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Previous research has shown that gold nanoparticles can increase the effectiveness of radiation on cancer cells. Improved radiation effectiveness would allow lower radiation doses given to patients, reducing adverse effects; alternatively, it would provide more cancer killing at current radiation doses. Damage from radiation and gold nanoparticles depends in part on the Auger effect, which is very localized; thus, it is important to place the gold nanoparticles on or in the cancer cells. In this work, we use the pH-sensitive, tumor-targeting agent, pH Low-Insertion Peptide (pHLIP), to tether 1.4-nm gold nanoparticles to cancer cells. We find that the conjugation of pHLIP to gold nanoparticles increases gold uptake in cells compared with gold nanoparticles without pHLIP, with the nanoparticles distributed mostly on the cellular membranes. We further find that gold nanoparticles conjugated to pHLIP produce a statistically significant decrease in cell survival with radiation compared with cells without gold nanoparticles and cells with gold alone. In the context of our previous findings demonstrating efficient pHLIP-mediated delivery of gold nanoparticles to tumors, the obtained results serve as a foundation for further preclinical evaluation of dose enhancement.

Purpose. Acidification of extracellular space promotes tumor development, progression and invasiveness. pH (Low) Insertion Peptides (pHLIP® peptides) belong to the class of pH-sensitive membrane peptides, which target acidic tumors and deliver imaging and/or therapeutic agents to cancer cells within tumors. Procedures. Ex vivo fluorescent imaging of tissue and organs collected at various time points after administration of different pHLIP® variants conjugated with fluorescent dyes of various polarity was performed. Methods of multivariate statistical analyses were employed to establish classification between fluorescently-labeled pHLIP® variants in multidimensional space of spectral parameters. Results. The fluorescently-labeled pHLIP® variants were classified based on their biodistribution profile and ability of targeting of primary tumors. Also, sub-millimeter sized metastatic lesions in lungs were identified by ex vivo imaging after intravenous administration of fluorescent pHLIP® peptide. Conclusions. Different cargo molecules conjugated with pHLIP® peptides can alter biodistribution and tumor targeting. The obtained knowledge is essential for the design of novel pHLIP® based diagnostic and therapeutic agents targeting primary tumors and metastatic lesions.

Cerebral palsy (CP) is caused by a variety of factors attributed to early brain damage, resulting in permanently impaired motor control, marked by weakness and muscle stiffness. To find out if altered physiology of spinal motoneurons (MNs) could contribute to movement deficits, we performed whole-cell patch-clamp in neonatal rabbit spinal cord slices after developmental injury at 79% gestation. After preterm hypoxia-ischemia (HI), rabbits are born with motor deficits consistent with a spastic phenotype including hypertonia and hyperreflexia. There is a range in severity, thus kits are classified as severely affected, mildly affected, or unaffected based on modified Ashworth scores and other behavioral tests. At postnatal day (P)0-5, we recorded electrophysiological parameters of 40 MNs in transverse spinal cord slices using whole-cell patch-clamp. We found significant differences between groups (severe, mild, unaffected and sham control MNs). Severe HI MNs showed more sustained firing patterns, depolarized resting membrane potential, and fired action potentials at a higher frequency. These properties could contribute to muscle stiffness, a hallmark of spastic CP. Interestingly altered persistent inward currents (PICs) and morphology in severe HI MNs would dampen excitability (depolarized PIC onset and increased dendritic length). In summary, changes we observed in spinal MN physiology likely contribute to the severity of the phenotype, and therapeutic strategies for CP could target the excitability of spinal MNs.