An ultrasound-guided fluorescence molecular tomography system is under development for in vivo quantification of
Protoporphyrin IX (PpIX) during Aminolevulinic Acid - Photodynamic Therapy (ALA-PDT) of Basal Cell Carcinoma.
The system is designed to combine fiber-based spectral sampling of PPIX fluorescence emission with co-registered
ultrasound images to quantify local fluorophore concentration. A single white light source is used to provide an estimate of the bulk optical properties of tissue. Optical data is obtained by sequential illumination of a 633nm laser source at 4 linear locations with parallel detection at 5 locations interspersed between the sources. Tissue regions from segmented ultrasound images, optical boundary data, white light-informed optical properties and diffusion theory are used to estimate the fluorophore concentration in these regions. Our system and methods allow interrogation of both superficial and deep tissue locations up to PpIX concentrations of 0.025ug/ml.
An ultrasound coupled handheld-probe-based optical fluorescence molecular tomography (FMT) system has been in
development for the purpose of quantifying the production of Protoporphyrin IX (PPIX) in aminolevulinic acid
treated (ALA), Basal Cell Carcinoma (BCC) <i>in vivo</i>. The design couples fiber-based spectral sampling of PPIX
fluorescence emission with a high frequency ultrasound imaging system, allowing regionally localized fluorescence
intensities to be quantified . The optical data are obtained by sequential excitation of the tissue with a 633nm
laser, at four source locations and five parallel detections at each of the five interspersed detection locations. This
method of acquisition permits fluorescence detection for both superficial and deep locations in ultrasound field. The
optical boundary data, tissue layers segmented from ultrasound image and diffusion theory are used to estimate the
fluorescence in tissue layers. To improve the recovery of the fluorescence signal of PPIX, eliminating tissue autofluorescence
is of great importance. Here the approach was to utilize measurements which straddled the steep Qband
excitation peak of PPIX, via the integration of an additional laser source, exciting at 637 nm; a wavelength
with a 2 fold lower PPIX excitation value than 633nm.The auto-fluorescence spectrum acquired from the 637 nm
laser is then used to spectrally decouple the fluorescence data and produce an accurate fluorescence emission signal,
because the two wavelengths have very similar auto-fluorescence but substantially different PPIX excitation levels.
The accuracy of this method, using a single source detector pair setup, is verified through animal tumor model
experiments, and the result is compared to different methods of fluorescence signal recovery.
The in vivo performance of a Fluorescence Molecular Tomography system as a function of pathophysiological
parameters that determine the penetration of nonbinding fluorescent nanoparticle was examined through imaging of
a series of three tumor models. The pathophysiological parameters examined were, vessel density, interstitial fluid
pressure (IFP), and collagen content. Drug delivery and IFP were measured in vivo via fluorescence spectroscopy
and a fiber-optic coupled pressure probe. Vessel density and collagen content were determined ex vivo through
histochemical analysis. The kinetics of the 40 nm,10000 KDa, fluorescent particles, which were injected into the tail
vein of the mice, was monitored by sequential excitation of the tissue on and off the tumor site through employment
of sixteen source detector pairs interspersed linearly in reflectance geometry. Each optical fluorescence data set was
collected at discrete time intervals in order to monitor drug uptake for a period of 45 minutes. The kinetics of the
drug delivery and the average nanoparticle uptake were correlated with the vessel density, interstitial pressure and
collagen content. The results of the correlations were verified to be consistent with the published relationship
between the three pathophysiological parameters and nanoparticle drug delivery.
Photodynamic therapy (PDT) for skin cancer is sometimes only partially effective, due to inadequate levels of the
fluorescent drug (photosensitizer, PS) and due to heterogeneous distribution of PS within the tissue. To image the PS
distribution within skin tumors, we have developed a fluorescence tomography system (FTS) that combines a
fluorescence detection array with a high frequency ultrasound (HFUS) transducer. In this paper we describe in vitro and
in vivo validation of this new system. The target fluorophore for detection was Protoporphyrin IX (PPIX). Validation
experiments were performed in vivo using a subcutaneous tumor model in which A431 tumor-bearing mice were treated
with 5-aminolevulinic acid to induce production of PPIX. FTS reconstructions were compared with standard histology
and with data from bulk tumor slices imaged ex vivo on a fluorescence scanner. Reconstructed images obtained from the
FTS were correlated with the histology and the ex vivo scans, confirming several-fold increases in PPIX fluorescence in
the skin and in the tumor relative to the surrounding tissues. Our data demonstrate the feasibility of using the FTS for
subsurface imaging of PPIX in skin carcinoma in vivo. Future aims are to use this device for individualized treatment
planning, in order to improve overall patient responses to PDT.