Light-induction of an anionic semiquinone (SQ) flavin radical in Drosophila cryptochrome (dCRY) alters the dCRY conformation to promote binding and degradation of the circadian clock protein Timeless (TIM). Specific peptide ligation with sortase A attaches a nitroxide spin-probe to the dCRY C-terminal tail (CTT) while avoiding deleterious side reactions. Pulse dipolar electron-spin resonance spectroscopy from the CTT nitroxide to the SQ shows that flavin photoreduction shifts the CTT ~1 nm and increases its motion, without causing full displacement from the protein. dCRY engineered to form the neutral SQ serves as a dark-state proxy to reveal that the CTT remains docked when the flavin ring is reduced but uncharged. Substitutions of flavin-proximal His378 promote CTT undocking in the dark or diminish undocking in the light, consistent with molecular dynamics simulations and TIM degradation activity. The His378 variants inform on recognition motifs for dCRY cellular turnover and strategies for developing optogenetic tools.

Prostate cancer, once it has progressed from its local to metastatic form, is a disease with poor prognosis and limited treatment options. Here we demonstrate an approach using nanoscale liposomes conjugated with E-selectin adhesion protein and Apo2L/TRAIL (TNF-related apoptosis-inducing ligand) apoptosis ligand that attach to the surface of leukocytes and rapidly clear viable cancer cells from circulating blood, both in human blood samples and in the living mouse. For the first time, it is shown that such an approach can be used to prevent the spontaneous formation and growth of metastatic tumors in an orthotopic xenograft model of prostate cancer, by greatly reducing the number of circulating tumor cells. We conclude that the use of circulating leukocytes as a carrier for the anti-cancer protein TRAIL could be an effective tool to directly target circulating tumor cells for the prevention of prostate cancer metastasis, and potentially other cancers that spread through the bloodstream.

Tumor draining lymph nodes are the first site of metastasis in most types of cancer. The extent of metastasis in the lymph nodes is often used in staging cancer progression. We previously showed that nanoscale TRAIL liposomes conjugated to human natural killer cells enhance their endogenous therapeutic potential in killing cancer cells cultured in engineered lymph node microenvironments. In this work, it is shown that liposomes decorated with apoptosis-inducing ligand TRAIL and an antibody against a mouse natural killer cell marker are carried to the tumor draining inguinal lymph nodes and prevent the lymphatic spread of a subcutaneous tumor in mice. It is shown that targeting natural killer cells with TRAIL liposomes enhances their retention time within the tumor draining lymph nodes to induce apoptosis in cancer cells. It is concluded that this approach can be used to kill cancer cells within the tumor draining lymph nodes to prevent the lymphatic spread of cancer.

Photodynamic therapy (PDT) is a promising non-invasive therapeutic modality that has been proposed for treating prostate cancer, but the procedure is associated with limited efficacy, tumor recurrence and photo-toxicity. In the present study, we proposed to develop a novel multifunctional nano-platform for targeted delivery of heat, reactive oxygen species (ROS) and heat shock protein 90 (Hsp90) inhibitor simultaneously for combination therapy against prostate cancer. This new nano-platform combines two newly developed entities: 1) a unique organic and biocompatible nanoporphyrin-based drug delivery system that can generate efficient heat and ROS simultaneously with light activation at the tumor sites for dual-modal photothermal- and photodynamic- therapy (PTT/PDT), and 2) new nano-formulations of Hsp90 inhibitors that can decrease the levels of pro-survival and angiogenic signaling molecules induced by phototherapy, therefore, further sensitizing cancer cells to phototherapy. Furthermore, the nanoparticles have activatable near infrared (NIR) fluorescence for optical imaging to conveniently monitor the real-time drug delivery in both subcutaneous and orthotopic mouse models bearing prostate cancer xenograft. This novel multifunctional nano-platform has great potential to improve the care of prostate cancer patients through targeted combination therapy.

The World Health Organization (WHO) recently reported that the total number of global cancer cases in 2013 reached 14 million, a 10% rise since 2008, while the total number of cancer deaths reached 8.2 million, a 5.2% increase since 2008. Metastasis is the major cause of death from cancer, accounting for 90% of all cancer related deaths. Tumor-draining lymph nodes (TDLN), the sentinel nodes, are the first organs of metastasis in several types of cancers. The extent of metastasis in the TDLN is often used in disease staging and prognosis evaluation in cancer patients. Here, we describe the microenvironment of the TDLN and review the recent literature on liposome-based therapies directed to immune cells within the TDLN with the intent to target cancer cells.