An image-guided precision proton radiation platform for preclinical in vivo research.

There are many unknowns in the radiobiology of proton beams and other particle beams. We describe the development and testing of an image-guided low-energy proton system optimized for radiobiological research applications. A 50 MeV proton beam from an existing cyclotron was modified to produce collimated beams (as small as 2 mm in diameter). Ionization chamber and radiochromic film measurements were performed and benchmarked with Monte Carlo simulations (TOPAS). The proton beam was aligned with a commercially-available CT image-guided x-ray irradiator device (SARRP, Xstrahl Inc.). To examine the alternative possibility of adapting a clinical proton therapy system, we performed Monte Carlo simulations of a range-shifted 100 MeV clinical beam. The proton beam exhibits a pristine Bragg Peak at a depth of 21 mm in water with a dose rate of 8.4 Gy min-1 (3 mm depth). The energy of the incident beam can be modulated to lower energies while preserving the Bragg peak. The LET was: 2.0 keV µm-1 (water surface), 16 keV µm-1 (Bragg peak), 27 keV µm-1 (10% peak dose). Alignment of the proton beam with the SARRP system isocenter was measured at 0.24 mm agreement. The width of the beam changes very little with depth. Monte Carlo-based calculations of dose using the CT image data set as input demonstrate in vivo use. Monte Carlo simulations of the modulated 100 MeV clinical proton beam show a significantly reduced Bragg peak. We demonstrate the feasibility of a proton beam integrated with a commercial x-ray image-guidance system for preclinical in vivo studies. To our knowledge this is the first description of an experimental image-guided proton beam for preclinical radiobiology research. It will enable in vivo investigations of radiobiological effects in proton beams.

SU-E-T-437: Interfractional Dosimetric Verification of Lung Patients Treated by Passive Double Scattering Proton Radiotherapy.

PURPOSE: Proton radiotherapy, with the ability to confine the dose at desired depth, can potentially benefit lung tumor patients by significantly sparing the healthy lung tissue. However, the superior proton dose distribution could be affected by tumor shrinkage due to the quick response and by motion especially related to the respiration. Thus the treatment should be frequently verified and be adjusted accordingly if necessary to achieve the initial treatment goal. MATERIAL AND METHODS: A cohort of 20 patients were selected from lung patients treated with passive proton radiotherapy. All those patients were evaluated via 4D-CT scans and found to have tumor motion less than 1 cm. The internal target volumes (ITV) were derived based on the full inspiration and expiration phases. The average of the 4D-CT scan, full inspiration and expiration phases were used for the initial treatment planning. The planning objective was 95% of the prescription dose to at least 95% volume of the ITV. Bi-weekly verification 4D-CT scans were performed to assess the robustness of the initial treatment plan and no replanning was required for target dose variations less then 3%. RESULTS: Compared with the initial treatment plan, the standard deviations of target coverage on inspiration, expiration, and average verification CT scans are within 3% for all the patients, with the maximum difference up to 7%. No statistically significant differences were found among the initial and verification plans (p>0.1). The percentage deviations of OAR sparing were highly variable, e.g., up to 40% for mean lung dose, 100% for mean heart dose, 50% for max cord dose, particularly for OARs receiving small amount of doses. However, the absolute dose deviations are all with OAR's tolerance. CONCLUSION: Overall, the passive double scattering proton modality allows for robust proton treatment planning and delivery to treat the lung tumors with limited motion.

SU-D-211-06: The Effect of Motion in RapidArc Lung SBRT Delivery.

PURPOSE: RapidArc is routinely used for stereotactic radiotherapy for lung cancer. While treatment dose is optimized and calculated on a static CT image, the motion of the target in conjunction with the motion of the MLC may Result in a delivered dose deviating from the planed dose. In this study, we investigate the dosimetric consequences of the inter-play effect by simulating dynamic dose delivery on a dynamic CT dataset of real patients. METHODS: The target motion in 20 patients was analyzed and 5 patients with >10 mm motion were chosen for this study. The RapidArc plan for eachpatient is optimized on a free-breathing CT using 2 arcs. Inherent in each plan is data on the associated parameters such as timestamp, MLC leave position, gantry angle and delivered beam MUs for each control point. Simulated dynamic delivery is performed by associating these parameters with each of the breathing phases of the 4D-CT. The starting breathing phase is selected randomly for each of the two arcs. Dose from the derived partial plans associated with each phase of the 4D-CT dose is recalculated in Eclipse. Accumulation of dose is performed using deformable image registration from each phase of the 4D-CT to the exhale phase of the 4D-CT. RESULTS: The coverage of the GTV and PTV shows negligible variations from the interplay effect. But the Homogeneity Index is affected by the motion. The prescription isodose volume is smaller than what was from the treatment plan dose. There were both intra- and inter-fraction effects seen inthe OARs dose in some patients. CONCLUSIONS: We investigated the motioneffect in RapidArc Lung SBRT delivery in 5 patients. Negligible variations were shown for target coverage. However the motion effects were observed in high dose distribution and volume. Some OARs dose distributions were affected by the motion.

Actin cytoskeletal function is spared, but apoptosis is increased, in WAS patient hematopoietic cells.

Mutations in the Wiskott-Aldrich syndrome protein (WASP) have been hypothesized to cause defective actin cytoskeletal function. This resultant dysfunction of the actin cytoskeleton has been implicated in the pathogenesis of Wiskott-Aldrich syndrome (WAS). In contrast, it was found that stimulated actin polymerization is kinetically normal in the hematopoietic lineages affected in WAS. It was also found that the actin cytoskeleton in WAS platelets is capable of producing the hallmark cytoarchitectural features associated with activation. Further analysis revealed accelerated cell death in WAS lymphocytes as evidenced by increased caspase-3 activity. This increased activity resulted in accelerated apoptosis of these cells. CD95 expression was also increased in these cells, suggesting an up-regulation in the FAS pathway in WAS lymphocytes. Additionally, inhibition of actin polymerization in lymphocytes using cytochalasin B did not accelerate apoptosis in these cells. This suggests that the accelerated apoptosis observed in WAS lymphocytes was not secondary to an underlying defect in actin polymerization caused by mutation of the WAS gene. These data indicate that WASP does not play a universal role in signaling actin polymerization, but does play a role in delaying cell death. Therefore, the principal consequence of mutations in the WAS gene is to accelerate lymphocyte apoptosis, potentially through up-regulation of the FAS-mediated cell death pathway. This accelerated apoptosis may ultimately give rise to the clinical manifestations observed in WAS. (Blood. 2000;95:1283-1292)

Molecular biology of the Wiskott-Aldrich syndrome.

The Wiskott-Aldrich syndrome (WAS) is an X-linked primary immunodeficiency associated with thrombocytopenia, bloody diarrhea, eczema, recurrent infections, and a high incidence of malignancies. X-linked thrombocytopenia (XLT) is a milder form with predominant platelet abnormalities. Both are caused by mutations of the cytoplasmic WAS protein (WASP). To date, mutations of WASP have been identified in over 340 families and consist of missense and nonsense mutations, deletions and insertions, and splice site mutations. There is a striking correlation between phenotype and genotype. The complex gene product of WASP has multiple functional domains that contribute to actin polymerization, cell motility, intracellular signaling, and apoptosis. Understanding the molecular basis of WAS/XLT not only explains the highly variable clinical phenotype, but also affects the medical management of this serious congenital disorder.

Regulation of oscillations in filamentous actin content in polymorphonuclear leukocytes stimulated with leukotriene B(4) and platelet-activating factor.

Stimulation of neutrophils with LTB(4) or PAF results in the production of a rapidly oscillating actin polymerization/depolymerization response. Treatment of neutrophils with inhibitors of PKC prior to stimulation with ligand resulted in a masking of the F-actin oscillations. Because myosin has been shown to be a substrate for neutrophil PKC, this protein was investigated as a potential downstream mediator of F-actin oscillations. Stimulation of neutrophils with LTB(4) resulted in myosin light chain being serine phosphorylated in a PKC-dependent manner. This phosphorylation was shown to occur in a manner that is kinetically distinct from the myosin phosphorylation induced by FMLP, a potent activator of actin polymerization that alone does not induce F-actin oscillations. Additionally, disruption of intracellular actin-myosin interactions resulted in inhibition of LTB(4)- as well as PAF-induced F-actin oscillations. These data suggest that PKC and downstream phosphorylation of myosin as well as actin-myosin interaction may play roles in mediating the production of neutrophil F-actin oscillations.

Rapid oscillations of actin polymerization/depolymerization in polymorphonuclear leukocytes stimulated by leukotriene B4 and platelet-activating factor.

We previously showed that activation of polymorphonuclear leukocytes by leukotriene B4 (LTB4) and platelet-activating factor produces a rapidly oscillating actin polymerization/depolymerization response. In this study, we show that 1) oscillations are not due to the stimulated cyclic release of autocoids that could bind to cell surface receptors and activate subsequent cycles; 2) oscillations are not related to oscillations of ligand binding; and 3) the particular kinetic pattern is a property of the receptor, not of the binding constants of the ligand. The major conclusion of these studies is that the oscillations are a property of the intrinsic signaling pathways triggered by these chemoattractants. We also questioned whether increased actin nucleation activity was induced by LTB4 and found that, although LTB4 induced a transient actin nucleation response, there was not a direct correlation between oscillations of the actin polymerization/depolymerization and the actin nucleation activity. This suggests that processes other than actin nucleation, such as release of monomeric actin from monomer sequestering proteins and regulation of depolymerization, are likely to be involved.