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Purpose: Positron emission tomography with the radioligand [¹¹C]D-deprenyl has shown an increased signal at the location of pain in patients with ankle sprains, rheumatoid arthritis and chronic whiplash injury, but the mechanism of this tracer uptake and its exact binding site in inflammation or tissue injury is still unclear. The aim of this study was to further evaluate [¹¹C]D-deprenyl´s usefulness as a marker of acute inflammation.

Methods: An animal PET/CT study was performed three days after the induction of a rat model of inflammatory or surgical pain. Fourteen adult male Sprague-Dawley rats and three tracers [¹¹C]D-deprenyl, [¹¹C]L-dideuterumdeprenyl and [¹⁸F]fluorodeoxyglucose were used.

Results: No [¹¹C]D-deprenyl accumulation was seen in a rat model of musculoskeletal pain. In the rat model of inflammatory pain all three ligands were shown to visualize the inflamed ankle joint with much lower uptake in the control ankle joint. The uptake was largest with [¹¹C]D-deprenyl and [¹¹C]L- dideuteriumdeprenyl, where approximately 1 % of the injected dose could be found in the affected ankle joint during the first minutes, whereas the uptake of [¹⁸F]FDG was approximately 0.5 % of the injected dose. However, the ratio of uptake of the injected ankle joint versus the control ankle joint was much higher for [¹⁸F]FDG (around 10 fold increase) than for the two deprenyl enantiomers (2 – 3 fold increase). The uptake pattern of [¹¹C]D-deprenyl and [¹¹C]L-dideuteriumdeprenyl did not show signs of specific binding or irreversible trapping.

Conclusions: Contrary to our expectations, of the three tracers only [¹⁸F]FDG may be used as markers of peripheral inflammation in a rat model of inflammatory pain. However, as a high site-specificity is required, [¹¹C]D-deprenyl and [¹¹C]L-dideyteriumdeprenyl deserve further exploration regarding sensitivity, specificity and uptake mechanisms in human pain syndromes.

The understanding of etiological mechanisms of whiplash associated disorder is still inadequate. Objective visualization and quantification of peripheral musculoskeletal injury and possible painful inflammation in whiplash associated disorder would facilitate diagnosis, strengthen patients’ subjective pain reports and aid clinical decisions eventually leading to better treatments. In the current study, we further evaluated the potential to use [¹¹C]D-deprenyl PET/CT to visualize inflammation after whiplash injury. Sixteen patients with whiplash injury grade II were recruited at the emergency department and underwent [¹¹C]D-deprenyl PET/CT in the acute phase and at 6 months after injury. Subjective pain levels, self rated neck disability and active cervical range of motion were recorded at each imaging session. Results showed that the molecular aspects of inflammation and possible tissue injuries after acute whiplash injury could be visualized, objectively quantified and followed over time with [¹¹C]-D-deprenyl PET/CT. An altered [¹¹C]D-deprenyl uptake in the cervical bone structures and facet joints was associated with subjective pain levels and self rated disability during both imaging occasions. These findings may contribute to a better understanding of affected peripheral structures in whiplash injury and strengthens the idea that PET/CT detectable organic lesions in peripheral tissue may be relevant for the development of persistent pain and disability in whiplash injury.

Perspective: This article presents a novel way of objectively visualizing possible structural damage and inflammation that cause pain and disability in whiplash injury. This PET method can bring an advance in pain research and eventually would facilitate the clinical management of patients in pain.

Background:

Diffusion-weighted imaging (DWI) has become increasingly valuable in lymph node imaging, yet the clinical utility of this technique in the staging of lymphoma has not been established.

Purpose:

To compare whole-body DWI with FDG-PET/CT in the staging of lymphoma patients.

Material and Methods:

Thirty-one patients, eight with Hodgkin lymphoma (HL) and 23 with non-Hodgkin's lymphoma (18 aggressive and five indolent) underwent both whole-body DWI, whole-body MRI (T1W and T2W-STIR) and FDG-PET/CT. Lesions on whole-body DWI were only considered positive if they correlated with lesions on T1W and T2W-STIR images. The staging given by each technique was compared, according to the Ann Arbor staging system. Differences in staging were solved using biopsy results, and clinical and CT follow-ups as standard of reference.

Results:

The staging was the same for DWI and FDG-PET/CT in 28 (90.3%) patients and different in three (9.7%). Of the 28 patients with the same staging, 11 had stage IV in both techniques and 17 had stages 0-III. No HL or aggressive non-Hodgkin's lymphoma patients had different staging. Three indolent small lymphocytic lymphoma/chronic lymphocytic leukemia (SLL/CLL) lymphoma had higher staging with DWI when compared with FDG-PET/CT. One small subcutaneous breast lymphoma was not seen but all other extranodal sites were detected by both techniques.

Conclusion:

Whole-body DWI is a promising technique for staging of both (aggressive and indolent) non-Hodgkin's lymphoma and HL.

Label free time-lapse video microscopy based monitoring of time evolving cell population morphology has potential to offer a simple and cost effective method for identity control of cell lines. Such morphology monitoring also has potential to offer discovery of chemically induced differential changes between pairs of cell lines of interest, for example where one in a pair of cell lines is normal/sensitive and the other malignant/resistant. A new simple algorithm, pixel histogram hierarchy comparison (PHHC), for comparison of time evolving morphologies (TEM) in phase contrast time-lapse microscopy movies was applied to a set of 10 different cell lines and three different iso-genic colon cancer cell line pairs, each pair being genetically identical except for a single mutation. PHHC quantifies differences in morphology by comparing pixel histogram intensities at six different resolutions. Unsupervised clustering and machine learning based classification methods were found to accurately identify cell lines, including their respective iso-genic variants, through time-evolving morphology. Using this experimental setting, drugs with differential activity in iso-genic cell line pairs were likewise identified. Thus, this is a cost effective and expedient alternative to conventional molecular profiling techniques and might be useful as part of the quality control in research incorporating cell line models, e.g. in any cell/tumor biology or toxicology project involving drug/agent differential activity in pairs of cell line models.

The thickness of a primary malignant melanoma tumor is the most important prognostic indicator for a patient with primary cutaneous malignant melanoma. To optimize the management and treatment of melanoma patients there is an unmet need to identify characteristics that can further stratify melanoma patients into high or low risk for progressive disease. Despite numerous studies no single marker has yet been shown to add significant prognostic information. An algorithmic approach, combining data from several markers provides an attractive model to identify patients of increased risk of dying from malignant melanoma. The primary aim of the present study was to analyze the correlation between clinical outcome and protein expression patterns of multiple proteins in malignant melanoma tumors using immunohistochemistry and tissue microarrays. Candidate proteins were identified based on a selective and differential expression pattern in melanoma tumors and tested in a cohort of 143 melanoma patients. Protein expression was analyzed using both manual scoring and automated image analysis-based algorithms. We found no single marker of prognosis that was independent of tumor thickness. When combining potential prognostic markers we could define a prognostic index, based on RBM3, MITF, SOX10 and Ki-67, that was independent of tumor thickness in multivariate analysis. Our findings suggest that a good prognosis signature can be identified in melanoma patients with tumors showing a low fraction of Ki-67 positive tumor cells and a high fraction of RBM3 positive tumor cells combined with low intensity levels of SOX10 and MITF.

Introduction: Molecular imaging of HER2-expression in malignant tumors provides potentially important information for patient management. Affibody molecules have shown to be suitable tracers for imaging applications using SPECT or PET. Results from an earlier evaluation of the application of site specific ^99mTc-labeling on the Affibody molecule, Z_HER2:2395-C, were favorable.

Methods: As a preparation for clinical application of this tracer we have developed and evaluated a robust single-vial freeze-dried kit, allowing labeling of the Affibody molecule, Z_HER2:2395-C, with ^99mTc.

Results: The composition of the kit (containing glucoheptonate, EDTA and tin(II)-chloride), as well as the protein amount and the pertechnetate volume were optimized for a high labeling yield (> 90 %) and minimal presence of reduced hydrolyzed technetium colloids (< 1 %). The specificity to HER2 receptors, the binding competence and the stability in PBS and murine serum were verified in vitro. The shelf-life was also evaluated in vitro, showing no reduction in labeling yield or binding capacity to HER2-expressing cells after over 400 days of storage of the single-vial freeze-dried kit.

Conclusions: Z_HER2:2395-C labeled with ^99mTc using the lyophilized kit was stable and resulted in a favorable biodistribution in an in vivo evaluation in normal NMRI mice.

Overexpression of HER2 in breast carcinomas predicts response to trastuzumab therapy. Affibody molecules based on a non-immunoglobulin scaffold have demon-strated high potential for in vivo molecular imaging of HER2-expressing tumors. Re-engineering of the molecular scaffold has led to a second generation of optimized Affibody molecules, having a surface distinctly different from the parental protein domain from staphylococcal protein A. The new tracer showed further increased melting point, stability and overall hydrophilicity compared to the parental molecule, and was shown to be more amenable for chemical peptide synthesis. The goal of this study was to assess potential effects of this extensive re-engineering on HER2 targeting, using ABY-025, a DOTA conjugated variant of the novel tracer.

Methods: ¹¹¹In-ABY-025 was compared with previously evaluated parent HER2-binding Affibody tracers in vitro and in vivo. The in vivo behavior was further evaluated in mice bearing SKOV-3 xenografts, in rats and in cynomolgus macaques.

Results: ¹¹¹In-ABY-025 bound specifically to HER2 in vitro and in vivo. Direct comparison with the previous generation of HER2-binding tracers showed that ABY-025 retained excellent targeting properties. Rapid blood clearance was shown in mice, rats and macaques. A highly specific tumor uptake of 16.7 ± 2.5 %IA/g was seen at 4 h after injection. The tumor-to-blood ratio was 6.3 at 0.5 h, 88 at 4 h, and increased up to 3 days after injection. Gamma camera imaging of tumors was already possible 0.5 h after injection. Furthermore, repeated i.v. administration of ABY-025 did not induce antibody formation in rats.

Conclusions: The biodistribution of ¹¹¹In-ABY-025 was in remarkably good agreement with the parent tracers, despite profound re-engineering of the non-binding surface. The molecule displayed rapid blood clearance in all species investigated and excellent targeting capacity in tumor bearing mice, leading to high tumor-to-organ-ratios and high contrast imaging shortly after injection.

The current way to increase efficacy of cancer therapy is the use of molecular recognition of aberrantly expressed gene products for selective treatment. However, only a fraction of the patients have tumors with a particular molecular target. Radionuclide imaging of molecular targets might help to stratify patient for cancer treatment. Affibody molecules are scaffold proteins, which can be selected for high affinity recognition of proteinaceous molecular targets. The capacity to re-fold under physiological conditions allows labeling of Affibody molecules in a broad range of pH and temperatures with preserved binding properties. Peptide synthesis or introduction of a unique cysteine enables site-specific labeling of Affibody molecules, resulting in uniform conjugates with well-defined pharmacological characteristics. The small size (7 kDa) of Affibody molecules provides rapid extravasation, rapid tumor penetration, and rapid clearance of unbound tracer from healthy organs and tissues. In combination with sub-nanomolar affinity, this results in high contrast in vivo imaging a few hours after injection. Excellent targeting has been demonstrated in pre-clinical studies with HER2-targeting Affibody molecules labeled with (99m)Tc and (111)In for single photon computed tomography (SPECT), and (18)F, (64)Cu, (124)I and (68)Ga for positron emission tomography (PET). Pilot clinical data confirm the high potential of Affibody molecules.