Chulhong Kim

Gold nanocages covered by smart polymers for controlled release with near-infrared light

Photosensitive caged compounds have enhanced our ability to address the complexity of biological systems by generating effectors with remarkable spatial/temporal resolutions1-3. The caging effect is typically removed by photolysis with ultraviolet light to liberate the bioactive species. Although this technique has been successfully applied to many biological problems, it suffers from a number of intrinsic drawbacks. For example, it requires dedicated efforts to design and synthesize a precursor compound to the effector. The ultraviolet light may cause damage to biological samples and is only suitable for in vitro studies because of its quick attenuation in tissue4. Here we address these issues by developing a platform based on the photothermal effect of gold nanocages. Gold nanocages represent a class of nanostructures with hollow interiors and porous walls5. They can have strong absorption (for the photothermal effect) in the near-infrared (NIR) while maintaining a compact size. When the surface of a gold nanocage is covered with a smart polymer, the pre-loaded effector can be released in a controllable fashion using a NIR laser. This system works well with various effectors without involving sophiscated syntheses, and is well-suited for in vivo studies due to the high transparency of soft tissue in NIR6.

In Vivo Molecular Photoacoustic Tomography of Melanomas Targeted by Bioconjugated Gold Nanocages

Early diagnosis, accurate staging, and image-guided resection of melanomas remain crucial clinical objectives for improving patient survival and treatment outcomes. Conventional techniques cannot meet this demand because of the low sensitivity, low specificity, poor spatial resolution, shallow penetration, and/or ionizing radiation. Here we overcome such limitations by combining high-resolution photoacoustic tomography (PAT) with extraordinarily optical absorbing gold nanocages (AuNCs). When bioconjugated with [Nle(4),D-Phe(7)]-alpha-melanocyte-stimulating hormone, the AuNCs can serve as a novel contrast agent for in vivo molecular PAT of melanomas with both exquisite sensitivity and high specificity. The bioconjugated AuNCs enhanced contrast approximately 300% more than the control, PEGylated AuNCs. The in vivo PAT quantification of the amount of AuNCs accumulated in melanomas was further validated with inductively coupled plasma mass spectrometry (ICP-MS).

Near-Infrared Gold Nanocages as a New Class of Tracers for Photoacoustic Sentinel Lymph Node Mapping on a Rat Model

This work demonstrated the use of Au nanocages as a new class of lymph node tracers for noninvasive photoacoustic (PA) imaging of a sentinel lymph node (SLN). Current SLN mapping methods based on blue dye and/or nanometer-sized radioactive colloid injection are intraoperative due to the need for visual detection of the blue dye and low spatial resolution of Geiger counters in detecting radioactive colloids. Compared to the current methods, PA mapping based on Au nanocages shows a number of attractive features: noninvasiveness, strong optical absorption in the near-infrared region (for deep penetration), and the accumulation of Au nanocages with a higher concentration than the initial solution for the injection. In an animal model, these features allowed us to identify SLNs containing Au nanocages as deep as 33 mm below the skin surface with good contrast. Most importantly, compared to methylene blue Au nanocages can be easily bioconjugated with antibodies for targeting specific receptors, potentially eliminating the need for invasive axillary staging procedures in addition to providing noninvasive SLN mapping.

Deeply penetrating in vivo photoacoustic imaging using a clinical ultrasound array system

Using a hand-held photoacoustic probe integrated with a clinical ultrasound array system, we successfully imaged objects deeply positioned in biological tissues. The optical contrasts were enhanced by methylene blue with a concentration of ~30 mM. The penetration depth reached ~5.2 cm in chicken breast tissue by using 650-nm wavelength, which is ~4.7 times the 1/e optical penetration depth. This imaging depth was achieved using a laser fluence on the tissue surface of only 3 mJ/cm2, which is 1/7 of the American National Standards Institute (ANSI) safety limit (20 mJ/cm2). The noise equivalent sensitivity at this depth was ~11 mM. Further, after intradermal injection of methylene blue in a rat, a sentinel lymph node was easily detected in vivo, beneath a 2-cm thick layer of chicken breast. Also, blood located 3.5 cm deep in the rat was clearly imaged with intrinsic contrast. We have photoacoustically guided insertion of a needle into a rat sentinel lymph node with accumulated methylene blue. These results highlight the clinical potential of photoacoustic image-guided identification and needle biopsy of sentinel lymph nodes for axillary staging in breast cancer patients.

Sentinel Lymph Nodes and Lymphatic Vessels: Noninvasive Dual-Modality in Vivo Mapping by Using Indocyanine Green in Rats—Volumetric Spectroscopic Photoacoustic Imaging and Planar Fluorescence Imaging

PURPOSEnTo noninvasively map sentinel lymph nodes (SLNs) and lymphatic vessels in rats in vivo by using dual-modality nonionizing imaging-volumetric spectroscopic photoacoustic imaging, which measures optical absorption, and planar fluorescence imaging, which measures fluorescent emission-of indocyanine green (ICG).nnnMATERIALS AND METHODSnInstitutional animal care and use committee approval was obtained. Healthy Sprague-Dawley rats weighing 250-420 g (age range, 60-120 days) were imaged by using volumetric photoacoustic imaging (n = 5) and planar fluorescence imaging (n = 3) before and after injection of 1 mmol/L ICG. Student paired t tests based on a logarithmic scale were performed to evaluate the change in photoacoustic signal enhancement of SLNs and lymphatic vessels before and after ICG injection. The spatial resolutions of both imaging systems were compared at various imaging depths (2-8 mm) by layering additional biologic tissues on top of the rats in vivo. Spectroscopic photoacoustic imaging was applied to identify ICG-dyed SLNs.nnnRESULTSnIn all five rats examined with photoacoustic imaging, SLNs were clearly visible, with a mean signal enhancement of 5.9 arbitrary units (AU) + or - 1.8 (standard error of the mean) (P < .002) at 0.2 hour after injection, while lymphatic vessels were seen in four of the five rats, with a signal enhancement of 4.3 AU + or - 0.6 (P = .001). In all three rats examined with fluorescence imaging, SLNs and lymphatic vessels were seen. The average full width at half maximum (FWHM) of the SLNs in the photoacoustic images at three imaging depths (2, 6, and 8 mm) was 2.0 mm + or - 0.2 (standard deviation), comparable to the size of a dissected lymph node as measured with a caliper. However, the FWHM of the SLNs in fluorescence images widened from 8 to 22 mm as the imaging depth increased, owing to strong light scattering. SLNs were identified spectroscopically in photoacoustic images.nnnCONCLUSIONnThese two modalities, when used together with ICG, have the potential to help map SLNs in axillary staging and to help evaluate tumor metastasis in patients with breast cancer.

Noninvasive in vivo spectroscopic nanorod-contrast photoacoustic mapping of sentinel lymph nodes

Sentinel lymph node (SLN) biopsy has increasingly become important in axillary staging of breast cancer patients since SLN biopsy alleviates the postoperative complications of previously practiced axillary lymph node dissections. Nevertheless, the procedures of SLN biopsy using blue dye and radioactive substance are still intraoperative, and the latter methods are also ionizing. In this pilot study, we have proposed noninvasive in vivo spectroscopic photoacoustic (PA) SLN mapping using gold nanorods as lymph node tracers in a rat model. Gold nanorods have biocompatibility, high optical absorption, and easily tuned surface plasmon resonance peak wavelength.

Handheld array-based photoacoustic probe for guiding needle biopsy of sentinel lymph nodes

By modifying a clinical ultrasound array system, we develop a novel handheld photoacoustic probe for image-guided needle biopsy. The integration of optical fiber bundles for pulsed laser light delivery enables photoacoustic image-guided insertion of a needle into rat axillary lymph nodes with accumulated indocyanine green (ICG). Strong photoacoustic contrast of the needle is achieved. After subcutaneous injection of the dye in the left forepaw, sentinel lymph nodes are easily detected, in vivo and in real time, beneath 2-cm-thick chicken breast overlaying the axillary region. ICG uptake in axillary lymph nodes is confirmed with fluorescence imaging both in vivo and ex vivo. These results demonstrate the clinical potential of this handheld photoacoustic system for facile identification and needle biopsy of sentinel lymph nodes for cancer staging and metastasis detection in humans.

Gold nanocages covered with thermally-responsive polymers for controlled release by high-intensity focused ultrasound

This paper describes the use of Au nanocages covered with smart, thermally-responsive polymers for controlled release with high-intensity focused ultrasound (HIFU). HIFU is a highly precise medical procedure that uses focused ultrasound to heat and destroy pathogenic tissue rapidly and locally in a non-invasive or minimally invasive manner. The released dosage could be remotely controlled by manipulating the power of HIFU and/or the duration of exposure. We demonstrated localized release within the focal volume of HIFU by using gelatin phantom samples containing dye-loaded Au nanocages. By placing chicken breast tissues on top of the phantoms, we further demonstrated the feasibility of this system for controlled release at depths up to 30 mm. Because it can penetrate more deeply into soft tissues than near-infrared light, HIFU is a potentially more effective external stimulus for rapid, on-demand drug release.

Multifunctional microbubbles and nanobubbles for photoacoustic and ultrasound imaging

We develop a novel dual-modal contrast agent-encapsulated-ink poly(lactic-co-glycolic acid) (PLGA) microbubbles and nanobubbles-for photoacoustic and ultrasound imaging. Soft gelatin phantoms with embedded tumor simulators of encapsulated-ink PLGA microbubbles and nanobubbles in various concentrations are clearly shown in both photoacoustic and ultrasound images. In addition, using photoacoustic imaging, we successfully image the samples positioned below 1.8-cm-thick chicken breast tissues. Potentially, simultaneous photoacoustic and ultrasound imaging enhanced by encapsulated-dye PLGA microbubbles or nanobubbles can be a valuable tool for intraoperative assessment of tumor boundaries and therapeutic margins.

In vivo photoacoustic mapping of lymphatic systems with plasmon-resonant nanostars

Plasmon-resonant nanostars (NSTs) provide excellent contrast enhancement for photoacoustic tomography. The high photoacoustic sensitivity of NSTs at near-infrared wavelengths enable their in vivo detection in rat sentinel lymph nodes and vessels, with direct application toward lymphangiography.