Yonathan Daniel, MD

Photodynamic therapy (PDT) is an alternative treatment for cancer based on cellular uptake of a photosensitizer that, when illuminated with a laser at an appropriate wavelength, generates oxidative radical species that lead to cell death. Chlorin e6 (Ce6), a second generation photosensitizer, has an absorption peak between 650-680nm and is characterized by its large heterocyclic aromatic ring, high levels of π-π bond conjugation,and its hydrophobicity. The aforementioned absorption peak is within the PDT therapeutic window and is ideal for biological application due to the low coefficient of tissue absorption at this range. A significant hurdle to the clinical application of Ce6 is its poor solubility in water. Some research groups have conjugated hydrophilic side chains to Ce6 in an attempt to increase its solubility. In this study, we took superparamagnetic iron oxide (SPIO) nanoclusters and mixed them with Ce6 using an inverse emulsion technique to create novel Ce6-coated SPIO nanoparticles. The resultant nanoparticles greatly increase the solubility of Ce6 without the additional conjugation of side chains. Transmission Electron Microscopy, UV-vis spectroscopy, Nuclear Magnetic Resonance spectroscopy, Fluorescence spectroscopy, and Zeta potential measurements were completed to characterize and confirm the structure of the Ce6-SPIO nanoparticles. The encapsulation efficiency of both Ce6 and SPIO within the formulated nanoparticles is ~100% and the total Ce6 Payload is 40-20% of the total weight (Ce6 +SPIO). The Ce6-SPIO nanoparticles demonstrate robust stability in physiological conditions and are within the ideal size range for tumor uptake via the enhanced permeability and retention effect. Along with the potential PDT capabilities of Ce6, previous research has shown that the SPIO nanoclusters are detectable by MRI and can be used as contrast agents for pre-operative MRI imaging. This indicates that the formulated Ce6-SPIO nanoparticles can potentially be used as a single therapeutic modality for both tumor imaging and treatment. The future direction of these nanoparticles is to now test cytotoxicity in vitro & in vivo along with MRI imaging studies to confirm the detectability of the nanoparticles.

Neuroblastoma is one of the most common extracranial tumors found in juveniles. Because of its generalized symptoms and the fact that most cases develop during infancy, it is often diagnosed at a late stage where common treatments are less effective. In Neuroblastoma, MYC-N genomic amplification and segmental chromosomal alterations are associated with greater disease progression and poorer clinical outcome. Segmental alterations are the greatest predictor of relapse and are attributed to erroneous repair of chromosomal breaks through a low fidelity alternative-non homologous end joining (alt-NHEJ) repair pathway. Interestingly, in tumors using this pathway, repair enzymes present in the alt-NHEJ are upregulated while classical NHEJ repair enzymes are down regulated. Poly ADP Ribose Polymerase-1 (PARP-1) plays a critical role in the alt-NHEJ pathway by using N-terminus zinc fingers to detect breaks in the genome and then signaling other enzymes that consequently bind to, and repair, the single strand breakages (SSB). On this basis, we sought to test the hypothesis that inhibiting PARP-1 in vitro would reduce the comparative survivability of high-risk neuroblastoma cells. We used cell lines: SKNSH, IMR5, BE2, BE2C, NB1691, NLF, SY5Y due to their characterization of high-risk neuroblastoma and performed growth inhibition assays using four PARP-1 inhibitors: talazoparib, niraparib, olaparib, and veliparib. Our data revealed that although all the cell lines studied showed clear responses to PARP-1 inhibition (PARPi), there was clear variation in which inhibitors worked best. The cell lines showed greatest sensitivity to talazoparib followed by niraparib, olaparib, and then veliparib. From our data we can definitively conclude that there is a clear differential in vitro response to PARP-1 inhibitors. Additionally, we noted that cells with a P53 mutation were often less sensitive to PARPi, although more research is necessary to definitively claim that. In future studies, we hope to look at PARP-1 expression and compare that to relative PARPi sensitivity in neuroblastoma cells. Additionally, radioligand binding studies may be useful in further understanding the interaction between PARP-1 and its inhibitors.