Aaron M. Mohs

Bioconjugated quantum dots for in vivo molecular and cellular imaging.

Semiconductor quantum dots (QDs) are tiny light-emitting particles on the nanometer scale, and are emerging as a new class of fluorescent labels for biology and medicine. In comparison with organic dyes and fluorescent proteins, they have unique optical and electronic properties, with size-tunable light emission, superior signal brightness, resistance to photobleaching, and broad absorption spectra for simultaneous excitation of multiple fluorescence colors. QDs also provide a versatile nanoscale scaffold for designing multifunctional nanoparticles with both imaging and therapeutic functions. When linked with targeting ligands such as antibodies, peptides or small molecules, QDs can be used to target tumor biomarkers as well as tumor vasculatures with high affinity and specificity. Here we discuss the synthesis and development of state-of-the-art QD probes and their use for molecular and cellular imaging. We also examine key issues for in vivo imaging and therapy, such as nanoparticle biodistribution, pharmacokinetics, and toxicology.

Tuning the optical and electronic properties of colloidal nanocrystals by lattice strain

Strain can have a large influence on the properties of materials at the nanoscale. The effect of lattice strain on semiconductor devices has been widely studied, but its influence on colloidal semiconductor nanocrystals is still poorly understood. Here we show that the epitaxial deposition of a compressive shell (ZnS, ZnSe, ZnTe, CdS or CdSe) onto a soft nanocrystalline core (CdTe) to form a lattice-mismatched quantum dot can dramatically change the conduction and valence band energies of both the core and the shell. In particular, standard type-I quantum-dot behaviour is replaced by type-II behaviour, which is characterized by spatial separation of electrons and holes, extended excited-state lifetimes and giant spectral shifts. Moreover, the strain induced by the lattice mismatch can be used to tune the light emission--which displays narrow linewidths and high quantum yields--across the visible and near-infrared part of the spectrum (500-1,050 nm). Lattice-mismatched core-shell quantum dots are expected to have applications in solar energy conversion, multicolour biomedical imaging and super-resolution optical microscopy.

Hand-held Spectroscopic Device for In Vivo and Intraoperative Tumor Detection: Contrast Enhancement, Detection Sensitivity, and Tissue Penetration

Surgery is one of the most effective and widely used procedures in treating human cancers, but a major problem is that the surgeon often fails to remove the entire tumor, leaving behind tumor-positive margins, metastatic lymph nodes, and/or satellite tumor nodules. Here we report the use of a hand-held spectroscopic pen device (termed SpectroPen) and near-infrared contrast agents for intraoperative detection of malignant tumors, based on wavelength-resolved measurements of fluorescence and surface-enhanced Raman scattering (SERS) signals. The SpectroPen utilizes a near-infrared diode laser (emitting at 785 nm) coupled to a compact head unit for light excitation and collection. This pen-shaped device effectively removes silica Raman peaks from the fiber optics and attenuates the reflected excitation light, allowing sensitive analysis of both fluorescence and Raman signals. Its overall performance has been evaluated by using a fluorescent contrast agent (indocyanine green, or ICG) as well as a surface-enhanced Raman scattering (SERS) contrast agent (pegylated colloidal gold). Under in vitro conditions, the detection limits are approximately 2-5 × 10(-11) M for the indocyanine dye and 0.5-1 × 10(-13) M for the SERS contrast agent. Ex vivo tissue penetration data show attenuated but resolvable fluorescence and Raman signals when the contrast agents are buried 5-10 mm deep in fresh animal tissues. In vivo studies using mice bearing bioluminescent 4T1 breast tumors further demonstrate that the tumor borders can be precisely detected preoperatively and intraoperatively, and that the contrast signals are strongly correlated with tumor bioluminescence. After surgery, the SpectroPen device permits further evaluation of both positive and negative tumor margins around the surgical cavity, raising new possibilities for real-time tumor detection and image-guided surgery.

Contrast-enhanced MRI with new biodegradable macromolecular Gd(III) complexes in tumor-bearing mice

The structures of polydisulfide‐based biodegradable macromolecular Gd(III) complexes were modified to improve their in vivo retention time and MRI contrast enhancement. Steric hindrance was introduced around the disulfide bonds to control their access to free thiols in order to alter the degradation rate of the copolymers. Two new macromolecular agents, (Gd‐DTPA)‐cystine copolymers (GDCP) and (Gd‐DTPA)‐cystine diethyl ester copolymers (GDCEP), were prepared. Both agents were readily degraded in vitro and in vivo by the disulfide‐thiol exchange reaction, but at a slow rate. The introduction of COOH and COOEt groups slowed down the degradation of the copolymers in the incubation with 15 μM cysteine. Metabolic degradation products were identified by matrix‐assisted laser desorption/ionization time‐of‐flight (MALDI‐TOF) mass spectrometry in the urine samples from rats injected with the agents. The T1 relaxivity (r1) was 5.43 mM−1s−1 for GDCP, and 5.86 mM−1s−1 for GDCEP, respectively, at 3T. MRI contrast enhancement of both agents was studied in nude mice bearing MDA‐BM‐231 human breast carcinoma xenografts, on a Siemens Trio 3T scanner. The modified agents resulted in more significant contrast enhancement in the blood pool and tumor periphery than (Gd‐DTPA)‐cystamine copolymers (GDCC) and a low‐molecular‐weight control agent, Gd‐(DTPA‐BMA), at a dose of 0.1 mmol‐Gd/kg. The results demonstrate that the structural modification of the biodegradable macromolecular Gd(III) complexes resulted in a relatively slow degradation of the macromolecules and significantly improved in vivo contrast enhancement. The modified agents show promise for use in investigations of blood pool and cancer by contrast‐enhanced (CE) MRI. Magn Reson Med 53:835–842, 2005.

Gadolinium(III)-based blood-pool contrast agents for magnetic resonance imaging: status and clinical potential.

Blood-pool MRI contrast agents have enormous potential to aid sensitive magnetic resonance detection and yield definitive diagnostic data of cancer and diseases of the cardiovascular system. Many attempts have been initiated to design macromolecular gadolinium (Gd[III]) complexes as magnetic resonance imaging blood-pool contrast agents, as macromolecules do not readily diffuse across healthy vascular endothelium, and remain intravascular. Although extremely efficacious in detecting and characterizing pathologic tissue, clinical development of these agents has been limited by potential toxicity concerns from incomplete Gd(III) clearance. Recent innovative technologies, such as reversible protein-binding contrast agents and biodegradable macromolecular contrast agents, may be valuable alternatives that combine the effective imaging characteristics of an intravascular contrast agent and the safety of clinically approved low-molecular-weight Gd(III) chelates.

Indocyanine green-loaded nanoparticles for image-guided tumor surgery

Detecting positive tumor margins and local malignant masses during surgery is critical for long-term patient survival. The use of image-guided surgery for tumor removal, particularly with near-infrared fluorescent imaging, is a potential method to facilitate removing all neoplastic tissue at the surgical site. In this study we demonstrate a series of hyaluronic acid (HLA)-derived nanoparticles that entrap the near-infrared dye indocyanine green, termed NanoICG, for improved delivery of the dye to tumors. Self-assembly of the nanoparticles was driven by conjugation of one of three hydrophobic moieties: aminopropyl-1-pyrenebutanamide (PBA), aminopropyl-5β-cholanamide (5βCA), or octadecylamine (ODA). Nanoparticle self-assembly, dye loading, and optical properties were characterized. NanoICG exhibited quenched fluorescence that could be activated by disassembly in a mixed solvent. NanoICG was found to be nontoxic at physiologically relevant concentrations and exposure was not found to inhibit cell growth. Using an MDA-MB-231 tumor xenograft model in mice, strong fluorescence enhancement in tumors was observed with NanoICG using a fluorescence image-guided surgery system and a whole-animal imaging system. Tumor contrast with NanoICG was significantly higher than with ICG alone.

Characterization of tumor angiogenesis with dynamic contrast‐enhanced MRI and biodegradable macromolecular contrast agents in mice

The efficacy of polydisulfide‐based biodegradable macromolecular contrast agents for characterizing tumor angiogenesis was investigated in a mouse model using dynamic contrast‐enhanced MRI (DCE‐MRI). Biodegradable macromolecular MRI contrast agents, gadopentetate dimeglumine (Gd‐DTPA) cystamine copolymers (GDCC), and Gd‐DTPA cystine copolymers (GDCP), with molecular weights of 20 and 70 kDa were used in the study. Gadodiamide (Gd [DTPA‐BMA]) and albumin labeled with Gd‐DTPA [albumin‐(Gd‐DTPA)] were used as the controls. The DCE‐MRI studies were performed in nude mice bearing prostate tumor xenografts from the MDA‐PCa‐2b cell line. Tumor angiogenic kinetic parameters, including endothelial transfer coefficient (KPS), fractional tumor plasma volume (fPV), and permeability surface area product (PS), were estimated from the DCE‐MRI data using a two‐compartment model. The KPS and fPV values estimated by the biodegradable macromolecular contrast agents were between those estimated by Gd(DTPA‐BMA) and albumin‐(Gd‐DTPA). The parameters estimated by the agent with a slow degradation rate and high molecular weight, GDCP‐70 (KPS = 2.09 ± 0.50 ml/min/100 cc and fPV = 0.075 ± 0.021), were closer to those by albumin‐(Gd‐DTPA) (KPS = 1.43 ± 0.64 ml/min/100 cc and fPV = 0.044 ± 0.007) than by other agents with relatively low molecular weight or rapid degradation rate. The polydisulfide‐based biodegradable macromolecular contrast agents are promising for characterizing tumor vascularity and angiogenesis with DCE‐MRI. Magn Reson Med 60:1347–1352, 2008.

Applications of Nanotechnology to Imaging and Therapy of Brain Tumors

In the past decade, numerous advances in the understanding of brain tumor physiology, tumor imaging, and tumor therapy have been attained. In some cases, these advances have resulted from refinements of pre-existing technologies (eg, improvements of contrast-enhanced magnetic resonance imaging). In other instances, advances have resulted from development of novel technologies. The development of nanomedicine (ie, applications of nanotechnology to the field of medicine) is an example of the latter. In this review, the authors explain the principles that underlay nanoparticle design and function as well as the means by which nanoparticles can be used for imaging and therapy of brain tumors.

Image-guided tumor surgery: will there be a role for fluorescent nanoparticles?

Image-guided surgery (IGS) using fluorescent nanoparticles (NPs) has the potential to substantially impact patient treatment. The use of fluorescence imaging provides surgeons with real-time feedback on the location of diseased tissue using safe, low-cost imaging agents and instrumentation. Fluorescent NPs are likely to play a role as they are capable of taking advantage of the enhanced permeability and retention (EPR) effect and can be modified to avoid clearance, increase circulation time, and specifically target tumors. Clinical trials of IGS using the FDA-approved fluorophores indocyanine green and methylene blue have already shown preliminary successes, and incorporation of fluorescent NPs will likely improve detection by providing higher signal to background ratio and reducing false-positive rates through active targeting. Preclinical development of fluorescent NP formulations is advancing rapidly, with strategies ranging from passive targeting to active targeting of cell surface receptors, creating pH-responsive NPs, and increasing cell uptake through cleavable proteins. This collective effort could lead to clinical trials using fluorescent NPs in the near future. WIREs Nanomed Nanobiotechnol 2016, 8:498-511. doi: 10.1002/wnan.1381 For further resources related to this article, please visit the WIREs website.

Near infrared fluorescent nanoparticles based on hyaluronic acid: Self-assembly, optical properties, and cell interaction.

UNLABELLED Fluorescent imaging agents that can specifically highlight tumor cells could have a significant impact on image-guided tumor removal. Here, fluorescent nanoparticles (NPs) derived from hyaluronic acid (HA) are investigated. HA is a ligand for the receptor CD44, which is a common biomarker present on many primary tumor cells, cancer-initiating cells, and tumor-associated fibroblasts. In addition, a family of enzymes that degrade HA, called hyaluronidases (HYALs), are also overexpressed with increased activity in many tumors. We report the design and development of a panel of targeted imaging agents using the near-infrared (NIR) dye, Cy7.5, that was directly conjugated to hydrophobically-modified HA. Two different molecular weights of HA, 10kDa and 100kDa, and three different degrees of hydrophobic moiety conjugation (0, 10, and 30mol%) were utilized to develop a panel of NPs with variable size that ranged from 50 to 400nm hydrodynamic diameter (HD) depending HA molecular weight, extent of fluorescence quenching (25-50%), kinetics of cellular uptake, and targeting to CD44+ cells. The kinetics and energy-dependence of cellular uptake in breast and prostate cancer cell lines, MDA-MB 231 and PC-3 cells, respectively, showed increased uptake with longer incubation times (at 4 and 8h compared to 1h), as well as uptake at 37°C but not 4°C, which indicated energy-dependent endocytosis. NP uptake studies in the presence of excess free HA showed that pre-treatment of cells with excess high molecular weight (MW) free HA decreased NP uptake by up to 50%, while no such trend was observed with low MW HA. These data lay the foundation for selection of optimized HA-derived NPs for image-guided surgery. STATEMENT OF SIGNIFICANCE Here, hyaluronic acid (HA), a well-studied biomacromolecule, is modified with a near infrared fluorophore and a hydrophobic moiety. The significance of this work, especially for imaging applications, is that the impact of HA molecular weight and the hydrophobic moiety conjugation degree on fluorescence and cell interaction can be predicted. With respect to existing literature, the eventual use of these HA-based NPs is image-guided surgery; thus, we focus on the dye, Cy7.5, for conjugation, which is more NIR than most existing HA literature. Furthermore, HA is a ligand for CD44, which is associated with cancer and tumor microenvironment cells. Systematic studies in this work highlight that HA can be tuned to maximize or minimize CD44 binding.