Cross-Polarization OCT for In Vivo Diagnostics and Prediction of Bladder Cancer

This chapter contains three parts covering recent efforts to increase the accuracy of optical coherence tomography (OCT) differential diagnostics of bladder pathologies. The first part compares the diagnostic efficacy of traditional OCT and cross-polarization OCT (CP OCT); CP OCT and fluorescence cystoscopy (FC) for detecting flat lesions in the bladder at the early stages of cancer. The second part contains a report on achievements in application of CP OCT for detection of recurrent carcinoma in the scar area that is a hardly distinguishable form of bladder cancer using an optimized CP OCT image analysis. The third part of the chapter reviews the results on CP OCT usage for in vivo diagnosis of the bladder cancer after radiation therapy of cervical cancer

Label-free macroscopic fluorescence lifetime imaging of brain tumors

Advanced stage glioma is the most aggressive form of malignant brain tumors with a short survival time. Real-time pathology assisted, or image guided surgical procedures that eliminate tumors promise to improve the clinical outcome and prolong the lives of patients. Our work is focused on the development of a rapid and sensitive assay for intraoperative diagnostics of glioma and identification of optical markers essential for differentiation between tumors and healthy brain tissues. We utilized fluorescence lifetime imaging (FLIM) of endogenous fluorophores related to metabolism of the glioma from freshly excised brains tissues. Macroscopic time-resolved fluorescence images of three intracranial animal glioma models and surgical samples of patients\u27 glioblastoma together with the white matter have been collected. Several established and new algorithms were applied to identify the imaging markers of the tumors. We found that fluorescence lifetime parameters characteristic of the glioma provided background for differentiation between the tumors and intact brain tissues. All three rat tumor models demonstrated substantial differences between the malignant and normal tissue. Similarly, tumors from patients demonstrated statistically significant differences from the peritumoral white matter without infiltration. While the data and the analysis presented in this paper are preliminary and further investigation with a larger number of samples is required, the proposed approach based on the macroscopic FLIM has a high potential for diagnostics of glioma and evaluation of the surgical margins of gliomas

Genetically Encoded Fluorescent Sensor for Poly-ADP-Ribose

Poly-(ADP-ribosyl)-ation (PARylation) is a reversible post-translational modification of proteins and DNA that plays an important role in various cellular processes such as DNA damage response, replication, transcription, and cell death. Here we designed a fully genetically encoded fluorescent sensor for poly-(ADP-ribose) (PAR) based on Förster resonance energy transfer (FRET). The WWE domain, which recognizes iso-ADP-ribose internal PAR-specific structural unit, was used as a PAR-targeting module. The sensor consisted of cyan Turquoise2 and yellow Venus fluorescent proteins, each in fusion with the WWE domain of RNF146 E3 ubiquitin ligase protein. This bipartite sensor named sPARroW (sensor for PAR relying on WWE) enabled monitoring of PAR accumulation and depletion in live mammalian cells in response to different stimuli, namely hydrogen peroxide treatment, UV irradiation and hyperthermia

Pharmacokinetics of Chlorin e6-Cobalt Bis(Dicarbollide) Conjugate in Balb/c Mice with Engrafted Carcinoma

The necessary precondition for efficient boron neutron capture therapy (BNCT) is control over the content of isotope 10B in the tumor and normal tissues. In the case of boron-containing porphyrins, the fluorescent part of molecule can be used for quantitative assessment of the boron content. Study Objective: We performed a study of the biodistribution of the chlorin e6-Cobalt bis(dicarbollide) conjugate in carcinoma-bearing Balb/c mice using ex vivo fluorescence imaging, and developed a mathematical model describing boron accumulation and release based on the obtained experimental data. Materials and Methods: The study was performed on Balb/c tumor-bearing mice (CT-26 tumor model). A solution of the chlorin e6-Cobalt bis(dicarbollide) conjugate (CCDC) was injected into the blood at a dose of 10 mg/kg of the animal’s weight. Analysis of the fluorescence signal intensity was performed at several time points by spectrofluorimetry in blood and by laser scanning microscopy in muscle, liver, and tumor tissues. The boron content in the same samples was determined by mass spectroscopy with inductively coupled plasma. Results: Analysis of a linear approximation between the fluorescence intensity and boron content in the tissues demonstrated a satisfactory value of approximation reliability with a Spearman’s rank correlation coefficient of r = 0.938, p < 0.01. The dynamics of the boron concentration change in various organs, calculated on the basis of the fluorescence intensity, enabled the development of a model describing the accumulation of the studied compound and its distribution in tissues. The obtained results reveal a high level of correspondence between the model and experimental data

DataSheet_1_Highly Invasive Fluorescent/Bioluminescent Patient-Derived Orthotopic Model of Glioblastoma in Mice.docx

Development of the novel diagnostic and therapeutic approaches in neuro-oncology requires tumor models that closely reproduce the biological features of patients’ tumors. Patient-derived xenografts (PDXs) are recognized as a valuable and the most “close-to-patient” tool for preclinical studies. However, their establishment is complicated by the factors related to both the surgical material and technique of the orthotopic implantation. The aim of this work was to develop a patient-derived glioblastoma multiform (GBM) model that stably co-expresses luciferase and a far-red fluorescent protein for monitoring of tumor progression in the brain and, using this model, to validate new diagnostic methods—macroscopic fluorescence lifetime imaging (macro-FLIM) and cross-polarization optical coherence tomography (CP OCT). The established model was similar to the original patient’s GBM in terms of histological and immunohistochemical features and possessed reproducible growth in nude mice, which could be observed by both fluorescence and bioluminescence imaging. Our results demonstrated the high potential of macro-FLIM and CP OCT for intraoperative differentiation of GBM from the white matter. Thus, the dual-labeled PDX model of GBM proved to be an excellent approach for observation of tumor development by optical methods.</p