José A. Richter

Assessment of biliary bicarbonate secretion in humans by positron emission tomography

BACKGROUND & AIMS Positron emission tomography (PET) allows imaging and quantitative analysis of organ functions in basal and stimulated conditions. We have applied this method to the study of biliary bicarbonate secretion in humans. METHODS PET was performed in 5 healthy subjects and 13 patients with hepatobiliary disorders after intravenous injection of NaH11CO3. In each case the study was performed in basal conditions and after secretin stimulation. Positron emission from the hepatic area was scanned, and normalized uptake values for parenchymal and hilar regions were estimated. RESULTS In healthy individuals, the injection of NaH11CO3 resulted in a peak uptake of the label in parenchymal and hilar regions 2-3 minutes after the injection. In both normal and cirrhotic subjects, secretin administration increased bicarbonate uptake in the parenchymal region, followed by accumulation of the label in the perihilar area. Normal basal uptake with absent response to secretin was registered in extrahepatic biliary obstruction and in untreated primary biliary cirrhosis (PBC). The secretin response was present in patients with PBC undergoing treatment with ursodeoxycholic acid. CONCLUSIONS PET allows investigation of biliary bicarbonate secretion in humans. An impaired response to secretin was observed in cholestatic conditions. Preliminary data suggest that ursodeoxycholic acid might improve the response to secretin in PBC.

Dendritic cells delivered inside human carcinomas are sequestered by interleukin-8

In the course of a clinical trial consisting of intratumoral injections of dendritic cells (DCs) transfected to produce interleukin‐12, the use of 111In‐labeled tracing doses of DCs showed that most DCs remained inside tumor tissue, instead of migrating out. In search for factors that could explain this retention, it was found that tumors from patients suffering hepatocellular carcinoma, colorectal or pancreatic cancer were producing IL‐8 and that this chemokine attracted monocyte‐derived dendritic cells that uniformly express both IL‐8 receptors CXCR1 and CXCR2. Accordingly, neutralizing antihuman IL‐8 monoclonal antibodies blocked the chemotactic attraction of DCs by recombinant IL‐8, as well as by the serum of the patients or culture supernatants of human colorectal carcinomas. In addition, tissue culture supernatants of colon carcinoma cells inhibited DC migration induced by MIP‐3β in an IL‐8‐dependent fashion. IL‐8 production in malignant tissue and the responsiveness of DCs to IL‐8 are a likely explanation of the clinical images, which suggest retention of DCs inside human malignant lesions. Impairment of DC migration toward lymphoid tissue could be involved in cancer immune evasion.

Reversion of epigenetically mediated BIM silencing overcomes chemoresistance in Burkitt lymphoma

In Burkitt lymphoma/leukemia (BL), achievement of complete remission with first-line chemotherapy remains a challenging issue, as most patients who respond remain disease-free, whereas those refractory have few options of being rescued with salvage therapies. The mechanisms underlying BL chemoresistance and how it can be circumvented remain undetermined. We previously reported the frequent inactivation of the proapoptotic BIM gene in B-cell lymphomas. Here we show that BIM epigenetic silencing by concurrent promoter hypermethylation and deacetylation occurs frequently in primary BL samples and BL-derived cell lines. Remarkably, patients with BL with hypermethylated BIM presented lower complete remission rate (24% vs 79%; P = .002) and shorter overall survival (P = .007) than those with BIM-expressing lymphomas, indicating that BIM transcriptional repression may mediate tumor chemoresistance. Accordingly, by combining in vitro and in vivo studies of human BL-xenografts grown in immunodeficient RAG2(-/-)γc(-/-) mice and of murine B220(+)IgM(+) B-cell lymphomas generated in Eμ-MYC and Eμ-MYC-BIM(+/-) transgenes, we demonstrate that lymphoma chemoresistance is dictated by BIM gene dosage and is reversible on BIM reactivation by genetic manipulation or after treatment with histone-deacetylase inhibitors. We suggest that the combination of histone-deacetylase inhibitors and high-dose chemotherapy may overcome chemoresistance, achieve durable remission, and improve survival of patients with BL.

Quantitative volumetric analysis of gliomas with sequential MRI and 11C-methionine PET assessment: patterns of integration in therapy planning

PurposeThe aim of the study was to evaluate the volumetric integration patterns of standard MRI and 11C-methionine positron emission tomography (PET) images in the surgery planning of gliomas and their relationship to the histological grade.MethodsWe studied 23 patients with suspected or previously treated glioma who underwent preoperative 11C-methionine PET because MRI was imprecise in defining the surgical target contour. Images were transferred to the treatment planning system, coregistered and fused (BrainLAB). Tumour delineation was performed by 11C-methionine PET thresholding (vPET) and manual segmentation over MRI (vMRI). A 3-D volumetric study was conducted to evaluate the contribution of each modality to tumour target volume. All cases were surgically treated and histological classification was performed according to WHO grades. Additionally, several biopsy samples were taken according to the results derived either from PET or from MRI and analysed separately.ResultsFifteen patients had high-grade tumours [ten glioblastoma multiforme (GBM) and five anaplastic), whereas eight patients had low-grade tumours. Biopsies from areas with high 11C-methionine uptake without correspondence in MRI showed tumour proliferation, including infiltrative zones, distinguishing them from dysplasia and radionecrosis. Two main PET/MRI integration patterns emerged after analysis of volumetric data: pattern vMRI-in-vPET (11/23) and pattern vPET-in-vMRI (9/23). Besides, a possible third pattern with differences in both directions (vMRI-diff-vPET) could also be observed (3/23). There was a statistically significant association between the tumour classification and integration patterns described above (p < 0.001, κ = 0.72). GBM was associated with pattern vMRI-in-vPET (9/10), low-grade with pattern vPET-in-vMRI (7/8) and anaplastic with pattern vMRI-diff-vPET (3/5).ConclusionThe metabolically active tumour volume observed in 11C-methionine PET differs from the volume of MRI by showing areas of infiltrative tumour and distinguishing from non-tumour lesions. Differences in 11C-methionine PET/MRI integration patterns can be assigned to tumour grades according to the WHO classification. This finding may improve tumour delineation and therapy planning for gliomas.

Promising Prospects for 44Sc-/47Sc-Based Theragnostics: Application of 47Sc for Radionuclide Tumor Therapy in Mice

In recent years, 47Sc has attracted attention because of its favorable decay characteristics (half-life, 3.35 d; average energy, 162 keV; Eγ, 159 keV) for therapeutic application and for SPECT imaging. The aim of the present study was to investigate the suitability of 47Sc for radionuclide therapy in a preclinical setting. For this purpose a novel DOTA-folate conjugate (cm10) with an albumin-binding entity was used. Methods: 47Sc was produced via the 46Ca(n,γ)47Ca→β−47Sc nuclear reaction at the high-flux reactor at the Institut Laue-Langevin. Separation of the 47Sc from the target material was performed by a semi-automated process using extraction chromatography and cation exchange chromatography. 47Sc-labeled cm10 was tested on folate receptor–positive KB tumor cells in vitro. Biodistribution and SPECT imaging experiments were performed in KB tumor–bearing mice. Radionuclide therapy was conducted with two groups of mice, which received either 47Sc-cm10 (10 MBq) or only saline. Tumor growth and survival time were compared between the two groups of mice. Results: Irradiation of 46Ca resulted in approximately 1.8 GBq of 47Ca, which subsequently decayed to 47Sc. Separation of 47Sc from 47Ca was obtained with 80% yield in only 10 min. The 47Sc was then available in a small volume (∼500 μL) of an ammonium acetate/HCl (pH 4.5) solution suitable for direct radiolabeling. 47Sc-cm10 was prepared with a radiochemical yield of more than 96% at a specific activity of up to 13 MBq/nmol. In vitro 47Sc-cm10 showed folate receptor–specific binding and uptake into KB tumor cells. In vivo SPECT/CT images allowed the visualization of accumulated radioactivity in KB tumors and in the kidneys. The therapy study showed a significantly delayed tumor growth in mice, which received 47Sc-cm10 (10 MBq, 10 Gy) resulting in a more than 50% increase in survival time, compared with untreated control mice. Conclusion: With this study, we demonstrated the suitability of using 47Sc for therapeutic purposes. On the basis of our recent results obtained with 44Sc-folate, the present work confirms the applicability of 44Sc/47Sc as an excellent matched pair of nuclides for PET imaging and radionuclide therapy.

Pilot Clinical Trial of Type 1 Dendritic Cells Loaded with Autologous Tumor Lysates Combined with GM-CSF, Pegylated IFN, and Cyclophosphamide for Metastatic Cancer Patients

Twenty-four patients with metastatic cancer received two cycles of four daily immunizations with monocyte-derived dendritic cells (DC). DC were incubated with preheated autologous tumor lysate and subsequently with IFN-α, TNF-α, and polyinosinic:polycytidylic acid to attain type 1 maturation. One DC dose was delivered intranodally, under ultrasound control, and the rest intradermally in the opposite thigh. Cyclophosphamide (day −7), GM-CSF (days 1–4), and pegIFN alpha-2a (days 1 and 8) completed each treatment cycle. Pretreatment with cyclophosphamide decreased regulatory T cells to levels observed in healthy subjects both in terms of percentage and in absolute counts in peripheral blood. Treatment induced sustained elevations of IL-12 in serum that correlated with the output of IL-12p70 from cultured DC from each individual. NK activity in peripheral blood was increased and also correlated with the serum concentration of IL-12p70 in each patient. Circulating endothelial cells decreased in 17 of 18 patients, and circulating tumor cells markedly dropped in 6 of 19 cases. IFN-γ–ELISPOT responses to DC plus tumor lysate were observed in 4 of 11 evaluated cases. Tracing DC migration with [111In] scintigraphy showed that intranodal injections reached deeper lymphatic chains in 61% of patients, whereas with intradermal injections a small fraction of injected DC was almost constantly shown to reach draining inguinal lymph nodes. Five patients experienced disease stabilization, but no objective responses were documented. This combinatorial immunotherapy strategy is safe and feasible, and its immunobiological effects suggest potential activity in patients with minimal residual disease. A randomized trial exploring this hypothesis is currently ongoing.

Impact of time-of-flight and point-spread-function in SUV quantification for oncological PET.

Background Accuracy in the quantification of the SUV is a critical point in PET because proper quantification of tumor uptake is essential for therapy monitoring and prognosis evaluation. Recent advances such as time-of-flight (TOF) and point-spread-function (PSF) reconstructions have dramatically improved detectability. However, first experiences with these techniques have shown a consistent tendency to measure markedly high SUV values, bewildering nuclear medicine physicians and referring clinicians. Purpose We investigated different reconstruction and quantification procedures to determine the optimum protocol for an accurate SUV quantification in last generation PET scanners. Methods Both phantom and patient images were evaluated. A complete set of experiments was performed using a body phantom containing 6 spheres with different background levels and contrasts. Whole-body FDG PET/CT of 20 patients with breast and lung cancer was evaluated. One hundred five foci were identified by 2 experienced nuclear medicine physicians. Each acquisition was reconstructed both with classical and advanced (TOF, PSF) reconstruction techniques. Each sphere and each in vivo lesion was quantified with different parameters as follows: SUVmax, SUVmean, and SUV50 (mean within a 50% isocontour). Results This study has confirmed that quantification with SUVmax produces important overestimation of metabolism in new generation PET scanners. This is a relevant result because, currently, SUVmax is the standard parameter for quantification. SUV50 has been shown as the best alternative, especially when applied to images reconstructed with PSF + TOF. Conclusions SUV50 provides accurate quantification and should replace SUVmax in PET tomographs incorporating advanced reconstruction techniques. PSF + TOF reconstruction is the optimum for both detection and accurate quantification.

Diagnostic accuracy of FDG PET in the follow-up of platinum-sensitive epithelial ovarian carcinoma

PurposeThis study was designed to retrospectively evaluate the diagnostic yield of FDG PET for the diagnosis of recurrent ovarian cancer.MethodsEighty FDG PET scans were performed on 55 patients owing to suspicion of relapse, and 45 FDG PET scans were performed on 31 patients who were clinically disease free. PET results were compared with the results of conventional radiological imaging (CIM) and serum CA 125 levels, and related to pathological findings in 54 cases or clinical follow-up in 71 cases.ResultsCIM correctly identified 49 cases with recurrence [sensitivity (SE) 53.3%] ,and there were 27 true negatives [specificity (SP) 81.8%] However, 43 cases were false negative and six were false positive. The positive predictive value (PPV), negative predictive value (NPV) and accuracy (ACC) of CIM were 89%, 38.6% and 60.8%, respectively. FDG PET correctly detected recurrent disease in 80/92 cases (SE 86.9%, p < 0.05) and ruled out relapse in 26/33 cases (SP = 78.8%). The PPV, NPV and ACC of PET were 91.9%, 68.4% and 84.8%, respectively. Standardised uptake values did not provide additional diagnostic accuracy compared with visual analysis. The CA 125 results showed an SE of 57.6%, an SP of 93.9% and an ACC of 67.2%. In 23 patients with positive serum CA 125 levels, but negative CIM, FDG PET was positive and relapse was confirmed. Furthermore, FDG PET was positive and relapse was confirmed in 11 patients with negative serum CA 125 levels and CIM.ConclusionFDG PET may detect recurrent ovarian cancer earlier than CIM, with higher sensitivity and even higher diagnostic accuracy.

Positron Emission Tomography Using 18F-Fluorodeoxyglucose for the Evaluation of Residual Hodgkin's Disease Mediastinal Masses

Given its obvious prognostic implications, the correct interpretation of the significance of any residual mediastinal mass following Hodgkins disease (HD) treatment keeps maintaining its paramount importance. In this respect, 18F-fluorodeoxyglucose positron emission tomography (PET) is proving very effective for both active disease detection and relapse prediction. Twenty-nine consecutive HD patients, in whom computed tomography (CT) scan performed after therapy completion had documented a residual mediatinal mass of at least 2 cm, prospectively entered the study and underwent PET within 1 week from CT scan. With a median follow-up of 28 months from PET execution, no relapse was recorded among the 17 patients presenting with a negative PET. On the contrary, 9 of the 12 patients presenting with a positive PET relapsed/progressed within one year from PET execution. PETs negative and positive predictive values at 1 year were 100% and 75%, respectively. A negative PET seems to possibly exclude relapse in HD patient with a residual mediastinal mass. On the contrary, a positive PET result indicates a significantly higher risk of relapse. However, due to possible false positive results, a closer follow-up for all and a pathologic study in few selected patients is warranted.

A fully automated one pot synthesis of 9-(4-[18F]fluoro-3-hydroxymethylbutyl) guanine for gene therapy studies.

PURPOSE To develop a new fully-automated method for the synthesis of 9-(4-[18F]fluoro-3-hydroxymethylbutyl)guanine ([18F]FHBG) amenable for its routine use in gene therapy monitoring studies. PROCEDURES A nuclear interface commercial synthesizer was substantially modified and adapted to the synthesis of the referred compound. After the fluorination reaction of the tosylate precursor, the intermediate product was purified by Solid Phase Extraction (SPE) before the hydrolysis. The final product was purified by semi-preparative high performance liquid chromatography (HPLC). RESULTS [18F]FHBG was obtained in 10-15% yield in 65 minutes including HPLC purification. The radiotracer was > 99% chemically and radiochemically pure, sterile and free from pyrogens. The synthesized compound was shown to accumulate in thymidine kinase (tk) expressing cells both in cell culture, and in laboratory animals infected with an adenoviral vector containing the herpes simplex virus type 1 thymidine kinase (HSV1-tk) gene. CONCLUSIONS This new procedure facilitates the compliance with the applicable regulatory guidelines for positron emission tomography (PET) radiopharmaceuticals and will assist the clinical application of [18F]FHBG-PET as a noninvasive way to monitor gene therapy in humans.