Development of Highly Potent and Selective Covalent FGFR4 Inhibitors Based on SNAr Electrophiles.

Fibroblast growth factor receptor 4 (FGFR4) is thought to be a driver in several cancer types, most notably in hepatocellular carcinoma. One way to achieve high potency and isoform selectivity for FGFR4 is covalently targeting a rare cysteine (C552) in the hinge region of its kinase domain that is not present in other FGFR family members (FGFR1-3). Typically, this cysteine is addressed via classical acrylamide electrophiles. We demonstrate that noncanonical covalent "warheads" based on nucleophilic aromatic substitution (SNAr) chemistry can be employed in a rational manner to generate highly potent and (isoform-)selective FGFR4 inhibitors with a low intrinsic reactivity. Key compounds showed low to subnanomolar potency, efficient covalent inactivation kinetics, and excellent selectivity against the other FGFRs, the kinases with an equivalent cysteine, and a representative subset of the kinome. Moreover, these compounds achieved nanomolar potencies in cellular assays and demonstrated good microsomal stability, highlighting the potential of SNAr-based approaches in covalent inhibitor design.

CT brain perfusion patterns and clinical outcome after successful cardiopulmonary resuscitation: A pilot study.

AIM: CT perfusion is a valuable tool for evaluating cerebrovascular diseases, but its role in patients with hypoxic ischaemic encephalopathy is unclear. This study aimed to investigate 1) the patterns of cerebral perfusion changes that may occur early on after successful resuscitation, and 2) their correlation with clinical outcome to explore their value for predicting outcome. METHODS: We conducted a retrospective analysis of perfusion maps from patients who underwent CT brain perfusion within 12 h following successful resuscitation. We classified the perfusion changes into distinct patterns. According to the cerebral performance category (CPC) score clinical outcome was categorised as favourable (CPC 1-2), or unfavourable (CPC 3-5). RESULTS: A total of 87 patients were included of whom 33 had a favourable outcome (60.6% male, mean age 60 ± 16 years), whereas 54 exhibited an unfavourable outcome (59.3% male, mean age 60 ± 19 years). Of the patients in the favourable outcome group, 30.3% showed no characteristic perfusion changes, in contrast to the unfavourable outcome group where all patients exhibit changes in perfusion. Eighteen perfusion patterns were identified. The most significant patterns for prediction of unfavourable outcome in terms of their high specificity and frequency were hypoperfusion of the brainstem as well as coexisting hypoperfusion of the brainstem and thalamus. CONCLUSION: This pilot study identified various perfusion patterns in patients after resuscitation, indicative of circulatory changes associated with post-cardiac-arrest brain injury. After validation, certain patterns could potentially be used in conjunction with other prognostic markers for stratifying patients and adjusting personalized treatment following cardiopulmonary resuscitation. Normal brain perfusion within 12 h after resuscitation is predictive of favourable outcome with high specificity.

MRI Extracellular Volume Fraction in Liver Fibrosis-A Comparison of Different Time Points and Blood Pool Measurements.

BACKGROUND: Extracellular volume (ECV) correlates with the degree of liver fibrosis. PURPOSE: To analyze the performance of liver MRI-based ECV evaluations with different blood pool measurements at different time points. STUDY TYPE: Prospective. SAMPLE: 73 consecutive patients (n = 31 females, mean age 56 years) with histopathology-proven liver fibrosis. FIELD STRENGTH/SEQUENCE: 3T acquisition within 90 days of biopsy, including shortened modified look-locker inversion recovery T1 mapping. ASSESSMENT: Polygonal regions of interest were manually drawn in the liver, aorta, vena cava, and in the main, left and right portal vein on four slices before and after Gd-DOTA administration at 5/10/15 minutes. ECV was calculated 1) on one single slice on portal bifurcation level, and 2) averaged over all four slices. STATISTICAL TESTS: Parameters were compared between patients with fibrosis grades F0-2 and F3-F4 with the Mann-Whitney U and fishers exact test. ROC analysis was used to assess the performance of the parameters to predict F3-4 fibrosis. A P-value <0.05 was considered statistically significant. RESULTS: ECV was significantly higher in F3-4 fibrosis (35.4% [33.1%-37.6%], 36.1% [34.2%-37.5%], and 37.0% [34.8%-39.2%] at 5/10/15 minutes) than in patients with F0-2 fibrosis (33.3% [30.8%-34.8%], 33.7% [31.6%-34.7%] and 34.9% [32.2%-36.0%]; AUC = 0.72-0.75). Blood pool T1 relaxation times in the aorta and vena cava were longer on the upper vs. lower slices at 5 minutes, but not at 10/15 minutes. AUC values were similar when measured on a single slice (AUC = 0.69-0.72) or based on blood pool measurements in the cava or portal vein (AUC = 0.63-0.67 and AUC = 0.65-0.70). DATA CONCLUSION: Liver ECV is significantly higher in F3-4 fibrosis compared to F0-2 fibrosis with blood pool measurements performed in the aorta, inferior vena cava, and portal vein at 5, 10, and 15 minutes. However, a smaller variability was observed for blood pool measurements between slices at 15 minutes. LEVEL OF EVIDENCE: 1 TECHNICAL EFFICACY: Stage 3.

Myocardial scar detection in free-breathing Dixon-based fat- and water-separated 3D inversion recovery late-gadolinium enhancement whole heart MRI.

The aim of this study was to investigate the diagnostic accuracy and reader confidence for late-gadolinium enhancement (LGE) detection of a novel free-breathing, image-based navigated 3D whole-heart LGE sequence with fat-water separation, compared to a free-breathing motion-corrected 2D LGE sequence in patients with ischemic and non-ischemic cardiomyopathy. Cardiac MRI patients including the respective sequences were retrospectively included. Two independent, blinded readers rated image quality, depiction of segmental LGE and documented acquisition time, SNR, CNR and amount of LGE. Results were compared using the Friedman or the Kruskal-Wallis test. For LGE rating, a jackknife free-response receiver operating characteristic analysis was performed with a figure of merit (FOM) calculation. Forty-two patients were included, thirty-two were examined with a 1.5 T-scanner and ten patients with a 3 T-scanner. The mean acquisition time of the 2D sequence was significantly shorter compared to the 3D sequence (07:12 min vs. 09:24 min; p < 0.001). The 3D scan time was significantly shorter when performed at 3 T compared to 1.5 T (07:47 min vs. 09:50 min; p < 0.001). There were no differences regarding SNR, CNR or amount of LGE. 3D imaging had a significantly higher FOM (0.89 vs. 0.78; p < 0.001). Overall image quality ratings were similar, but 3D sequence ratings were higher for fine anatomical structures. Free-breathing motion-corrected 3D LGE with high isotropic resolution results in enhanced LGE-detection with higher confidence and better delineation of fine structures. The acquisition time for 3D imaging was longer, but may be reduced by performing on a 3 T-scanner.

3D In Vivo Models for Translational Research on Pancreatic Cancer: The Chorioallantoic Membrane (CAM) Model.

Pancreatic ductal adenocarcinoma (PDAC) is a highly aggressive cancer with adverse outcomes that have barely improved over the last decade. About half of all patients present with metastasis at the time of diagnosis, and the 5-year overall survival rate across all stages is only 6%. Innovative in vivo research models are necessary to combat this cancer and to discover novel treatment strategies. The chorioallantoic membrane (CAM) model represents one 3D in vivo methodology that has been used in a large number of studies on different cancer types for over a century. This model is based on a membrane formed within fertilized chicken eggs that contain a dense network of blood vessels. Because of its high cost-efficiency, simplicity, and versatility, the CAM model appears to be a highly valuable research tool in the pursuit of gaining more in-depth insights into PDAC. A summary of the current literature on the usage of the CAM model for the investigation of PDAC was conducted and subdivided into angiogenesis, drug testing, modifications, personalized medicine, and further developments. On this comprehensive basis, further research should be conducted on PDAC in order to improve the abysmal prognosis of this malignant disease.

Methodologies and MR Parameters in Quantitative Magnetic Resonance Neurography: A Scoping Review Protocol.

Magnetic resonance neurography (MRN), the MR imaging of peripheral nerves, is clinically used for assessing and monitoring peripheral neuropathies based on qualitative, weighted MR imaging. Recently, quantitative MRN has been increasingly reported with various MR parameters as potential biomarkers. An evidence synthesis mapping the available methodologies and normative values of quantitative MRN of human peripheral nerves, independent of the anatomical location and type of neuropathy, is currently unavailable and would likely benefit this young field of research. Therefore, the proposed scoping review will include peer-reviewed literature describing methodologies and normative values of quantitative MRN of human peripheral nerves. The literature search will include the databases MEDLINE (PubMed), EMBASE (Ovid), Web of Science, and Scopus. At least two independent reviewers will screen the titles and abstracts against the inclusion criteria. Potential studies will then be screened in full against the inclusion criteria by two or more independent reviewers. From all eligible studies, data will be extracted by two or more independent reviewers and presented in a diagrammatic or tabular form, separated by MR parameter and accompanied by a narrative summary. The reporting will follow the guidelines of the Preferred Reporting Items for Systematic reviews and Meta-Analyses extension for Scoping Reviews (PRISMA-ScR). Upon completion, the scoping review will provide a map of the available literature, identify possible gaps, and inform on possible future research. SCOPING REVIEW REGISTRATION: Open Science Framework 9P3ZM.

Increasing the efficiency of hyperthermic intraperitoneal chemotherapy (HIPEC) by combination with a photosensitive drug in pediatric rhabdomyosarcoma in an animal model.

BACKGROUND: Cytoreductive surgery (CRS) in combination with hyperthermic intraperitoneal chemotherapy (HIPEC) is an option in advanced peritoneal sarcomatosis. Nevertheless, CRS and HIPEC are not successful in all patients. An enhancement of HIPEC using photodynamic therapy (PDT) might be beneficial. Therefore, a combination of the photosensitizer hypericin (HYP) with HIPEC was evaluated in an animal model. PROCEDURE: An established HIPEC animal model for rhabdomyosarcoma (NOD/LtSz-scid IL2Rγnullmice, n = 80) was used. All groups received HYP (100 µg/200 µl) intraperitoneally with and without cisplatin-based (30 or 60 mg/m2 ) HIPEC (37°C or 42°C, for 60 minutes) (five groups, each n = 16). Peritoneal cancer index (PCI) was documented visually and by HYP-based photodynamic diagnosis (PDD). HYP-based PDT of the tumor was performed. Tissue samples were evaluated regarding proliferation (Ki-67) and apoptosis (TUNEL). RESULTS: HYP uptake was detected even in smallest tumor nodes (<1 mm) with improved tumor detection during PDD (PCI with PDD vs. PCI without PDD: 8.5 vs. 7, p < .001***). Apoptotic effects after PDT without HIPEC were limited to the tumor surface, whereas PDT after HIPEC (60 mg/m2 , 42°C) showed additional reduction of tumor proliferation in the top nine to 11 cell layers (50 µm). CONCLUSION: HYP as fluorescent photosensitizer offers an intraoperative diagnostic advantage detecting intraperitoneal tumor dissemination. The combination of HYP and cisplatin-based HIPEC was feasible in vivo, showing enhanced effects on tumor proliferation and apoptosis induction across the tumor surface. Further studies combining HYP and HIPEC will follow to establish a clinical application.

Quantitative water T2 relaxometry in the early detection of neuromuscular diseases: a retrospective biopsy-controlled analysis.

OBJECTIVES: To assess quantitative water T2 relaxometry for the early detection of neuromuscular diseases (NMDs) in comparison to standard qualitative MR imaging in a clinical setting. METHODS: This retrospective study included 83 patients with suspected NMD who underwent multiparametric MRI at 3 T with a subsequent muscle biopsy between 2015 and 2019. Qualitative T1-weighted and T2-TIRM images were graded by two neuroradiologists to be either pathological or normal. Mean and median water T2 relaxation times (water T2) were obtained from manually drawn volumes of interests in biopsied muscle from multi-echo sequence. Histopathologic pattern of corresponding muscle biopsies was used as a reference. RESULTS: In 34 patients, the T1-weighted images showed clear pathological alternations indicating late-stage fatty infiltration in NMDs. In the remaining 49 patients without late-stage changes, T2-TIRM grading achieved a sensitivity of 56.4%, and mean and median water T2 a sensitivity of 87.2% and 97.4% to detect early-stage NMDs. Receiver operating characteristic (ROC) analysis revealed an area under the curve (AUC) of 0.682, 0.715, and 0.803 for T2-TIRM, mean water T2, and median water T2, respectively. Median water T2 ranged between 36 and 42 ms depending on histopathologic pattern. CONCLUSIONS: Quantitative water T2 relaxometry had a significantly higher sensitivity in detecting muscle abnormalities than subjective grading of T2-TIRM, prior to late-stage fatty infiltration signal alternations in T1-weighted images. Normal-appearing T2-TIRM does not rule out early-stage NMDs. Our findings suggest considering water T2 relaxometry complementary to T2-TIRM for early detection of NMDs in clinical diagnostic routine. KEY POINTS: • Quantitative water T2 relaxometry is more sensitive than subjective assessment of fat-suppressed T2-weighted images for the early detection of neuromuscular diseases, prior to late-stage fatty infiltration signal alternations in T1-weighted images. • Normal-appearing muscles in fat-suppressed T2-weighted images do not rule out early-stage neuromuscular diseases. • Quantitative water T2 relaxometry should be considered complementary to subjectively rated fat-suppressed T2-weighted images in clinical practice.

HIPEC-Induced Acute Kidney Injury: A Retrospective Clinical Study and Preclinical Model.

BACKGROUND: Hyperthermic intraperitoneal chemotherapy (HIPEC) combined with cytoreductive surgery (CRS) is the treatment of choice for selected patients with peritoneal malignancies. HIPEC is accompanied by moderate-to-high patient morbidity, including acute kidney injury. The significance of nephrotoxic agents such as cisplatin versus hyperthermia in HIPEC-induced nephrotoxicity has not been defined yet. PATIENTS AND METHODS: A total of 153 patients treated with HIPEC were divided into groups with (AKI+) and without (AKI-) kidney injury. Laboratory parameters and data concerning patient demographics, underlying disease, surgery, complications, and HIPEC were gathered to evaluate risk factors for HIPEC-induced AKI. A preclinical mouse model was applied to assess the significance of cisplatin and hyperthermia in HIPEC-induced AKI, as well as protective effects of the cytoprotective agent amifostine. RESULTS: AKI occurred in 31.8% of patients undergoing HIPEC. Treatment with cisplatin-containing HIPEC regimens represented a major risk factor for HIPEC-related AKI (p < 0.001). Besides, angiotensin receptor blockers and increased preoperative creatinine and urea levels were independent risk factors for AKI after HIPEC. In a preclinical mouse model, intraperitoneal perfusion with cisplatin induced AKI, whereas hyperthermia alone, or in combination with cisplatin, did not induce or enhance renal injury. Amifostine failed to confer nephroprotective effects in a miniaturized HIPEC model. CONCLUSIONS: AKI is a frequent complication after HIPEC. The risk of renal injury is particularly high in patients treated with cisplatin-containing HIPEC regimens. Hyperthermic perfusion of the abdomen by itself does not seem to induce or aggravate HIPEC-induced renal injury.

A new valid rhabdomyosarcoma spheroid culture model for in vitro evaluation of hypericin-based photodynamic therapy.

BACKGROUND: Advanced stages of pediatric alveolar rhabdomyosarcoma (RMA) are associated with an unfavorable outcome at established therapeutic strategies, accentuating the need for novel treatment options. Photodynamic therapy (PDT) with hypericin (HYP) has shown strong cytotoxic effects in two-dimensional (2D) cell culture. In order to more accurately mimic in vivo tissue architecture and better predict pharmaceutical response, the aim of this study was to establish a spheroid culture model by which PDT efficacy could be assessed in a three-dimensional (3D) context. MATERIALS AND METHODS: 3D multicellular tumor spheroids were generated using various scaffold-based and scaffold-free techniques. On two reproducible methods, HYP-PDT was performed varying spheroid sizes, photosensitizer concentrations, and illumination times. The ability for HYP uptake within the spheroid was analyzed assessing the substrate's autofluorescence. Antitumorigenic treatment effects were evaluated investigating cell viability, spheroid morphology, proliferative activity, and induction of apoptosis. RESULTS: Magnetic spheroid printing and orbital shaking methods were established as reproducible culturing systems producing uniform spheroids. Within assessed incubation times, HYP showed good penetration depth in spheroids containing 50,000 cells. PDT was causing metabolic and molecular impairment of RMA cells, resulting in viability decrease, reduction of cell proliferation, and induction of apoptosis. CONCLUSION: Assessing HYP-based PDT in a 3D culture model, we were able to gain an insight on how parameters like photosensitizer, oxygen, and light distribution contribute to the phototoxic effect. Compared to 2D cell culture, a higher treatment resistance was detected, which can be related to spheroid structure and mechanisms of intercellular communication, signal transduction, and gene expression.

Establishment of a new valid animal model for the evaluation of hyperthermic intraperitoneal chemotherapy (HIPEC) in pediatric rhabdomyosarcoma.

BACKGROUND: Cytoreductive surgery in combination with hyperthermic intraperitoneal chemotherapy has been established as a novel treatment approach for peritoneal sarcomatosis. Despite promising clinical reports, there is still a lack of knowledge regarding optimal drug usage and local effects. Therefore, we intended to establish a murine animal model for further evaluation. PROCEDURE: Alveolar rhabdomyosarcoma cells were xenotransplanted into NOD/LtSz-scid IL2Rγnullmice (n = 100). The mice received a continuous intraperitoneal lavage with isotonic saline solution as control or with cisplatin (30 or 60 mg/m2 ) as treatment group for 60 minutes at 37°C or 42°C (6 subgroups, each n = 16). Tumor spread was documented by an adapted peritoneal cancer index and MRI (n = 4). Tumor and tissue samples, harvested at the end of the perfusion, were evaluated regarding morphology, proliferation, and apoptosis (H&E-, Ki-67-, cleaved caspase 3-staining, TUNEL assay). RESULTS: Extensive peritoneal sarcomatosis in over 91% of the cases was observed. HIPEC was feasible without acute side effects. Ki-67 staining revealed concentration- or temperature-dependent effects of cisplatin-based HIPEC on the tumors. Although cleaved caspase-3 showed only sporadic apoptotic effects. TUNEL assay detected concentration- or temperature-dependent apoptotic effects at the outer tumor surface. MRI scans confirmed the observed tumor dissemination. CONCLUSION: This is the first animal model for evaluation of HIPEC in pediatric RMS in mice. Cisplatin-based HIPEC had early effects on the proliferation whereas circumscribed apoptotic effects could be detected at the tumor surface. This model allows further insights on the possible efficiency of HIPEC in RMS. Further studies using other drug combinations and treatment will follow.

Hypericin and its radio iodinated derivatives - A novel combined approach for the treatment of pediatric alveolar rhabdomyosarcoma cells in vitro.

BACKGROUND: Alveolar rhabdomyosarcoma (RMA) is a highly malignant soft tissue tumor in children with poor prognosis and failure of established therapies in advanced stages. Therefore, novel treatment options are required. Photodynamic therapy (PDT) has been found useful for the treatment of different tumor entities and might represent such a novel treatment option. A major limitation of PDT remains the restriction to superficial tumor cell layers as illumination with light is essential for the generation of reactive oxygen species. Current research focusses on the development of modified Hypericin (HYP)-based photosensitizers, as well as combining PDT and targeted internal radiotherapy with 131I, to generate an additive anti-tumor effect. METHODS: A standardized protocol for in vitro Hypericin-PDT was established in RMA cells. The anti-tumor properties of this photosensitizer were analyzed on molecular and metabolic levels. Changes in cell morphology were visualized using bright field-, fluorescence- and scanning-electron microscopy. Iodinated Hypericin derivatives with both radioactive and non-radioactive isotopes 131I/127I were employed to establish a targeted radionuclide therapy and investigate the potential of a combined treatment with PDT. RESULTS: In vitro photodynamic treatment with Hypericin showed a strong anti-tumor efficiency with favorable cellular uptake and compromised cancer cells on metabolic and molecular levels. Iodination of the photosensitizer did not impair the photosensitizer´s properties. Targeted radiotherapy with 131I-HYP led to distinct reductions of tumor viability. A simultaneously performed PDT leads to a reduction of cell viability that begins earlier in time. However, an additive enhancement of the cell viability was not observed in the selected dose range. CONCLUSION: In this in vitro study, we got a first insight of a possible potential of Hypericin for the treatment of pediatric soft tissue sarcoma. By coupling with radioiodine, we developed a novel approach for a combined anti-tumor treatment. The in vitro experiments lay the foundation for further in vivo experiments, which are needed to study the effects of a sequential administration of 131I-HYP and HYP.

Segmentation of Peripheral Nerves From Magnetic Resonance Neurography: A Fully-Automatic, Deep Learning-Based Approach.

Diagnosis of peripheral neuropathies relies on neurological examinations, electrodiagnostic studies, and since recently magnetic resonance neurography (MRN). The aim of this study was to develop and evaluate a fully-automatic segmentation method of peripheral nerves of the thigh. T2-weighted sequences without fat suppression acquired on a 3 T MR scanner were retrospectively analyzed in 10 healthy volunteers and 42 patients suffering from clinically and electrophysiologically diagnosed sciatic neuropathy. A fully-convolutional neural network was developed to segment the MRN images into peripheral nerve and background tissues. The performance of the method was compared to manual inter-rater segmentation variability. The proposed method yielded Dice coefficients of 0.859 ± 0.061 and 0.719 ± 0.128, Hausdorff distances of 13.9 ± 26.6 and 12.4 ± 12.1 mm, and volumetric similarities of 0.930 ± 0.054 and 0.897 ± 0.109, for the healthy volunteer and patient cohorts, respectively. The complete segmentation process requires less than one second, which is a significant decrease to manual segmentation with an average duration of 19 ± 8 min. Considering cross-sectional area or signal intensity of the segmented nerves, focal and extended lesions might be detected. Such analyses could be used as biomarker for lesion burden, or serve as volume of interest for further quantitative MRN techniques. We demonstrated that fully-automatic segmentation of healthy and neuropathic sciatic nerves can be performed from standard MRN images with good accuracy and in a clinically feasible time.

Radiostereometric migration analysis of the Cerafit femoral stem: 28 patients followed for 2 years.

Radiostereometric analysis (RSA) is the gold standard evaluating micromovements after total hip arthroplasty. The aim of this study was to investigate the migratory pattern of an uncemented femoral stem during the first 2 years after surgery. We followed 28 patients with a mean age of 57 (SD 13) years for the first two postoperative years. Radiostereometric analysis was used to measure the translation and rotation of the femoral component. The Harris hip score (HHS) was determined to evaluate the clinical outcome. No stem had to be revised. The mean HHS advanced from 35 (SD 11) preoperative to 89 (SD 10) 1 year after surgery. At the end of the observation period, mean subsidence of the stem was 0.26 mm (SD 0.82). Maximum total point motion (MTPM) was 1.23 mm (SD 1.22). The main distal migration took place up to 6 weeks after surgery with nearly no further subsidence up to 2 years postoperatively. All the measured migrations of the hip stem were very small. Results of the HHS demonstrate good clinical outcome. Long-term RSA is necessary to assess possible late migration of the Cerafit standard femoral stem.