Proteasome Inhibitors Block DNA Repair and Radiosensitize Non-Small Cell Lung Cancer

Despite optimal radiation therapy (RT), chemotherapy and/or surgery, a majority of patients with locally advanced non-small cell lung cancer (NSCLC) fail treatment. To identify novel gene targets for improved tumor control, we performed whole genome RNAi screens to identify knockdowns that most reproducibly increase NSCLC cytotoxicity. These screens identified several proteasome subunits among top hits, including the topmost hit PSMA1, a component of the core 20 S proteasome. Radiation and proteasome inhibition showed synergistic effects. Proteasome inhibition resulted in an 80–90% decrease in homologous recombination (HR), a 50% decrease in expression of NF-κB-inducible HR genes BRCA1 and FANCD2, and a reduction of BRCA1, FANCD2 and RAD51 ionizing radiation-induced foci. IκBα RNAi knockdown rescued NSCLC radioresistance. Irradiation of mice with NCI-H460 xenografts after inducible PSMA1 shRNA knockdown markedly increased murine survival compared to either treatment alone. Proteasome inhibition is a promising strategy for NSCLC radiosensitization via inhibition of NF-κB-mediated expression of Fanconi Anemia/HR DNA repair genes.American Society for Radiation Oncology (Junior Faculty Career Research Training Award)Harvard University. Joint Center for Radiation Therapy (Foundation Grant)Dana-Farber/Harvard Cancer Center (SPORE Developmental Research Project Award in Lung Cancer Research)National Cancer Institute (U.S.) (Award K08CA172354

The proteasome inhibitor MG-132 sensitizes PC-3 prostate cancer cells to ionizing radiation by a DNA-PK-independent mechanism

BACKGROUND: By modulating the expression levels of specific signal transduction molecules, the 26S proteasome plays a central role in determining cell cycle progression or arrest and cell survival or death in response to stress stimuli, including ionizing radiation. Inhibition of proteasome function by specific drugs results in cell cycle arrest, apoptosis and radiosensitization of many cancer cell lines. This study investigates whether there is also a concomitant increase in cellular radiosensitivity if proteasome inhibition occurs only transiently before radiation. Further, since proteasome inhibition has been shown to activate caspase-3, which is involved in apoptosis, and caspase-3 can cleave DNA-PKcs, which is involved in DNA-double strand repair, the hypothesis was tested that caspase-3 activation was essential for both apoptosis and radiosensitization following proteasome inhibition. METHODS: Prostate carcinoma PC-3 cells were treated with the reversible proteasome inhibitor MG-132. Cell cycle distribution, apoptosis, caspase-3 activity, DNA-PKcs protein levels and DNA-PK activity were monitored. Radiosensitivity was assessed using a clonogenic assay. RESULTS: Inhibition of proteasome function caused cell cycle arrest and apoptosis but this did not involve early activation of caspase-3. Short-time inhibition of proteasome function also caused radiosensitization but this did not involve a decrease in DNA-PKcs protein levels or DNA-PK activity. CONCLUSION: We conclude that caspase-dependent cleavage of DNA-PKcs during apoptosis does not contribute to the radiosensitizing effects of MG-132

The genomic landscape of balanced cytogenetic abnormalities associated with human congenital anomalies

Despite the clinical significance of balanced chromosomal abnormalities (BCAs), their characterization has largely been restricted to cytogenetic resolution. We explored the landscape of BCAs at nucleotide resolution in 273 subjects with a spectrum of congenital anomalies. Whole-genome sequencing revised 93% of karyotypes and demonstrated complexity that was cryptic to karyotyping in 21% of BCAs, highlighting the limitations of conventional cytogenetic approaches. At least 33.9% of BCAs resulted in gene disruption that likely contributed to the developmental phenotype, 5.2% were associated with pathogenic genomic imbalances, and 7.3% disrupted topologically associated domains (TADs) encompassing known syndromic loci. Remarkably, BCA breakpoints in eight subjects altered a single TAD encompassing MEF2C, a known driver of 5q14.3 microdeletion syndrome, resulting in decreased MEF2C expression. We propose that sequence-level resolution dramatically improves prediction of clinical outcomes for balanced rearrangements and provides insight into new pathogenic mechanisms, such as altered regulation due to changes in chromosome topology

A systematic review of upper extremity casting for children and adults with central nervous system motor disorders

Objective: To summarize evidence on the use of upper extremity casting designed to achieve reductions in contracture, tone, pain, function, oedema or spasticity in the elbow, wrist or hand of adults and children with neurological conditions. Data sources: A search was conducted of the Cochrane Database of Systematic Reviews; the electronic databases MEDLINE, EMBASE, CINAHL, PEDro, OT-Seeker; Google Scholar; reference lists of retrieved trial reports and review articles. Review methods: Two independent reviewers determined whether retrieved study abstracts met inclusion criteria: human subjects; \u3e50% of participants children or adults described as having brain injury, cerebral palsy or stroke. Methodological quality of randomized controlled trials was rated using the PEDro scale (1—10 highest). Results: Thirty-one papers were retrieved and 23 studies appraised: three were randomized controlled trials and four were systematic reviews. Over three-quarters of the studies, excluding systematic reviews, were lower level evidence (n = 4 level V; n = 4 level IV; n = 1 level III). Methodological quality of randomized controlled trials was high (PEDro 8, 8 and 9) and there were modest positive short-term outcomes for two trials, although they did not include no-stretch comparison conditions. Safety issues typically included pain or skin breakdown; two adverse events were not cast related. Conclusion: While theoretical rationales suggest upper limb casting should be effective there is insufficient high-quality evidence regarding impact or long-term effects to either support or abandon this practice. High variability in casting protocols indicates little consistency or consensus in practice. As maximum or low-load stretch are rationales for cast application, the absence of no-stretch conditions in existing trials is a major weakness in current evidence

Primitive Geometric Operations on Planar Algebraic Curves with Gaussian Approximations

We present a curve approximation method which approximates each planar algebraic curve segment by discrete curve points at each of which the curve has its gradient from a set of uniformly distributed normals. This method, called Gaussian Approximation (GAP), provides efficient algorithms for various primitive geometric operations, especially for those related with gradients such as common tangent and convolution computations, on planar algebraic curve segments. The hierarchy of unit gradients gives the corresponding hierarchy of CAP. The approximation error at each level of the hierarchy can be modeled in the representation of GAP itself, and we can use this structure to dynamically control the precision and efficiency of geometric computation with CAP. We implemented various primitive geometric operations on planar algebraic curve segments with GAP representations on SUN4/Sparc station using C