A laboratory version of an LED-array light source centered at 664 nm was reviewed last year in Biomedical Optics '93. Since that time, primary development efforts have been directed toward the development of two LED-array light sources for preclinical trials, one centered at 630 nm and the other at 800 nm. The first array is targeted as an alternative to the argon- pumped dye laser for photodynamic therapy (PDT) procedures based on the Photofrin II photosensitizer. The second array at 800 nm is directed at deep tissue penetration with a new photosensitizer recently developed at the University of Utah, bacteriochlorin derivative (bcd). While the first array is viewed as a lower-cost, more reliable, user-friendly replacement for traditional PDT dye lasers, the second is being developed in unison with bcd to significantly expand the range of applications of PDT to previously undeveloped cancer treatment modalities such as breast cancer. The 800 nm array has demonstrated an optical output of 5.13 watts while the 630 nm source radiates with a maximum optical power output of 1.63 watts.
Two types of N-(2-hydroxypropyl)methacrylamide (HPMA) copolymer containing meso- chlorin e<SUB>6</SUB> monoethylene diamine disodium salt (Mce<SUB>6</SUB>) were synthesized. The Mce<SUB>6</SUB> was bound via pendant enzymatically degradable oligopeptide side chains (G-F-L-G) in one copolymer and was attached through noncleavable side chains (G) in the other. Preliminary experiments have been undertaken to compare their localization/retention behavior and their tumorcidal activity in vivo (A/J mice; C1300 neuroblastoma). Results of localization/retention experiments indicated that the Mce<SUB>6</SUB> bound to the noncleavable copolymer was retained in the tumor and other tissues for a prolonged time period compared with free Mce<SUB>6</SUB> or the Mce<SUB>6</SUB> bound to the cleavable copolymer. Light activation of the Mce<SUB>6</SUB> from the cleavable copolymer rendered a substantially more potent biological response in vivo than did the permanently bound Mce<SUB>6</SUB>. It is hypothesized and indirectly supported by photophysical data that both of the polymer-photosensitizer complexes are aggregated (or conformationally altered) under physiological conditions due to their hydrophilic/hydrophobic properties. In buffer at pH 7.4, the quantum yield of singlet oxygen generation by free Mce<SUB>6</SUB> is three-fold higher than by the Mce<SUB>6</SUB> bound to a noncleavable copolymer; adding detergent (CTAB), increased the quantum yield of singlet oxygen generation to a value consistent with that of the free Mce<SUB>6</SUB>. In vivo, if a sufficient time lag is allowed after drug administration for tumor cell lysosomal enzymes to cleave the Mce<SUB>6</SUB> from the polymer containing degradable side chains, the Mce<SUB>6</SUB> would be released in free form and behave with properties akin to the free drug. Due to the difference in cellular uptake mechanisms for free and bound drugs (and the targeting potential of the copolymer), a much higher local concentration in the tumor compared with surrounding tissue can be achieved with the polymer bound drug than the free photosensitizer. Side effects characteristics of current PDT treatment such as light ultrasensitivity in skin may be reduced by improving localization selectivity.
Locally absorbing microvolumes (10Å-10?m) much smaller than the radiation wavelength in
size are characteristic of heterogeneous microstructure in cells and living systems and can be
studied and controlled with ultrafast pulses of light. The ultrafast transient absorption and
heating of local microvolumes absorbing through endogenous of exogenous chromophores at the
radiation wavelength can be used to study size, structure and function of locally overheated
microstructures. Pulse-heated microvolumes with altered refractive index and scattering and
altered fluorescence are probed with a second light pulse. Also, the pulsed heating of the
desired kind of microvolumes in cells and tissues with ultrafast laser pulses of a certain
wavelength pulse duration and intensity opens up new possibilities for photothermotherapy.
Ultrafast transient overheating of microvolumes may be substantial (?T=1-100 deg) while the
time-and space-averaged heating of irradiated macrovolume is much lower. The fast transient
perturbation of living systems with ultrafast, tunable laser pulses that significantly effect
biological processes form the basis for new therapeutic applications. Ultrashort laser pulses
(fs-ns) are shorter in duration than the time it takes for heat to diffuse from microregions even
as small as 10-IOOA° across and coupled with their wide-band tunability make it possible to
investigate local absorption microregions using endogenous or exogenous chromophores to
determine optimum wavelength for spectroscopy and phototherapy. We have demonstrated
remarkable effects on cell growth with femtosecond laser pulses (620nm) at an average
intensity of 5.5x10-4 W/cm2 and dose of 0.33 J/cm2.
An electronic optical imaging system consisting of a computer-controlled CCD array with 576 x 384
detection elements and 14 bits of digitization is presented. The system is used to obtain internal and
external light intensity distributions for diffusing optical fiber tips used with photodynamic therapy
and laser angioplasty. Significant intensity distribution variations were observed between fiber tips
from 5 different sources. The effect of launch numerical aperture on the measured light distributions
is also presented.
An open-loop temperature control was introduced to control evolution of the maximum
temperature on the tissue surface to be within upper and lower limits. For this purpose, the temperature
evolutions of sample shots were analyzed and optimal sequences of laser pulses were computed. The
1.06 tm pulsed Nd:YAG laser was used and the thermal camera measured temperature. Experiment
on animals in vivo and in vitro was performed to test the technique. Upper and lower temperature
limits during laser irradiation were set below 100 °C since thermal coagulation was ofprimary concern.
Usually, difference between the upper and lower limits was set to 1 5°C during experiment. However,
this difference depended on the laser specifications such as power, pulse width, and repetition rates, as
well as on tissue properties. Coagulation studies showed a clear relation of temperature versus
cogulation depth. Therefore, the heating temperature and the duration time can be used as primary
parameters instead of laser power and exposure time or energy.
Fiber tip breakage during urinary and biliary laser lithotripsy has
been recognized to occur with several laser types. This phenomenon
has also been seen with Q-switched Nd:YAG laser lithotripsy. Our aim
was to determine the biological consequences of this event in the
canine ureter and bile duct.
In an excised tissue preparation, urinary and biliary stones were
impacted in a canine ureter and common bile duct. Three and four
hundred micron quartz laser fibers were placed in direct contact with
the stone. Normal saline coaxial irrigation was initiated at 75
xal/min. A Q-switched Nd:YAG laser was activated at repetition rates
from 1O-30 Hz. and pulse energies from 10-30 mJ. The tissue was
sectioned and microscopic examination of the fragmentation site was
Histological exam revealed the persistence of large numbers of fiber
fragments in the lumen and imnbedded in the epithelium at the
lithotripsy site. Fragments varied greatly in size and appeared to
have angular, sharp edges
We conclude that irrigation can not be relied upon to remove the
fiber fragments from the lithotripsy sites and that the biological
consequences of fiber fragmentation may be greater than previously
believed. Glass fragments "blown" into the epitheliumu will often
result in glass granulomas, which may eventually cause obstruction of
closed lumninal structures. This raises serious concern for the
presence of any fiber breakage during laser lithotripsy.
An optical system for digitized imaging (OSDI) consisting of a computer-controlled charge-coupled-device (CCD) array with 576 x 384 detection elements and 1.4 bits of digitization is used for in-vivo measurements of light distributions from diffusing tips of optical fibers in tumors. A macro lens assembly allows imaging with a spatial resolution of approximately 46 pm. Radiation-induced fibrosarcoma were implanted and grown subcutaneously in C3H mice. A fiber with a cylindrical diffuser tip (1.5 x 5 mm) was inserted into tumors. Tumor light intensity distribution images were made with the OSDI at wavelengths varying from 457.9 to 800 nm with argon, argon-dye and titanium-sapphire lasers. These tumor images show that the volume of tumor exposed to actinic light intensities increases for wavelengths between 400 and 800 run and reaches a maximum at about 800 nm. The uniformity of light distribution also increases at longer wavelengths. At shorter wavelengths, blood vessels in the tumor are clearly delineated as dark lines and networks of lines that might shield sensitized tumor cells from adequate light exposure. The light-acti.vated drug, Photofrin II (PF II), 20 mg/kgbw, was administered intravenously to anesthetized mice. With optical excitation at 457.9 nm consecutive (0, 1., 2.5, 4 hours) fluorescence-only images were made of PF II fluorescence distribution in the tumor and surrounding the diffusing optical fiber. Serial images after PF II injection showed drug fluorescence increasing with time in the tumor around the fiber. The OSDI provides a way to measure actual light intensity distributions and could be used in vivo to guide adjustments in light intensity and drug distributions before and during tumor phototherapy.