Tumor Uptake of 64Cu-DOTA-Trastuzumab in Patients with Metastatic Breast Cancer.
ABSTRACT: The goal of this study was to characterize the relationship between tumor uptake of 64Cu-DOTA-trastuzumab as measured by PET/CT and standard, immunohistochemistry (IHC)-based, histopathologic classification of human epidermal growth factor receptor 2 (HER2) status in women with metastatic breast cancer (MBC). Methods: Women with biopsy-confirmed MBC and not given trastuzumab for 2 mo or more underwent complete staging, including 18F-FDG PET/CT. Patients were classified as HER2-positive (HER2+) or -negative (HER2-) based on fluorescence in situ hybridization (FISH)-supplemented immunohistochemistry of biopsied tumor tissue. Eighteen patients underwent 64Cu-DOTA-trastuzumab injection, preceded in 16 cases by trastuzumab infusion (45 mg). PET/CT was performed 21-25 (day 1) and 47-49 (day 2) h after 64Cu-DOTA-trastuzumab injection. Radiolabel uptake in prominent lesions was measured as SUVmax Average intrapatient SUVmax (max>pt) was compared between HER2+ and HER2- patients. Results: Eleven women were HER2+ (8 immunohistochemistry 3+; 3 immunohistochemistry 2+/FISH amplified), whereas 7 were HER2- (3 immunohistochemistry 2+/FISH nonamplified; 4 immunohistochemistry 1+). Median max>pt for day 1 and day 2 was 6.6 and 6.8 g/mL for HER 2+ and 3.7 and 4.3 g/mL for HER2- patients (P < 0.005 either day). The distributions of max>pt overlapped between the 2 groups, and interpatient variability was greater for HER2+ than HER2- disease (P < 0.005 and 0.001, respectively, on days 1 and 2). Conclusion: By 1 d after injection, uptake of 64Cu-DOTA-trastuzumab in MBC is strongly associated with patient HER2 status and is indicative of binding to HER2. The variability within and among HER2+ patients, as well as the overlap between the HER2+ and HER2- groups, suggests a role for 64Cu-DOTA-trastuzumab PET/CT in optimizing treatments that include trastuzumab.
Project description:The human epidermal growth factor receptor-2 (HER2) is overexpressed in 20-30% of all breast cancer cases, leading to increased cell proliferation, growth and migration. The monoclonal antibody, trastuzumab, binds to HER2 and is used for treatment of HER2-positive breast cancer. Trastuzumab has previously been labelled with copper-64 by conjugation of a 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) chelator. The aim of this study was to optimise the (64) Cu-labelling of DOTA-trastuzumab and as the first to produce and compare with its 1,4,7-triazacyclononane, 1-glutaric acid-5,7 acetic acid (NODAGA) analogue in a preliminary HER2 tumour mouse model. The chelators were conjugated to trastuzumab using the activated esters DOTA mono-N-hydroxysuccinimide (NHS) and NODAGA-NHS. (64) Cu-labelling of DOTA-trastuzumab was studied by varying the amount of DOTA-trastuzumab used, reaction temperature and time. Full (64) Cu incorporation could be achieved using a minimum of 10-µg DOTA-trastuzumab, but the fastest labelling was obtained after 15?min at room temperature using 25?µg of DOTA-trastuzumab. In comparison, 80% incorporation was achieved for (64) Cu-labelling of NODAGA-trastuzumab. Both [(64) Cu]DOTA-trastuzumab and [(64) Cu]NODAGA-trastuzumab were produced after purification with radiochemical purities of >97%. The tracers were injected into mice with HER2 expressing tumours. The mice were imaged by positron emission tomography and showed high tumour uptake of 3-9% ID/g for both tracers.
Project description:Women with human epidermal growth factor receptor 2 (HER2)-positive breast cancer are candidates for treatment with the anti-HER2 antibody trastuzumab. Assessment of HER2 status in recurrent disease is usually made by core needle biopsy of a single lesion, which may not represent the larger tumor mass or other sites of disease. Our long-range goal is to develop PET of radiolabeled trastuzumab for systemically assessing tumor HER2 expression and identifying appropriate use of anti-HER2 therapies. The purpose of this study was to evaluate PET/CT of (64)Cu-DOTA-trastuzumab for detecting and measuring tumor uptake of trastuzumab in patients with HER2-positive metastatic breast cancer.Eight women with biopsy-confirmed HER2-positive metastatic breast cancer and no anti-HER2 therapy for 4 mo or longer underwent complete staging, including (18)F-FDG PET/CT. For 6 of the 8 patients, (64)Cu-DOTA-trastuzumab injection (364-512 MBq, 5 mg of trastuzumab) was preceded by trastuzumab infusion (45 mg). PET/CT (PET scan duration 1 h) was performed 21-25 (day 1) and 47-49 (day 2) h after (64)Cu-DOTA-trastuzumab injection. Scan fields of view were chosen on the basis of (18)F-FDG PET/CT. Tumor detection sensitivity and uptake analyses were limited to lesions identifiable on CT; lesions visualized relative to adjacent tissue on PET were considered PET-positive. Radiolabel uptake in prominent lesions was measured as maximum single-voxel standardized uptake value (SUVmax).Liver uptake of (64)Cu was reduced approximately 75% with the 45-mg trastuzumab predose, without significant effect on tumor uptake. The study included 89 CT-positive lesions. Detection sensitivity was 77%, 89%, and 93% for day 1, day 2, and (18)F-FDG, respectively. On average, tumor uptake was similar for (64)Cu-DOTA-trastuzumab and (18)F-FDG (SUVmax and range, 8.1 and 3.0-22.5 for day 1 [n = 48]; 8.9 and 0.9-28.9 for day 2 [n = 38]; 9.7 and 3.3-25.4 for (18)F-FDG [n = 56]), but same-lesion SUVmax was not correlated between the 2 radiotracers. No toxicities were observed, and estimated radiation dose from (64)Cu-DOTA-trastuzumab was similar to (18)F-FDG.(64)Cu-DOTA-trastuzumab visualizes HER2-positive metastatic breast cancer with high sensitivity and is effective in surveying disseminated disease. A 45-mg trastuzumab predose provides a (64)Cu-DOTA-trastuzumab biodistribution favorable for tumor imaging. (64)Cu-DOTA-trastuzumab PET/CT warrants further evaluation for assessing tumor HER2 expression and individualizing treatments that include trastuzumab.
Project description:<h4>Background</h4>Epidermal growth factor receptors (EGFR) are overexpressed on many head and neck squamous cell carcinoma (HNSCC). Radioimmunotherapy (RIT) with F(ab')<sub>2</sub> of the anti-EGFR monoclonal antibody panitumumab labeled with the β-particle emitter, <sup>177</sup>Lu may be a promising treatment for HNSCC. Our aim was to assess the feasibility of a theranostic strategy that combines positron emission tomography (PET) with [<sup>64</sup>Cu]Cu-DOTA-panitumumab F(ab')<sub>2</sub> to image HNSCC and predict the radiation equivalent doses to the tumour and normal organs from RIT with [<sup>177</sup>Lu]Lu-DOTA-panitumumab F(ab')<sub>2</sub>.<h4>Results</h4>Panitumumab F(ab')<sub>2</sub> were conjugated to DOTA and complexed to <sup>64</sup>Cu or <sup>177</sup>Lu in high radiochemical purity (95.6 ± 2.1% and 96.7 ± 3.5%, respectively) and exhibited high affinity EGFR binding (K<sub>d</sub> = 2.9 ± 0.7 × 10<sup>- 9</sup> mol/L). Biodistribution (BOD) studies at 6, 24 or 48 h post-injection (p.i.) of [<sup>64</sup>Cu]Cu-DOTA-panitumumab F(ab')<sub>2</sub> (5.5-14.0 MBq; 50 μg) or [<sup>177</sup>Lu]Lu-DOTA-panitumumab F(ab')<sub>2</sub> (6.5 MBq; 50 μg) in NRG mice with s.c. HNSCC patient-derived xenografts (PDX) overall showed no significant differences in tumour uptake but modest differences in normal organ uptake were noted at certain time points. Tumours were imaged by microPET/CT with [<sup>64</sup>Cu]Cu-DOTA-panitumumab F(ab')<sub>2</sub> or microSPECT/CT with [<sup>177</sup>Lu]Lu-DOTA-panitumumab F(ab')<sub>2</sub> but not with irrelevant [<sup>177</sup>Lu]Lu-DOTA-trastuzumab F(ab')<sub>2</sub>. Tumour uptake at 24 h p.i. of [<sup>64</sup>Cu]Cu-DOTA-panitumumab F(ab')<sub>2</sub> [14.9 ± 1.1% injected dose/gram (%ID/g) and [<sup>177</sup>Lu]Lu-DOTA-panitumumab F(ab')<sub>2</sub> (18.0 ± 0.4%ID/g) were significantly higher (P < 0.05) than [<sup>177</sup>Lu]Lu-DOTA-trastuzumab F(ab')<sub>2</sub> (2.6 ± 0.5%ID/g), demonstrating EGFR-mediated tumour uptake. There were no significant differences in the radiation equivalent doses in the tumour and most normal organs estimated for [<sup>177</sup>Lu]Lu-DOTA-panitumumab F(ab')<sub>2</sub> based on the BOD of [<sup>64</sup>Cu]Cu-DOTA-panitumumab F(ab')<sub>2</sub> compared to those estimated directly from the BOD of [<sup>177</sup>Lu]Lu-DOTA-panitumumab F(ab')<sub>2</sub> except for the liver and whole body which were modestly underestimated by [<sup>64</sup>Cu]Cu-DOTA-panitumumab F(ab')<sub>2</sub>. Region-of-interest (ROI) analysis of microPET/CT images provided dose estimates for the tumour and liver that were not significantly different for the two radioimmunoconjugates. Human doses from administration of [<sup>177</sup>Lu]Lu-DOTA-panitumumab F(ab')<sub>2</sub> predicted that a 2 cm diameter HNSCC tumour in a patient would receive 1.1-1.5 mSv/MBq and the whole body dose would be 0.15-0.22 mSv/MBq.<h4>Conclusion</h4>A PET theranostic strategy combining [<sup>64</sup>Cu]Cu-DOTA-panitumumab F(ab')<sub>2</sub> to image HNSCC tumours and predict the equivalent radiation doses in the tumour and normal organs from RIT with [<sup>177</sup>Lu]Lu-DOTA-panitumumab F(ab')<sub>2</sub> is feasible. RIT with [<sup>177</sup>Lu]Lu-DOTA-panitumumab F(ab')<sub>2</sub> may be a promising approach to treatment of HNSCC due to frequent overexpression of EGFR.
Project description:<h4>Background and aims</h4>The objective of this study was to investigate the effects of semaglutide, a long acting glucagon-like peptide-1 receptor agonist, on atherosclerotic inflammation and calcification using a multimodality positron emission tomography and computed tomography (PET/CT) approach.<h4>Methods</h4>Atherosclerotic New Zealand White rabbits were randomized to an intervention- (n = 12) or placebo group (n = 11) receiving either semaglutide or saline-placebo. PET/CT imaging was done before and after 16-weeks of intervention. Three different radiotracers were used: [<sup>64</sup>Cu]Cu-DOTATATE for imaging of activated macrophages, [<sup>18</sup>F]FDG imaging cellular metabolism and [<sup>18</sup>F]NaF PET visualizing micro-calcifications. Tracer uptake was quantified by maximum standardized uptake value (SUV<sub>max</sub>) and target-to-background-ratio (TBR<sub>max</sub>). Animals were euthanized for autoradiographic imaging and histological analyses.<h4>Results</h4>A reduction in activated macrophage tracer-uptake was observed in the semaglutide group (SUV<sub>max</sub>: p = 0.001 and TBR<sub>max</sub>: p = 0.029). When imaging cellular metabolism, an attenuation of SUV<sub>max</sub> and TBR<sub>max</sub> was observed in the semaglutide group (p = 0.034 and p = 0.044). We found no difference in uptake of the micro-calcification tracer between the two groups (SUV<sub>max</sub>: p = 0.62 and TBR<sub>max</sub>: p = 0.36). Values of macrophage density in the vessel wall were significantly correlated with SUV<sub>max</sub> values of the activated macrophage (r = 0.54, p = 0.0086) and cellular metabolism tracers (r = 0.51, p = 0.013).<h4>Conclusions</h4>Semaglutide decreased vascular uptake of tracers imaging activated macrophages and cellular metabolism but not micro-calcifications compared to a saline placebo. This supports the hypothesis that semaglutide reduces atherosclerotic inflammation by means of decreased activated macrophage activity.
Project description:<h4>Objective</h4>To identify the impact of segmentation methods and intensity discretization on radiomic features (RFs) extraction from <sup>68</sup>Ga-DOTA-TOC PET images in patients with neuroendocrine tumors.<h4>Methods</h4>Forty-nine patients were retrospectively analyzed. Tumor contouring was performed manually by four different operators and with a semi-automatic edge-based segmentation (SAEB) algorithm. Three SUV<sub>max</sub> fixed thresholds (20, 30, 40%) were applied. Fifty-one RFs were extracted applying two different intensity rescale factors for gray-level discretization: one absolute (AR60 = SUV from 0 to 60) and one relative (RR = min-max of the VOI SUV). Dice similarity coefficient (DSC) was calculated to quantify segmentation agreement between different segmentation methods. The impact of segmentation and discretization on RFs was assessed by intra-class correlation coefficients (ICC) and the coefficient of variance (COV<sup>L</sup>). The RFs' correlation with volume and SUV<sub>max</sub> was analyzed by calculating Pearson's correlation coefficients.<h4>Results</h4>DSC mean value was 0.75 ± 0.11 (0.45-0.92) between SAEB and operators and 0.78 ± 0.09 (0.36-0.97), among the four manual segmentations. The study showed high robustness (ICC >?0.9): (a) in 64.7% of RFs for segmentation methods using AR60, improved by applying SUV<sub>max</sub> threshold of 40% (86.5%); (b) in 50.9% of RFs for different SUV<sub>max</sub> thresholds using AR60; and (c) in 37% of RFs for discretization settings using different segmentation methods. Several RFs were not correlated with volume and SUV<sub>max</sub>.<h4>Conclusions</h4>RFs robustness to manual segmentation resulted higher in NET <sup>68</sup>Ga-DOTA-TOC images compared to <sup>18</sup>F-FDG PET/CT images. Forty percent SUV<sub>max</sub> thresholds yield superior RFs stability among operators, however leading to a possible loss of biological information. SAEB segmentation appears to be an optimal alternative to manual segmentation, but further validations are needed. Finally, discretization settings highly impacted on RFs robustness and should always be stated.
Project description:<h4>Background</h4>The monocyte chemoattractant protein-1/CCR2 (chemokine receptor 2) axis plays an important role in abdominal aortic aneurysm (AAA) pathogenesis, with effects on disease progression and anatomic stability. We assessed the expression of CCR2 in a rodent model and human tissues, using a targeted positron emission tomography radiotracer (<sup>64</sup>Cu-DOTA-ECL1i).<h4>Methods</h4>AAAs were generated in Sprague-Dawley rats by exposing the infrarenal, intraluminal aorta to PPE (porcine pancreatic elastase) under pressure to induce aneurysmal degeneration. Heat-inactivated PPE was used to generate a sham operative control. Rat AAA rupture was stimulated by the administration of ?-aminopropionitrile, a lysyl oxidase inhibitor. Biodistribution was performed in wild-type rats at 1 hour post tail vein injection of <sup>64</sup>Cu-DOTA-ECL1i. Dynamic positron emission tomography/computed tomography imaging was performed in rats to determine the in vivo distribution of radiotracer.<h4>Results</h4>Biodistribution showed fast renal clearance. The localization of radiotracer uptake in AAA was verified with high-resolution computed tomography. At day 7 post-AAA induction, the radiotracer uptake (standardized uptake value [SUV]=0.91±0.25) was approximately twice that of sham-controls (SUV=0.47±0.10; <i>P</i><0.01). At 14 days post-AAA induction, radiotracer uptake by either group did not significantly change (AAA SUV=0.86±0.17 and sham-control SUV=0.46±0.10), independent of variations in aortic diameter. Competitive CCR2 receptor blocking significantly decreased AAA uptake (SUV=0.42±0.09). Tracer uptake in AAAs that subsequently ruptured (SUV=1.31±0.14; <i>P</i><0.005) demonstrated uptake nearly twice that of nonruptured AAAs (SUV=0.73±0.11). Histopathologic characterization of rat and human AAA tissues obtained from surgery revealed increased expression of CCR2 that was co-localized with CD68<sup>+</sup> macrophages. Ex vivo autoradiography demonstrated specific binding of <sup>64</sup>Cu-DOTA-ECL1i to CCR2 in both rat and human aortic tissues.<h4>Conclusions</h4>CCR2 positron emission tomography is a promising new biomarker for the noninvasive assessment of AAA inflammation that may aid in associated rupture prediction.
Project description:A modular system for the construction of radiometalated antibodies was developed based on the bioorthogonal cycloaddition reaction between 3-(4-benzylamino)-1,2,4,5-tetrazine and the strained dienophile norbornene. The well-characterized, HER2-specific antibody trastuzumab and the positron emitting radioisotopes (64)Cu and (89)Zr were employed as a model system. The antibody was first covalently coupled to norbornene, and this stock of norbornene-modified antibody was then reacted with tetrazines bearing the chelators 1,4,7,10-tetraazacyclo-dodecane-1,4,7,10-tetraacetic acid (DOTA) or desferrioxamine (DFO) and subsequently radiometalated with (64)Cu and (89)Zr, respectively. The modification strategy is simple and robust, and the resultant radiometalated constructs were obtained in high specific activity (2.7-5.3 mCi/mg). For a given initial stoichiometric ratio of norbornene to antibody, the (64)Cu-DOTA- and (89)Zr-DFO-based probes were shown to be nearly identical in terms of stability, the number of chelates per antibody, and immunoreactivity (>93% in all cases). In vivo PET imaging and acute biodistribution experiments revealed significant, specific uptake of the (64)Cu- and (89)Zr-trastuzumab bioconjugates in HER2-positive BT-474 xenografts, with little background uptake in HER2-negative MDA-MB-468 xenografts or other tissues. This modular system-one in which the divergent point is a single covalently modified antibody stock that can be reacted selectively with various chelators-will allow for both greater versatility and more facile cross-comparisons in the development of antibody-based radiopharmaceuticals.
Project description:Trastuzumab (Herceptin®) is an approved immunotherapeutic agent used for the treatment of metastatic breast cancer over-expressing HER2 antigen receptors. The aim of the present work is to standardize the formulation protocol of [<sup>177</sup>Lu]Lu-trastuzumab addressing various reaction parameters, evaluating the efficacy of the radiolabeled product by <i>in vitro</i> investigations, scaling-up the preparation for administration in patients and performing preliminary clinical studies in patients suffering from metastatic breast cancer. Trastuzumab was conjugated with a suitable bi-functional chelating agent namely, <i>p</i>-NCS-benzyl-DOTA. On average 6.15 ± 0.92 <i>p</i>-NCS-benzyl-DOTA molecules were observed to be attached to each trastuzumab moiety. [<sup>177</sup>Lu]Lu-trastuzumab could be prepared with >95% radiochemical purity (% RCP) employing the optimized radiolabeling procedure. <i>In vitro</i> studies revealed the affinity of [<sup>177</sup>Lu]Lu-trastuzumab towards HER2 +ve cancer cell lines as well as against HER2 protein (<i>K</i> <sub>d</sub> = 13.61 nM and 11.36 nM, respectively). The value for percentage immunoreactive fraction (% IRF) for [<sup>177</sup>Lu]Lu-trastuzumab was observed to be 76.92 ± 2.80. Bio-distribution studies in Swiss mice revealed non-specific uptake in the blood, liver, lungs and heart followed by gradual clearance of activity predominantly through the hepatobiliary route. Preliminary clinical studies carried out in 8 cancer patients with immunohistochemically proven HER2 positive metastatic breast cancer revealed preferential localization of [<sup>177</sup>Lu]Lu-trastuzumab in breast cancer lesions, which was in concordance with [<sup>18</sup>F]FDG-PET scans recorded earlier in the same patient indicating the potential of the agent towards radioimmunotheranostic applications.
Project description:Trastuzumab is a monoclonal antibody targeting human epidermal growth factor 2 (HER2), which has been successfully used in the treatment of patients with breast cancer and gastric cancer; however, problems concerning its cardiotoxicity, drug resistance, and unpredictable efficacy still remain. Herein, we constructed novel organic dopamine-melanin nanoparticles (dMNs) as a carrier and then surface-loaded them with trastuzumab to construct a multifunctional nanoprobe named Her-PEG-dMNPs. We used micro-PET/CT and PET/MRI multimodality imaging to evaluate the retention effect of the nanoprobe in HER2 expression in gastric cancer patient-derived xenograft (PDX) mice models after labeling of the radionuclides <sup>64</sup>Cu or <sup>124</sup>I and MRI contrast agent Mn<sup>2+</sup>. The nanoprobes can specifically target the HER2-expressing SKOV-3 cells <i>in vitro</i> (3.61 ± 0.74 vs. 1.24 ± 0.43 for 2 h, <i>P </i>= 0.002). <i>In vivo</i>, micro-PET/CT and PET/MRI showed that the <sup>124</sup>I-labeled nanoprobe had greater contrast and retention effect in PDX models than unloaded dMNPs as carrier (1.63 ± 0.07 vs. 0.90 ± 0.04 at 24 h, <i>P </i>= 0.002), a similarity found in <sup>64</sup>Cu-labeled Her-PEG-dMNPs. Because <sup>124</sup>I has a longer half-life and matches the pharmacokinetics of the nanoparticles, we focused on the further evaluation of <sup>124</sup>I-Her-PEG-dMNPs. Furthermore, immunohistochemistry staining confirmed the overexpression of HER2 in the animal model. This study developed and validated novel HER2-specific multimodality imaging nanoprobes for quantifying HER2 expression in mice. Through the strong retention effect of the tumor site, it can be used for the promotion of monoclonal antibody treatment effect and process monitoring.
Project description:We compared the imaging characteristics and hypoxia selectivity of 4 hypoxia PET radiotracers ((18)F-fluoromisonidazole [(18)F-FMISO], (18)F-flortanidazole [(18)F-HX4], (18)F-fluoroazomycin arabinoside [(18)F-FAZA], and (64)Cu-diacetyl-bis(N4-methylsemicarbazone) [(64)Cu-ATSM]) in a single murine xenograft tumor model condition using small-animal PET imaging and combined ex vivo autoradiography and fluorescence immunohistochemistry.Nude mice bearing SQ20b xenograft tumors were administered 1 of 4 hypoxia PET tracers and images acquired 80-90 min after injection. Frozen sections from excised tumors were then evaluated for tracer distribution using digital autoradiography and compared with histologic markers of tumor hypoxia (pimonidazole, carbonic anydrase 9 [CA9]) and vascular perfusion (Hoechst 33342).The highest tumor uptake was observed with (64)Cu-ATSM (maximum standardized uptake values [SUV(max)], 1.26 ± 0.13) and the lowest with (18)F-FAZA (SUVmax, 0.41 ± 0.24). (18)F-FMISO and (18)F-HX4 had similar intermediate tumor uptake (SUV(max), 0.76 ± 0.38 and 0.65 ± 0.19, respectively). Digital autoradiographs of hypoxia tracer distribution were compared pixel by pixel with images of immunohistochemistry stains. The fluorinated nitroimidazoles all showed radiotracer uptake increasing with pimonidazole and CA9 staining. (64)Cu-ATSM showed the opposite pattern, with highest radiotracer uptake observed in regions with the lowest pimonidazole and CA9 staining.The fluorinated nitroimidazoles showed similar tumor distributions when compared with immunohistochemistry markers of hypoxia. Variations in tumor standardized uptake value and normal tissue distribution may determine the most appropriate clinical setting for each tracer. (64)Cu-ATSM showed the highest tumor accumulation and little renal clearance. However, the lack of correlation between (64)Cu-ATSM distribution and immunohistochemistry hypoxia markers casts some doubt on the hypoxia selectivity of (64)Cu-ATSM.