F-deoxyglucose (FDG) or ¹⁸F-deoxythymidine (FLT) uptake by positron emission tomography (PET), may provide earlier, more robust evaluation of treatment efficacy.

Methods

A prospective trial evaluated on-treatment changes in FDG and FLT PET imaging among patients with relapsed or recurrent non-small cell lung cancer treated with erlotinib to assess the relationship between PET-evaluated response and clinical outcomes. We describe an audit of compliance with the study imaging charter, to establish the feasibility of achieving methodological consistency in a multicentre setting.

Results

Patients underwent PET scans at baseline and approximately day 14 and day 56 of treatment (n?=?73, 66 and 51 studies, and n?=?73, 63 and 50 studies for FDG PET and FLT PET, respectively). Blood glucose levels were within the target range for all FDG PET scans. Charter-specified uptake times were achieved in 86% (63/73) and 89% (65/73) of baseline FDG and FLT scans, respectively. On-treatment scans were less consistent: 72% (84/117) and 68% (77/113), respectively, achieved the target of ±5?min of baseline uptake time. However, 96% (112/117) and 94% (106/113) of FDG and FLT PET studies, respectively, were within ±15?min. Bland-Altman analysis of intra-individual hepatic average standardized uptake value (SUV_ave), to assess reproducibility, showed only a small difference in physiological uptake (?0.006?±?0.224 in 118 follow-up FDG scans and 0.09?±?0.81 in 111 follow-up FLT scans).

Conclusion

It is possible to achieve high reproducibility of scan acquisition methodology, provided that strict imaging compliance guidelines are mandated in the study protocol.     

PET with [18F]fluorothymidine for imaging of primary breast cancer: a pilot study

The aim of this study was to evaluate the use of [¹⁸F]fluorothymidine (FLT) as a positron emission tomography (PET) tracer for the diagnosis of breast cancer. To this end, 12 patients with 14 primary breast cancer lesions (T2–T4) were studied by FLT-PET. For comparison, [¹⁸F]fluorodeoxyglucose (FDG) PET scans were performed in six patients. Thirteen of the 14 primary tumours demonstrated focally increased FLT uptake (SUV_mean=3.4±1.1). Seven out of eight patients with histologically proven axillary lymph node metastases showed focally increased FLT uptake in the corresponding areas (SUV_mean=2.4±1.2). The lowest SUV (mean =0.7) was observed in one of two inflammatory cancers. The contrast between primary tumours or metastases and surrounding tissue was high in most cases. In direct comparison to FDG-PET, the SUVs of primary tumours (5/6) and axillary lymph node metastases (3/4) were lower in FLT-PET (SUV_FLT: 3.2 vs SUV_FDG: 4.7 in primary tumours and SUV_FLT: 2.9 vs SUV_FDG: 4.6 in lymph node metastases). Since FLT uptake in surrounding breast tissue was also lower, tumour contrast was comparable to that with FDG. It is of note that normal FLT uptake was very low in the mediastinum, resulting in a higher tumour-to-mediastinum ratio as compared to FDG (P=0.03). FLT-PET is suitable for the diagnosis of primary breast cancer and locoregional metastases. High image contrast may facilitate the detection of small foci, especially in the mediastinum.     

3′-Deoxy-3′-[18F]fluorothymidine (FLT) uptake in breast cancer cells as a measure of proliferation after doxorubicin and docetaxel treatment

IntroductionThe nucleoside analogue [¹⁸F]fluorothymidine (FLT) has been designed as a marker of cell proliferation that can be imaged in vivo by positron emission tomography. Clinical pilot studies have demonstrated decreasing FLT uptake following antiproliferative chemotherapy of breast cancer. However, the significance of posttreatment FLT uptake has not been evaluated at the cell level. The aim of this study was to investigate whether FLT uptake detects proliferation inhibition induced by docetaxel or doxorubicin treatment in an in vitro breast cancer model.MethodsBreast cancer cells (MCF-7) were treated with docetaxel or doxorubicin for 24 h at drug doses inducing 25–99% inhibition of clonogenic survival (IC₂₅ to IC₉₉). Cellular FLT uptake was estimated at 4 h and at 1, 3 and 5 days interval from chemotherapy. [³H]Thymidine incorporation and S-phase fraction were measured for comparison. Analysis of variance and the Bland–Altman difference plot were employed for statistical analysis.ResultsAfter treatment, FLT uptake was declined in dependence of the proliferation inhibition mediated by both chemotherapeutic agents (all P<.0001). The decrease of FLT was greater after doxorubicin treatment than after the corresponding docetaxel dose. With doxorubicin (IC₉₉), FLT accumulation was reduced by 70% as early as 4 h after treatment. FLT uptake was closely correlated to [³H]thymidine incorporation and S-phase fraction (r=.84 to .93).ConclusionsRight after docetaxel or doxorubicin treatment, FLT uptake corresponds to the reduction of tumor cell proliferation induced. [¹⁸F]FLT appears promising for monitoring chemosensitivity in breast cancer.     

Predictive value of early and late residual 18F-fluorodeoxyglucose and 18F-fluorothymidine uptake using different SUV measurements in patients with non-small-cell lung cancer treated with erlotinib

Purpose

To evaluate the predictive value of early and late residual ¹⁸F-fluorodeoxyglucose (FDG) and ¹⁸F-fluorothymidine (FLT) uptake using different SUV measurements in PET in patients with advanced non-small-cell lung cancer (NSCLC) treated with erlotinib.

Methods

We retrospectively reviewed data from 30 patients with untreated stage IV NSCLC who had undergone a combined FDG PET and FLT PET scan at 1?week (early) and 6?weeks (late) after the start of erlotinib treatment. Early and late residual FDG and FLT uptake were measured in up to five lesions per scan with different quantitative standardized uptake values (SUV_max, SUV_2Dpeak, SUV_3Dpeak, SUV₅₀, SUV_A50, SUV_A41) and compared with short-term outcome (progression vs. nonprogression after 6?weeks of erlotinib treatment). Receiver-operating characteristics (ROC) curve analysis was used to determine the optimal cut-off value for detecting nonprogression after 6?weeks. Kaplan-Meier analysis and the log-rank test were used to evaluate the association between residual uptake and progression-free survival (PFS).

Results

Nonprogression after 6?weeks was associated with a significantly lower early and late residual FDG uptake, measured with different quantitative parameters. In contrast, nonprogression after 6?weeks was not associated with early and late residual FLT uptake. Furthermore, patients with a lower early residual FDG uptake measured in terms of SUV_max and SUV_2Dpeak had a significantly prolonged PFS (282?days vs. 118?days; p?=?0.022) than patients with higher values. Similarly, lower late residual FDG uptake and early residual FLT uptake measured in terms of SUV_3Dpeak, SUV_A50 and SUV_A41, and late FLT uptake measured in terms of SUV_3Dpeak and SUV_A50 was associated with an improved PFS.

Conclusion

Early and late residual FDG uptake, measured using different quantitative SUV parameters, are predictive factors for short-term outcome in patients with advanced NSCLC treated with erlotinib. Additionally, low residual FDG and FLT uptake early and late in the course of erlotinib treatment is associated with improved PFS.     

Diagnostic performance of 18F-fluorothymidine PET/CT for primary colorectal cancer and its lymph node metastasis: comparison with 18F-fluorodeoxyglucose PET/CT

Purpose

To examine the diagnostic performance of ¹⁸F-fluorothymidine (FLT) PET/CT in primary and metastatic lymph node colorectal cancer foci in comparison with ¹⁸F-fluorodeoxyglucose (FDG) PET/CT.

Methods

The study population comprised 28 patients with 30 newly diagnosed colorectal cancers who underwent surgical resection of the primary lesion and regional lymph nodes after both FLT and FDG PET/CT. The associations between SUVmax levels and pathological factors were evaluated using the Mann-Whitney U or Kruskal-Wallis test. Differences in diagnostic indexes for detecting nodal metastasis between the two tracers were estimated using the McNemar exact or χ ² test.

Results

All 30 primary cancers (43.0?±?20.0 mm, range 14 – 85 mm) were visualized by both tracers, but none of the FLT SUVmax values exceeded the FDG SUVmax values in any of the primary cancers (6.6?±?2.4 vs. 13.6?±?5.8, p?<?0.001). The sensitivity, specificity and accuracy for detecting nodal metastasis were 41 % (15/37), 98.8 % (493/499) and 94.8 % (508/536) for FDG PET/CT, and 32 % (12/37), 98.8 % (493/499) and 94.2 % (505/536) for FLT PET/CT, respectively. The sensitivity (p?=?0.45), specificity (p?=?0.68) and accuracy (p?=?0.58) were not different between the tracers. Nodal uptake of FLT and FDG was discordant in 7 (19 %) of 37 metastatic nodes. There were ten concordant true-positive nodes of which six showed higher FDG SUVmax and four showed higher FLT SUVmax, but the difference between FDG and FLT SUVmax was not significant (5.56?±?3.55 and 3.62?±?1.45, respectively; p?=?0.22).

Conclusion

FLT has the same potential as FDG in PET/CT for the diagnosis of primary and nodal foci of colorectal cancer despite significantly lower FLT uptake in primary foci.