Frequency-domain fluorescence lifetime imaging for endoscopic clinical cancer photodetection: Apparatus design and preliminary results

We describe a new fluorescence imaging device for clinical cancer photodetection in hollow organs in which the tumor/normal tissue contrast is derived from the fluorescence lifetime of endogenous or exogenous fluorochromes. This fluorescence lifetime contrast gives information about the physicochemical properties of the environment which are different between normal and certain diseased tissues. The excitation light from a CW laser is modulated in amplitude at a radio frequency by an electrooptical modulator and delivered by an optical fiber through an endoscope to the hollow organ. The image of the tissue collected by the endoscope is separated in two spectral windows, one being the backscattered excitation light and the other the fluorescence of the fluorochrome. Each image is then focused on the photocathode of image intensifiers (II) whose optical gain is modulated at the same frequency as the excitation intensity, resulting in homodyne phase-sensitive images. By acquiring stationary phase-sensitive frames at different phases between the excitation and the detection, it is possible to calculate in quasi-real time the apparent fluorescence lifetime of the corresponding tissue region for each pixel. A result obtained by investigating the endogenous fluorochromes present in the mucous membrane of an excised human bladder is presented to illustrate this method and most of the optical parameters which are of major importance for this photodetection modality have been evaluate

Endoscopic tissue characterization by frequency-domain fluorescence lifetime imaging (FD-FLIM)

Tissue characterization by endoscopic fluorescence imaging of endogenous or exogenous fluorochromes is a promising method for early cancer detection. However, the steady-state fluorescence contrast between healthy tissue and lesions such as early-stage carcinomas is generally poor. The authors propose to improve this contrast by using the additional information contained in the fluorescence lifetime (FLT). The FLT of several fluorochromes is sensitive to their physico-chemical environment. The FLT can be measured by frequency-domain methods. The excitation light from a continuous wave (CW) laser is modulated in amplitude at radio-frequencies by an electro-optic modulator, and delivered to the tissue via an optical fibre. The endoscopie site is imaged by an endoscope on to an optical device. The gain of the fluorescence image detector is also modulated at the same frequency for homodyning. The tissue fluorescence image is recorded at several phases between the excitation and the detection modulations during an acquisition cycle. With these images, an image processor calculates the apparent FLT for each pixel and constructs a lifetime image of the endoscopie site. This process is performed at quasi-video frequencies. The influence of various physical parameters (modulation frequency, number of images by cycle, shot noise, tissue optical properties etc.) has been investigated by analytical analysis, simulation methods and experimentation. Preliminary results obtained on human tissues are also presented to illustrate the potentiality of the metho

Frequency-domain fluorescence lifetime imaging for endoscopic clinical cancer photodetection: Apparatus design and preliminary results

We describe a new fluorescence imaging device for clin. cancer photodetection in hollow organs in which the tumor/normal tissue contrast is derived from the fluorescence lifetime of endogenous or exogenous fluorochromes. This fluorescence lifetime contrast gives information about the physicochem. properties of the environment which are different between normal and certain diseased tissues. The excitation light from a CW laser is modulated in amplitude at a radiofrequency by an electrooptical modulator and delivered by an optical fiber through an endoscope to the hollow organ. The image of the tissue collected by the endoscope is sepd. in two spectral windows, one being the backscattered excitation light and the other the fluorescence of the fluorochrome. Each image is then focused on the photocathode of image intensifiers (II) whose optical gain is modulated at the same frequency as the excitation intensity, resulting in homodyne phase-sensitive images. By acquiring stationary phase-sensitive frames at different phases between the excitation and the detection, it is possible to calc. in quasi-real time the apparent fluorescence lifetime of the corresponding tissue region for each pixel. A result obtained by investigating the endogenous fluorochromes present in the mucous membrane of an excised human bladder is presented to illustrate this method and most of the optical parameters which are of major importance for this photodetection modality have been evaluated

Miniaturized side-viewing imaging probe for fluorescence lifetime imaging (FLIM): validation with fluorescence dyes, tissue structural proteins and tissue specimens

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Clinical assessment of fluorescence cystoscopy during transurethral bladder resection in superficial bladder cancer

The prognosis of superficial bladder cancer in terms of recurrence and disease progression is related to bladder tumor multiplicity and the presence of concomitant "plane” tumors such as high-grade dysplasia and carcinoma in situ. This study in 33 patients aimed to demonstrate the role of fluorescence cystoscopy in transurethral resection of superficial bladder cancer. The method is based on the detection of protoporphyrin-IX-induced fluorescence in urothelial cancer cells by topical administration of 5-aminolevulinic acid. The sensitivity and the specificity of this procedure on apparently normal mucosa in superficial bladder cancer are estimated to be 82.9% and 81.3%, respectively. Thus, fluorescence cytoscopy is a simple and reliable method for mapping the bladder mucosa, especially in the case of multifocal bladder disease, and it facilitates the screening of occult dysplasi

Clinical assessment of fluorescence cytoscopy during transurethral bladder resection in superficial bladder cancer.

The prognosis of superficial bladder cancer in terms of recurrence and disease progression is related to bladder tumor multiplicity and the presence of concomitant "plane" tumors such as high-grade dysplasia and carcinoma in situ. This study in 33 patients aimed to demonstrate the role of fluorescence cystoscopy in transurethral resection of superficial bladder cancer. The method is based on the detection of protoporphyrin-IX-induced fluorescence in urothelial cancer cells by topical administration of 5-aminolevulinic acid. The sensitivity and the specificity of this procedure on apparently normal mucosa in superficial bladder cancer are estimated to be 82.9% and 81.3%, respectively. Thus, fluorescence cytoscopy is a simple and reliable method for mapping the bladder mucosa, especially in the case of multifocal bladder disease, and it facilitates the screening of occult dysplasia

Cylindrical fiberoptic light diffuser for medical applications

BACKGROUND AND OBJECTIVE: A cylindrical light diffuser has been developed mainly for medical applications, including photodynamic therapy (PDT), in particular interstitial PDT, PDT of the bronchi, or intravascular PDT. STUDY DESIGN/MATERIALS AND METHODS: The diffuser is based on a polymer optical fiber. The coupling of the light out of the core is controlled by the roughness mechanically induced on the surface of the core. The light is then isotropically diffused by a thin layer of a scattering medium. The active length can be 100 mm or more, whereas the outer diameter is 1 mm or less. The diffuser is flexible and can be introduced in tissue though a hypodermic needle. RESULTS: The main property of this light diffuser is the homogeneity of the light intensity emitted along its whole active length and around its circumference (360 degrees). Various intensity profiles can be made, including M-shape profiles for a homogeneous irradiance (+/- 10%) at a certain depth in the tissue. Furthermore, the diffuser is essentially isotropic and its optical properties are hardly dependent on wavelength. CONCLUSIONS: A diffuser for interstitial and intraluminal PDT has been developed. It is flexible, homogeneous, independent on wavelength, and can be made with a very high length-to-diameter ratio

Endoscopic frequency-domain fluorescence lifetime imaging for clinical cancer photodetection: apparatus design

We describe a new fluorescence imaging device for clinical cancer photodetection in hollow organs in which the image contrast is derived from the fluorescence lifetime of the fluorochrome at each point in a 2D image. Lifetime images are created from a series of images obtained from two gain-modulated image intensifiers. One of them (II-1) detects the light-induced tissue fluorescence, whereas the other (II-2) detects the backscattered fluorescence excitation light. This light is modulated at the same frequency as the detectors, resulting in homodyne phase-sensitive image. These stationary phase-sensitive images are collected using two CCD cameras, digitized and manipulated with a mathematical operator in real time. A series of such images, obtained with both image intensifiers at various phase shifts between their gain modulation and the modulation of the excitation light, is used to determine phase angle and/or the modulation of the fluorescence emission at each pixel. The reference values of these phase angles and modulations are obtained with II-2, whereas II-1 enables the measurement of the phase and modulation of the fluorescence. Phase and modulation are related to the fluorescence lifetime of the fluorochrome. An advantage of the experimental method proposed here is that pixel-to-pixel scanning is not required to obtain the fluorescence lifetime image, as the information from all pixels is obtained at the same time

Photodynamic therapy for early squamous cell carcinomas of the esophagus, bronchi, and mouth with m-tetra(hydroxyphenyl) chlorin

Objective: To clinically evaluate a new photosensitizer, m- tetra(hydroxyphenyl) chlorin (m-THPC), for the photodynamic therapy of early squamous cell carcinomas of the upper aerodigestive tract. Design: Phase 1 included evaluation of the innocuousness of the compound after intravenous injection (control of vital parameters and blood analysis before and after injection) and evaluation of the duration of skin photosensitization. Phase 2 included assessment of optimal conditions for treatment (injected dose, drug- light interval, light dose, wavelength, etc), on 33 early squamous cell carcinomas of the mouth, esophagus, and bronchi, with a mean follow-up of 14 months; irradiation tests on healthy and neoplastic mucosae to determine the irradiation conditions that lead to tumor eradication with minimal damage to the surrounding normal mucosa and muscle layers; and localization of the dye in various tissue compartments and cells at different time intervals after the injection of the photosensitizer, by using a fluorescence microscope to analyze 46 biopsy specimens taken during the treatment sessions and 8 resected specimens of early cancers, excised with the carbon dioxide laser. Setting: Endoscopic medical center of an otolaryngology-head and neck surgery department. Patients: Twenty-five patients treated previously for a head and neck cancer with a synchronous or metachronous early second primary cancer. Patients with porphyria were excluded from the trial. Results: The best results in the bronchi and mouth were obtained with an injected dose of 0.15 mg of m-THPC per kilogram of body weight 4 days before irradiation. The fluence was 7 to 16J/cm 2, and the fluence rate was between 100 and 150 mW/cm 2 using red light at 652 nm. In the esophagus, green light at 514 nm is preferred to the red light to avoid fistulas. Optimal irradiation conditions at this wavelength, which was also used in the trachea, were found at a fluence of 75 to 100J/cm 2 and a fluence rate between 70 and 100 mW/cm 2. Of 33 lesions treated thus far by photodynamic therapy with m-THPC, 28 show no recurrence with a mean follow-up of 14 months. Photosensitivity to sunlight does not exceed 6 weeks. Conclusions: m-Tetra(hydroxyphenyl) chlorin is a second-generation photosensitizer that has several significant advantages as compared with the first-generation porphyrin mixtures hematoporphyrin derivative and porfimer sodium (Photofrin II). It is a pure compound that is 100 times more phototoxic at 652 nm and 10 times more phototoxic at 514 nm, has better selectivity for early carcinomas, and a shorter duration of skin photosensitivity. The therapeutic results indicate a recurrence rate thai is similar to that obtained with Photofrin II, ie, about 15%

Photodynamic therapy for early squamous cell carcinomas of the esophagus, bronchi, and mouth with m-tetra (hydroxyphenyl) chlorin1

OBJECTIVE: To clinically evaluate a new photosensitizer, m-tetra(hydroxyphenyl) chlorin (m-THPC), for the photodynamic therapy of early squamous cell carcinomas of the upper aerodigestive tract. DESIGN: Phase 1 included evaluation of the innocuousness of the compound after intravenous injection (control of vital parameters and blood analysis before and after injection) and evaluation of the duration of skin photosensitization. Phase 2 included assessment of optimal conditions for treatment (injected dose, drug-light interval, light dose, wavelength, etc), on 33 early squamous cell carcinomas of the mouth, esophagus, and bronchi, with a mean follow-up of 14 months; irradiation tests on healthy and neoplastic mucosae to determine the irradiation conditions that lead to tumor eradication with minimal damage to the surrounding normal mucosa and muscle layers; and localization of the dye in various tissue compartments and cells at different time intervals after the injection of the photosensitizer, by using a fluorescence microscope to analyze 46 biopsy specimens taken during the treatment sessions and 8 resected specimens of early cancers, excised with the carbon dioxide laser. SETTING: Endoscopic medical center of an otolaryngology-head and neck surgery department. PATIENTS: Twenty-five patients treated previously for a head and neck cancer with a synchronous or metachronous early second primary cancer. Patients with porphyria were excluded from the trial. RESULTS: The best results in the bronchi and mouth were obtained with an injected dose of 0.15 mg of m-THPC per kilogram of body weight 4 days before irradiation. The fluence was 7 to 16 J/cm2, and the fluence rate was between 100 and 150 mW/cm2 using red light at 652 nm. In the esophagus, green light at 514 nm is preferred to the red light to avoid fistulas. Optimal irradiation conditions at this wavelength, which was also used in the trachea, were found at a fluence of 75 to 100 J/cm2 and a fluence rate between 70 and 100 mW/cm2. Of 33 lesions treated thus far by photodynamic therapy with m-THPC, 28 show no recurrence with a mean follow-up of 14 months. Photosensitivity to sunlight does not exceed 6 weeks. CONCLUSIONS: m-Tetra(hydroxyphenyl) chlorin is a second-generation photosensitizer that has several significant advantages as compared with the first-generation porphyrin mixtures hematoporphyrin derivative and porfimer sodium (Photofrin II). It is a pure compound that is 100 times more phototoxic at 652 nm and 10 times more photoxic at 514 nm, has better selectivity for early carcinomas, and a shorter duration of skin photosensitivity. The therapeutic results indicate a recurrence rate that is similar to that obtained with Photofrin II, ie, about 15