Prediction of Bone Marrow Biopsy Results From MRI in Multiple Myeloma Patients Using Deep Learning and Radiomics.

OBJECTIVES: In multiple myeloma and its precursor stages, plasma cell infiltration (PCI) and cytogenetic aberrations are important for staging, risk stratification, and response assessment. However, invasive bone marrow (BM) biopsies cannot be performed frequently and multifocally to assess the spatially heterogenous tumor tissue. Therefore, the goal of this study was to establish an automated framework to predict local BM biopsy results from magnetic resonance imaging (MRI). MATERIALS AND METHODS: This retrospective multicentric study used data from center 1 for algorithm training and internal testing, and data from center 2 to 8 for external testing. An nnU-Net was trained for automated segmentation of pelvic BM from T1-weighted whole-body MRI. Radiomics features were extracted from these segmentations, and random forest models were trained to predict PCI and the presence or absence of cytogenetic aberrations. Pearson correlation coefficient and the area under the receiver operating characteristic were used to evaluate the prediction performance for PCI and cytogenetic aberrations, respectively. RESULTS: A total of 672 MRIs from 512 patients (median age, 61 years; interquartile range, 53-67 years; 307 men) from 8 centers and 370 corresponding BM biopsies were included. The predicted PCI from the best model was significantly correlated ( P ≤ 0.01) to the actual PCI from biopsy in all internal and external test sets (internal test set: r = 0.71 [0.51, 0.83]; center 2, high-quality test set: r = 0.45 [0.12, 0.69]; center 2, other test set: r = 0.30 [0.07, 0.49]; multicenter test set: r = 0.57 [0.30, 0.76]). The areas under the receiver operating characteristic of the prediction models for the different cytogenetic aberrations ranged from 0.57 to 0.76 for the internal test set, but no model generalized well to all 3 external test sets. CONCLUSIONS: The automated image analysis framework established in this study allows for noninvasive prediction of a surrogate parameter for PCI, which is significantly correlated to the actual PCI from BM biopsy.

A rare cause for lower back pain: a case of an IgG4-related periaortitis.

IgG4-related disease (IgG4-RD) are a group of autoinflammatory diseases often presenting as tumor-like lesions because of their infiltrative or mass forming behavior. They are characterized by a typical histology consisting of storiform fibrosis, high numbers of infiltrating IgG4-positive plasma cells, obliterative phlebitis, and a moderate presence of eosinophilic cells. Serum IgG4 levels can be elevated. We present a case of a 57 year-old male patient with immobilizing lower back pain, fever, and night sweats. We diagnosed IgG4-related periaortitis using serum IgG4 levels, abdominal ultrasound, PET/CT, and histology. We successfully treated the patient with glucocorticoids (GC) and azathioprine. Periaortitis is a rare presentation of IgG4-RD and therefore noteworthy. It has to be considered in patients with a retroperitoneal mass.

Early effects of FOLFOX treatment of colorectal tumour in an animal model: assessment of changes in gene expression and FDG kinetics.

PURPOSE: The very early chemotherapeutic effects of the FOLFOX (fluorouracil, folinic acid, oxaliplatin) protocol were assessed in mice implanted with a human colorectal cell line. The aim of this study was to identify changes in gene expression patterns and to detect combinations of PET parameters that may be helpful in identifying treated tumours early after chemotherapy using dynamic PET studies. METHODS: A human colorectal cell line (HCT 116) was used in nude mice. Dynamic PET studies were performed in untreated (n = 13) and treated (n = 12) animals. The data were assessed using compartmental and noncompartmental analysis. The removed tumour specimens were assessed by gene array analysis to obtain quantitative information on gene expression. RESULTS: One chemotherapeutic treatment using the FOLFOX protocol resulted in an upregulation of 2,078 gene probes by more than 25%, while 2,254 probes were downregulated following treatment. The gene array data demonstrated primarily an enhancement of genes related to apoptosis. In particular, the apoptosis antigen 1 (APO-1), p21 and the G protein-coupled receptor 87 (G-87) were 2.6- to 3.3-fold upregulated as compared to the expression in untreated animals. There was a 100% separation of untreated and treated animals on the basis of these three genes. The SUV and the FDG kinetic parameters obtained by compartmental and noncompartmental fitting were not significantly different when individual parameters were compared between groups. However, classification analysis of the combination of the PET parameters VB, K1, k3, and influx revealed an overall accuracy of 84%. We were able to identify 91.7% (11/12) of the treated animals and 76.9% (10/13) of the untreated animals correctly using the classification analysis of PET data. CONCLUSION: Even one chemotherapeutic treatment using FOLFOX has an impact on gene expression and significantly modulates FDG kinetics. Quantitative assessment of the tracer kinetics and the application of classification analysis to the data are promising tools to identify those tumours that demonstrate a chemotherapeutic effect very early following treatment.

Changes in glucose metabolism and gene expression after transfer of anti-angiogenic genes in rat hepatoma.

PURPOSE: Human troponin I (TROP), the soluble receptor for vascular endothelial growth factor (sFLT) and angiostatin (ASTAT) are potent inhibitors of endothelial cell proliferation, angiogenesis and tumour growth in vivo. Transfer of these genes into tumours may induce changes not only in perfusion, but also more general ones such as changes in metabolism. The aim of this study was to assess these reactions using FDG-PET and high-throughput methods such as gene profiling. METHODS: We established Morris hepatoma (MH3924A) cell lines expressing TROP, sFLT or ASTAT and quantified (18)F-fluorodeoxyglucose ((18)FDG) uptake by dynamic positron emission tomography (PET) after tumour inoculation in ACI rats. Furthermore, expression of glucose transporter-1 and -3 (GLUT-1 and GLUT-3) as well as hexokinase-1 and -2 were investigated by RT-PCR and immunohistomorphometry. In addition, gene array analyses were performed. RESULTS: (18)FDG uptake, vascular fraction and distribution volume were significantly higher in all genetically modified tumours. Immunohistomorphometry showed an increased percentage of hexokinase-1 and -2 as well as GLUT-1 and -3 immunoreactive (ir) cells. Using gene arrays and comparing all three groups of genetically modified tumours, we found upregulated expression of 36 genes related to apoptosis, signal transduction, stress or metabolism. CONCLUSION: TROP-, sFLT- or ASTAT-expressing MH3924A tumours show enhanced influx of (18)FDG, which seems to be caused by several factors: enhanced exchange of nutrients between blood and tumour, increased amounts of glucose transporters and hexokinases, and increased expression of genes related to apoptosis, matrix and stress, which induce an increased demand for glucose.

Troponin I overexpression inhibits tumor growth, perfusion, and vascularization of morris hepatoma.

UNLABELLED: Antiangiogenic gene transfer inhibiting growth of new blood vessels is a promising approach in cancer therapy. Human troponin I (TnI) efficiently inhibits endothelial cell proliferation, migration, as well as angiogenesis and tumor growth in vivo. However, little is known about its effects on perfusion and tumor biology. METHODS: Stable Morris hepatoma (MH3924A) cells overexpressing human TnI (TnI-MH3924A) were cocultured with human umbilical vein endothelial cells (HUVECs) followed by measurements of endothelial apoptosis and proliferation. Furthermore, tumor growth and perfusion were determined using H(2)(15)O and (68)Ga-DOTA-albumin (DOTA is 1,4,7,10-tetraazacyclododecane-N,N',N'',N'''-tetraacetic acid) PET as well as functional MRI. Additionally, histologic measurements of vascularization, apoptosis, proliferation, and gene array analyses were performed. RESULTS: Apoptosis of HUVECs was increased and proliferation was decreased after coculture with TnI-MH3924A cells. TnI-MH3924A tumors showed a significant inhibition of growth (90%) and a decreased perfusion (25%), whereas blood volume remained unchanged. MRI investigations demonstrated a significant decrease of the rate constant k(ep). Immunohistochemical analyses showed decreased microvessel density and proliferation and significant induction of apoptosis. Furthermore, TnI-expressing hepatomas demonstrated changes in the expression of genes related to angiogenesis, apoptosis, signal transduction, or stress. CONCLUSION: TnI regulates tumor growth by modulating vascularization including apoptosis induction and decrease of proliferation. In addition, changes in expression of genes associated with angiogenesis, apoptosis, signal transduction, or stress were found. The upregulation of angiogenesis and stress-related genes indicates a cross-talk of different mechanisms as part of the tumor's reaction to TnI. Because the decrease of vascularization led to lower perfusion values as measured by PET and MRI, these noninvasive methods are promising tools for the monitoring of antiangiogenic gene therapy.

Angiopoietin-2 overexpression in morris hepatoma results in increased tumor perfusion and induction of critical angiogenesis-promoting genes.

UNLABELLED: Monitoring of angiogenesis-relevant approaches with functional imaging and histomorphometric analyses is desirable to evaluate the biologic effects. In this study we wished to examine the complex effects of angiopoietin-2 (Ang-2) gene transfer in a rat hepatoma model. METHODS: Using a bicistronic retroviral vector for Ang-2, Morris hepatoma (MH3924A) cell lines with Ang-2 expression were generated (Ang-2-MH3924A). In human umbilical vein endothelial cells (HUVECs) cocultured with Ang-2-MH3924A, the proliferative action with or without growth factors were determined. Furthermore, animal experiments were performed to measure effects on tumor growth and perfusion. Finally, tumors were examined by immunohistochemistry and DNA chip analysis. RESULTS: Ang-2-expressing MH3924A enhanced basic fibroblast growth factor-mediated endothelial cell proliferation. Perfusion, as measured by H(2)(15)O PET, was increased in genetically modified tumors. Consistent with the increased perfusion, micro- and macrovascularization were increased. However, tumor growth was similar to wild-type MH3924A (WT-MH3924A). Proliferating cell nuclear antigen and TUNEL (terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling) staining revealed an increased number of positive cells, indicating a compensation of increased proliferation by enhanced apoptosis. DNA chip analysis showed an induction of angiogenesis-promoting genes, including crucial vascular growth factor receptors, as well as genes related to extracellular matrix (ECM), apoptosis, signal transduction, and oxidative stress. CONCLUSION: Our results suggest that Ang-2 expression increases perfusion or vascularization, especially in interaction with the vascular growth factor system, without affecting tumor growth. Simultaneous, enhanced expression of genes for ECM, apoptosis, and signal transduction indicates Ang-2's versatile role in angiogenesis including its destabilizing function on ECM and endothelium.

Angiostatin overexpression in Morris hepatoma results in decreased tumor growth but increased perfusion and vascularization.

UNLABELLED: Growth of malignant tumors is dependent on sufficient blood supply. Thus, inhibition of tumor angiogenesis is emerging as a promising target in the treatment of malignancies. Human angiostatin (hANG) is one of the most potent inhibitors of endothelial cell proliferation, angiogenesis, and tumor growth in vivo. However, its mechanisms operating in vivo are not well understood. METHODS: To obtain more information about functional changes in the angiogenic process, we established Morris hepatoma (MH3924A) cell lines expressing hANG (hANG-MH3924A). The effects of hANG expression on proliferation and apoptosis of human umbilical vein endothelial cells (HUVECs) were measured in coculture experiments in vitro. To evaluate changes in tumor perfusion and blood volume, H2 15O and 68Ga-DOTA-albumin (DOTA is 1,4,7,10-tetraazacyclododecane-N,N',N'',N'''-tetraacetic acid) were used for PET studies in vivo. Additionally, immunohistologic quantification of vascularization, apoptosis, and proliferation as well as gene array analyses were performed. RESULTS: Our in vitro experiments demonstrate reduced proliferation and increased apoptosis in HUVECs when being cocultured with hANG-MH3924A. In support, tumor growth of hANG-MH3924A is diminished by 95% in vivo. However, tumor perfusion and blood volume are increased in hANG-MH3924A corresponding to an increased microvessel density. Furthermore, hANG-transfected tumors show changes in expression of genes related to apoptosis, stress, signal transduction, and metabolism. CONCLUSION: hANG expression leads to inhibition of tumor growth, increased apoptosis, and changes in the expression of multiple genes involved in stress reactions, signal transduction, and apoptosis, which indicates a multifactorial reaction of tumors. An enhanced microvessel density is seen as part of these reactions and is associated with increased perfusion as measured by PET.

Gallium-68-DOTA-albumin as a PET blood-pool marker: experimental evaluation in vivo.

Investigations into tumor angiogenesis and antiangiogenic treatment have renewed interest in tumor perfusion. To image tumor blood-pool by PET, suitable tracers are not generally available. In this experimental study, we characterized a 68Ga-labeled 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) conjugate of rat serum albumin (68Ga-DOTA-RSA) in vivo using a generator-produced isotope. Biodistribution was determined in ACI rats after intravenous administration of 3-6 MBq of 68Ga-DOTA-RSA. Three ACI rats were imaged over 1 h by dynamic PET after intravenous administration of 15-25 MBq of 68Ga-DOTA-RSA while the blood-pool activity was recorded simultaneously in a closed extracorporeal loop (ECL) between the carotid artery and the jugular vein. Time-activity curves (TACs) were obtained from volume of interest (VOI) analysis and from the ECL data. Stability and metabolites in plasma and urine were analyzed by size exclusion HPLC (SE-HPLC) 1 h after intravenous injection of 67Ga-DOTA-RSA. Blood radioactivity decreased by 10% and 18% from 10 to 60 min p.i. by biodistribution and PET or ECL, respectively. Tissue sampling between 10 and 60 min p.i. showed slight increases in the uptake of spleen, myocardium, kidney and skeletal muscle while hepatic accretion remained unchanged. Total urinary excretion after 60 min amounted to 9% of the injected dose. HPLC demonstrated a single urinary metabolite corresponding in size to gallium-labeled DOTA. 68Ga-DOTA-RSA is a blood-pool tracer whose physical and biological half-life is well suited for PET. Our findings support clinical imaging using 68Ga-DOTA-labeled human serum albumin (HSA). The generator-produced label makes 68Ga-DOTA-labeled albumin continuously available even to centers lacking an in-house cyclotron.

Improved correlation of histological data with DCE MRI parameter maps by 3D reconstruction, reslicing and parameterization of the histological images.

Due to poor correlation of slice thickness and orientation, verification of radiological methods with histology is difficult. Thus, a procedure for three-dimensional reconstruction, reslicing and parameterization of histological data was developed, enabling a proper correlation with radiological data. Two different subcutaneous tumors were examined by MR microangiography and DCE-MRI, the latter being post-processed using a pharmacokinetic two-compartment model. Subsequently, tumors were serially sectioned and vessels stained with immunofluorescence markers. A ray-tracing algorithm performed three-dimensional visualization of the histological data, allowing virtually reslicing to thicker sections analogous to MRI slice geometry. Thick slices were processed as parameter maps color coding the marker density in the depth of the slice. Histological 3D reconstructions displayed the diffuse angioarchitecture of malignant tumors. Resliced histological images enabled specification of high enhancing areas seen on MR microangiography as large single vessels or vessel assemblies. In orthogonally reconstructed histological slices, single vessels were delineated. ROI analysis showed significant correlation between histological parameter maps of vessel density and MR parameter maps (r=0.83, P=0.05). The 3D approach to histology improves correlation of histological and radiological data due to proper matching of slice geometry. This method can be used with any histological stain, thus enabling a multivariable correlation of non-invasive data and histology.

Transfer of the sFLT-1 gene in Morris hepatoma results in decreased growth and perfusion and induction of genes associated with stress response.

PURPOSE: Inhibition of tumor angiogenesis is emerging as a promising target in the treatment of malignancies. Therefore, monitoring of antiangiogenic approaches with functional imaging and histomorphometrical analyses are desirable to evaluate the biological effects caused by this treatment modality. EXPERIMENTAL DESIGN: Using a bicistronic retroviral vector for transfer of the soluble receptor for the vascular endothelial growth factor (sFLT) hepatoma (MH3924A) cell lines with sFLT expression were generated. In human umbilical vein endothelial cells cultured with conditioned medium of sFLT-expressing hepatoma cells, the inhibitory action of secreted sFLT was determined using a Coulter counter and a thymidine incorporation assay. Furthermore, in vivo experiments were done to measure the effects on tumor growth and perfusion. Finally, the tumors were examined by immunohistochemistry (including computer-assisted morphometry) and DNA chip analysis. RESULTS: Stable sFLT-expressing hepatoma cells inhibited endothelial cell proliferation in vitro. In vivo, growth and perfusion, as measured by H(2)(15)O positron emission tomography, were reduced in genetically modified tumors. However, the immunohistochemically quantified microvascularization and macrovascularization, as indicated by CD31- and alpha-actin-positive area, revealed no significant changes, whereas the number of apoptotic cells was increased in sFLT-expressing tumors, although not significantly. DNA chip analysis of tumors with gene transfer showed an increase of genes related to apoptosis, signal transduction, and oxidative stress. CONCLUSION: Our results suggest that sFLT expression inhibits tumor growth and perfusion and enhances expression of apoptosis-related genes in this model. Enhanced expression of genes for signal transduction, stress, and metabolism indicates tumor defense reactions.

Conjugation of DOTA using isolated phenolic active esters: the labeling and biodistribution of albumin as blood pool marker.

A convenient method for the functionalization of proteins with DOTA (1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid) has been developed. For this purpose DOTA was converted into a series of different monoreactive activated phenolic esters. The esters were prepared in a single step from commercially available DOTA, using 1-ethyl-3-[3-(dimethylamino)propyl]carbodiimide or 1,3-dicyclohexylcarbodiimide as coupling agent. The resulting activated esters were isolated by HPLC, lyophilized, and stored for future applications. In solid form the compounds exhibit high hydrolytic stability. The reactions with proteins proceeded in good yields. The conjugation and subsequent radiolabeling of the 4-nitrophenol ester of DOTA with 67Ga was investigated with rat serum albumin. A time-dependent biodistribution study in tumor bearing rats was conducted to demonstrate the integrity of the albumin conjugate. These results suggest that phenolic esters of DOTA represent versatile reagents to conjugate DOTA with proteins and other biomolecules in high yields.

Pharmacological properties of hydrophilic and lipophilic derivatives of octreotate.

Derivatives of somatostatin (SST) represent the most important peptides for receptor targeting in oncological applications. Whereas the pharmacophor in somatostatin receptor-affine substances has been thoroughly investigated, the influence of modifications at the N-terminal has not yet been systematically studied. In order to investigate the influence of hydrophilic versus lipophilic modifications at the N-terminal end, a series of homologous derivatives of Tyr3-octreotate modified with oligomers of ethylene glycol or fatty acids were synthesized. For this purpose, Tyr3-octreotate was assembled using solid phase peptide synthesis and the fatty acids or oligomers of ethylene glycol were conjugated to the N-terminal end. The oligomers of ethylene glycol were activated by 4-nitrophenylchloroformate to obtain carbamate-linked hydrophilic compounds. The receptor affinities of these compounds were determined by competition experiments with [125I]Tyr3-octreotide on rat cortex membranes. The hydrophilic derivatives and the short chain lipophilic derivatives revealed IC50 values between 0.66 +/- 0.02 nM and 2.16 +/- 0.31 nM respectively. After labeling with (125)I the organ distribution of selected derivatives was investigated in Lewis rats bearing the rat pancreatic tumor CA20948. All of the compounds showed high tumor uptake. The peptides conjugated to oligomers of ethylene glycol showed low uptake into the liver and kidneys. Increasing the length of the fatty acids resulted in a remarkable decrease in kidney uptake. In conclusion, the systematic modifications at the N-terminal result in a low effect on the receptor affinity but allow the modulation of the pharmacokinetic properties of octreotide derivatives.

A multivessel model describing replenishment kinetics of ultrasound contrast agent for quantification of tissue perfusion.

To improve the quantification of tissue perfusion using intermittent sonography, a new model describing replenishment kinetics of microbubbles is proposed. The new approach takes into account the variability of blood flow velocities found in vivo, especially in tumors, and consistently describes the refilling process of microbubbles. Based upon this model, blood volume, blood velocity, blood flow and perfusion in 17 experimental tumors were calculated, and compared with the results obtained with the established, phenomenologically derived exponential kinetic model. In contrast to the existing model, our approach describes tissue vascularization more physiologically and allows deduction of a consistent new hyperbolic model for quantification of intermittent sonography. Blood volume and mean blood velocity did significantly correlate between both the new and the established model (k=0.99; k=0.94, both p<0.001). However, mean tumor blood velocity was lower (-19%, p<0.01) with the established model compared to the newly developed model. In addition, the range and distribution of blood flow velocities found in vivo can be estimated with the new model. Furthermore, it uses simpler mathematical fitting routines and allows easier data acquisition, which may allow a more practicable clinical application of intermittent sonography. In conclusion, a more valid, detailed and accurate calculation of perfusion parameters, especially of tumors, can be derived in vivo with the new multivessel model of intermittent sonography.

Preoperative ultrasonographic identification of the sentinel lymph node in patients with malignant melanoma.

BACKGROUND: Dissection of the "sentinel lymph node" (SLN) as identified by lymphoscintigraphy is becoming increasingly important in the treatment of patients with malignant melanoma. The purpose of the current study was to determine whether the SLN also could be identified by ultrasound. METHODS: Sixty-seven patients with malignant melanoma (40 females and 27 males, with an average age of 48.8 years) in whom extirpation of the SLN was indicated underwent ultrasonography of the regional lymph nodes prior to preoperative lymphoscintigraphy. The location of the melanoma was the legs in 30 patients, the arms in 14 patients, and the trunk in 23 patients. During regional ultrasonography, the location of the lymph nodes that differed in the cortex/medulla ratio from the surrounding lymph nodes was marked on the skin corresponding to the planes of insonation (M1) when the probe was held vertically to the skin surface. After lymphoscintigraphy using technetium-99m, the position of a gamma probe at which the highest count rate vertical to the skin surface was recorded also was marked (M2). RESULTS: In the inguinal region, the agreement between M1 and M2 was found to be 100% (40 of 40 SLNs) and was 72.5% in the axilla (29 of 40 SLNs). In patients with melanomas located on the leg, the location of M1 and M2 agreed in 97% of cases (36 of 37 lymph nodes in 30 patients); in patients with melanomas located on the arms, the agreement was 76% (13 of 17 lymph nodes in 14 patients) and in patients with melanomas located on the trunk, the agreement was 75% (21 of 28 lymph nodes in 23 patients). The position documented by ultrasound relative to the neighboring structures of the SLN was confirmed intraoperatively in all cases. CONCLUSIONS: The results of the current study indicate that the SLN in patients with melanoma located on the limbs, especially the legs, are characterized by a specific sonomorphologic pattern. Preoperative sonography might constitute an important addition to lymphoscintigraphy in the planning of SLN biopsy.