Indocyanine green binds to DOTAP liposomes for enhanced optical properties and tumor photoablation.

Indocyanine green (ICG) is a clinically-approved near infrared (NIR) dye used for optical imaging. The dye is only slightly soluble in water and is prone to aggregation in saline solutions, so that alternative formulations can improve photophysical performance. Numerous nanoscale formulations of ICG have been described in the literature, but we sought to develop an approach that does not require additional purification steps. Pre-formed liposomes incorporating 45 mol% of the cationic lipid 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP) rapidly bind ICG, resulting in enhanced NIR optical properties. ICG binding is dependent on the amount of DOTAP incorporated in the liposomes. A dye-to-lipid mass ratio of [0.5 : 25] is sufficient for full complexation, without additional purification steps following mixing. NIR absorption, fluorescence intensity, and photoacoustic signals are increased for the liposome-bound dye. Not only is the optical character enhanced by simple mixing of ICG with liposomes, but retention in 4T1 mammary tumors is observed following intratumor injection, as assessed by fluorescence and photoacoustic imaging. Subsequent photothermal therapy with 808 nm laser irradiation is effective and results in tumor ablation without regrowth for at least 30 days. Thus, ICG optical properties and photothermal ablation outcomes can be improved by mixing the dye with pre-formed DOTAP liposomes in conditions that result in full dye-binding to the liposomes.

Highly-Soluble Cyanine J-aggregates Entrapped by Liposomes for In Vivo Optical Imaging around 930 nm.

Near infrared (NIR) dyes are useful for in vivo optical imaging. Liposomes have been used extensively for delivery of diverse cargos, including hydrophilic cargos which are passively loaded in the aqueous core. However, most currently available NIR dyes are only slightly soluble in water, making passive entrapment in liposomes challenging for achieving high optical contrast. Methods: We modified a commercially-available NIR dye (IR-820) via one-step Suzuki coupling with dicarboxyphenylboronic acid, generating a disulfonated heptamethine; dicarboxyphenyl cyanine (DCP-Cy). DCP-Cy was loaded in liposomes and used for optical imaging. Results: Owing to increased charge in mildly basic aqueous solution, DCP-Cy had substantially higher water solubility than indocyanine green (by an order of magnitude), resulting in higher NIR absorption. Unexpectedly, DCP-Cy tended to form J-aggregates with pronounced spectral red-shifting to 934 nm (from 789 nm in monomeric form). J-aggregate formation was dependent on salt and DCP-Cy concentration. Dissolved at 20 mg/mL, DCP-Cy J-aggregates could be entrapped in liposomes. Full width at half maximum absorption of the liposome-entrapped dye was just 25 nm. The entrapped DCP-Cy was readily detectable by fluorescence and photoacoustic NIR imaging. Upon intravenous administration to mice, liposomal DCP-Cy circulated substantially longer than the free dye. Accumulation was largely in the spleen, which was visualized with fluorescence and photoacoustic imaging. Conclusions: DCP-Cy is simple to synthesize and exhibits high aqueous solubility and red-shifted absorption from J-aggregate formation. Liposomal dye entrapment is possible, which facilitates in vivo photoacoustic and fluorescence imaging around 930 nm.

Binding of an amphiphilic phthalocyanine to pre-formed liposomes confers light-triggered cargo release.

Liposomes are able to load a range of cargos and have been used for drug delivery applications, including for stimuli-triggered drug release. Here, we describe an approach for imparting near infrared (NIR) light-triggered release to pre-formed liposomes, using a newly-synthesized cationic, amphiphilic phthalocyanine. When simply mixed in aqueous solution with cargo-loaded liposomes, the cationic amphiphilic phthalocyanine, but not a cationic hydrophilic azaphthalocyanine, spontaneously incorporates into the liposome bilayer. This enables subsequent release of loaded cargo (doxorubcin or basic orange) upon irradiation with NIR light. The rate of release could be altered by varying the amount of photosensitizer added to the liposomes. In the absence of NIR light exposure, stable cargo loading of the liposomes was maintained. Introduction.

Multifunctional Liposomes for Image-Guided Intratumoral Chemo-Phototherapy.

Intratumoral (IT) drug injections reduce systemic toxicity, but delivered volumes and distribution can be inconsistent. To improve IT delivery paradigms, porphyrin-phospholipid (PoP) liposomes are passively loaded with three hydrophilic cargos: sulforhodamine B, a fluorophore; gadolinium-gadopentetic acid, a magnetic resonance (MR) agent; and oxaliplatin, a colorectal cancer chemotherapeutic. Liposome composition is optimized so that cargo is retained in serum and storage, but is released in less than 1 min with exposure to near infrared light. Light-triggered release occurs with PoP-induced photooxidation of unsaturated lipids and all cargos release concurrently. In subcutaneous murine colorectal tumors, drainage of released cargo is delayed when laser treatment occurs 24 h after IT injection, at doses orders of magnitude lower than systemic ones. Delayed light-triggering results in substantial tumor shrinkage relative to controls a week following treatment, although regrowth occurs subsequently. MR imaging reveals that over this time frame, pools of liposomes within the tumor migrate to adjacent regions, possibly leading to altered spatial distribution during triggered drug release. Although further characterization of cargo loading and release is required, this proof-of-principle study suggests that multimodal theranostic IT delivery approaches hold potential to both guide injections and interpret outcomes, in particular when combined with chemo-phototherapy.

Chemophototherapy: An Emerging Treatment Option for Solid Tumors.

Near infrared (NIR) light penetrates human tissues with limited depth, thereby providing a method to safely deliver non-ionizing radiation to well-defined target tissue volumes. Light-based therapies including photodynamic therapy (PDT) and laser-induced thermal therapy have been validated clinically for curative and palliative treatment of solid tumors. However, these monotherapies can suffer from incomplete tumor killing and have not displaced existing ablative modalities. The combination of phototherapy and chemotherapy (chemophototherapy, CPT), when carefully planned, has been shown to be an effective tumor treatment option preclinically and clinically. Chemotherapy can enhance the efficacy of PDT by targeting surviving cancer cells or by inhibiting regrowth of damaged tumor blood vessels. Alternatively, PDT-mediated vascular permeabilization has been shown to enhance the deposition of nanoparticulate drugs into tumors for enhanced accumulation and efficacy. Integrated nanoparticles have been reported that combine photosensitizers and drugs into a single agent. More recently, light-activated nanoparticles have been developed that release their payload in response to light irradiation to achieve improved drug bioavailability with superior efficacy. CPT can potently eradicate tumors with precise spatial control, and further clinical testing is warranted.

Mechanisms of Light-induced Liposome Permeabilization.

Liposomes have been widely studied for drug delivery applications. The inclusion of photoactive molecules into liposomes opens the possibility of light-controlled cargo release to enhance drug biodistribution or bioavailability at target sites. Membrane permeabilization induced by light can be an effective strategy for enhancing cargo delivery with spatial and temporal control, which holds potential for chemophototherapy approaches. Several diverse mechanisms have been reported including light-induced oxidation, photocrosslinking, photoisomerization, photocleavage and photothermal release. Here, we review selected recent reports of light-triggered cargo release from liposomes.

Reduced folate carrier-1 G80a gene polymorphism is associated with neuroblastoma's development.

Neuroblastoma is a malignant embryonal tumor of neural crest cells that give rise to the sympathetic nervous system, responsible for 10-70% of all cases of childhood cancer. Because of its early appearance, it has been suggested that risk factors active in the prenatal can be associated with the pathogenesis of neuroblastoma. The aim of this study was to investigate whether the genetic polymorphisms MTHFR C677T and A1298C, MTR A2756G, TYMS 2R/3R and SLC19A1 G80A, involved in folate metabolism, increase the risk of neuroblastoma in Brazilian children. This study comprised 31 Brazilian children (0-14 years old) diagnosed with neuroblastoma compared with 92 controls. Investigation of polymorphisms MTHFR C677T, MTR A2756G and SLC19A1 A80G was performed using PCR-RFLP, the TYMS 2R/3R using PCR and MTHFR A1298C using AS-PCR. The SLC19A1 A80A genotype was significantly associated with the development of neuroblastoma, compared with the control group (Williams G-Test = 0.0286; OR = 5.1667; 95% CI = 1.4481-18.4338; p = 0.0175). When analyzed together, the 80AG+AA genotypes showed a trend toward association (OR = 3.3033; 95% CI = 1.0586-10.3080; p = 0.0563). Our results suggest that individuals carriers of genotype AA for the SLC19A1 gene present risk for the development of neuroblastoma and possibly have difficulty in absorption of folic acid by the cells, and this may adversely affect the metabolism of folate causing genomic instability and promoting the development of cancer. This is the first retrospective/prospective study to examine the relationship between polymorphisms of folate pathway genes and risk of neuroblastoma.