Correction factors to convert microdosimetry measurements in silicon to tissue in \u3csup\u3e12\u3c/sup\u3eC ion therapy

Silicon microdosimetry is a promising technology for heavy ion therapy (HIT) quality assurance, because of its sub-mm spatial resolution and capability to determine radiation effects at a cellular level in a mixed radiation field. A drawback of silicon is not being tissue-equivalent, thus the need to convert the detector response obtained in silicon to tissue. This paper presents a method for converting silicon microdosimetric spectra to tissue for a therapeutic 12C beam, based on Monte Carlo simulations. The energy deposition spectra in a 10 μm sized silicon cylindrical sensitive volume (SV) were found to be equivalent to those measured in a tissue SV, with the same shape, but with dimensions scaled by a factor κ equal to 0.57 and 0.54 for muscle and water, respectively. A low energy correction factor was determined to account for the enhanced response in silicon at low energy depositions, produced by electrons. The concept of the mean path length (lPath) to calculate the lineal energy was introduced as an alternative to the mean chord length (l) because it was found that adopting Cauchy\u27s formula for the (l) was not appropriate for the radiation field typical of HIT as it is very directional (lPath) can be determined based on the peak of the lineal energy distribution produced by the incident carbon beam. Furthermore it was demonstrated that the thickness of the SV along the direction of the incident 12C ion beam can be adopted as (lPath). The tissue equivalence conversion method and (lPath) were adopted to determine the RBE10, calculated using a modified microdosimetric kinetic model, applied to the microdosimetric spectra resulting from the simulation study. Comparison of the RBE10 along the Bragg peak to experimental TEPC measurements at HIMAC, NIRS, showed good agreement. Such agreement demonstrates the validity of the developed tissue equivalence correction factors and of the determination of (lPath)

Identification of regions in multiple sequence alignments thermodynamically suitable for targeting by consensus oligonucleotides: application to HIV genome

BACKGROUND: Computer programs for the generation of multiple sequence alignments such as "Clustal W" allow detection of regions that are most conserved among many sequence variants. However, even for regions that are equally conserved, their potential utility as hybridization targets varies. Mismatches in sequence variants are more disruptive in some duplexes than in others. Additionally, the propensity for self-interactions amongst oligonucleotides targeting conserved regions differs and the structure of target regions themselves can also influence hybridization efficiency. There is a need to develop software that will employ thermodynamic selection criteria for finding optimal hybridization targets in related sequences. RESULTS: A new scheme and new software for optimal detection of oligonucleotide hybridization targets common to families of aligned sequences is suggested and applied to aligned sequence variants of the complete HIV-1 genome. The scheme employs sequential filtering procedures with experimentally determined thermodynamic cut off points: 1) creation of a consensus sequence of RNA or DNA from aligned sequence variants with specification of the lengths of fragments to be used as oligonucleotide targets in the analyses; 2) selection of DNA oligonucleotides that have pairing potential, greater than a defined threshold, with all variants of aligned RNA sequences; 3) elimination of DNA oligonucleotides that have self-pairing potentials for intra- and inter-molecular interactions greater than defined thresholds. This scheme has been applied to the HIV-1 genome with experimentally determined thermodynamic cut off points. Theoretically optimal RNA target regions for consensus oligonucleotides were found. They can be further used for improvement of oligo-probe based HIV detection techniques. CONCLUSIONS: A selection scheme with thermodynamic thresholds and software is presented in this study. The package can be used for any purpose where there is a need to design optimal consensus oligonucleotides capable of interacting efficiently with hybridization targets common to families of aligned RNA or DNA sequences. Our thermodynamic approach can be helpful in designing consensus oligonucleotides with consistently high affinity to target variants in evolutionary related genes or genomes

Antisense-induced ribosomal frameshifting

Programmed ribosomal frameshifting provides a mechanism to decode information located in two overlapping reading frames by diverting a proportion of translating ribosomes into a second open reading frame (ORF). The result is the production of two proteins: the product of standard translation from ORF1 and an ORF1–ORF2 fusion protein. Such programmed frameshifting is commonly utilized as a gene expression mechanism in viruses that infect eukaryotic cells and in a subset of cellular genes. RNA secondary structures, consisting of pseudoknots or stem–loops, located downstream of the shift site often act as cis-stimulators of frameshifting. Here, we demonstrate for the first time that antisense oligonucleotides can functionally mimic these RNA structures to induce +1 ribosomal frameshifting when annealed downstream of the frameshift site, UCC UGA. Antisense-induced shifting of the ribosome into the +1 reading frame is highly efficient in both rabbit reticulocyte lysate translation reactions and in cultured mammalian cells. The efficiency of antisense-induced frameshifting at this site is responsive to the sequence context 5′ of the shift site and to polyamine levels

3D silicon microdosimetry and RBE study using C-12 ion of different energies

This paper presents a new version of the 3D mesa "bridge" microdosimeter comprised of an array of 4248 silicon cells fabricated on 10 µm thick silicon-on-insulator substrate. This microdosimeter has been designed to overcome limitations existing in previous generation silicon microdosimeters and it provides well-defined sensitive volumes and high spatial resolution. The charge collection characteristics of the new 3D mesa microdosimeter were investigated using the ANSTO heavy ion microprobe, utilizing 5.5 MeV He2+ ions. Measurement of microdosimetric quantities allowed for the determination of the Relative Biological Effectiveness of 290 MeV/u and 350 MeV/u 12C heavy ion therapy beams at the Heavy Ion Medical Accelerator in Chiba (HIMAC), Japan. The microdosimetric RBE obtained showed good agreement with the tissue-equivalent proportional counter. Utilizing the high spatial resolution of the SOI microdosimeter, the LET spectra for 70 MeV 12C+6 ions, like those present at the distal edge of 290 and 350 MeV/u beams, were obtained as the ions passed through thin layers of polyethylene film. This microdosimeter can provide useful information about the lineal energy transfer (LET) spectra downstream of the protective layers used for shielding of electronic devices for single event upset prediction

Thin Silicon Microdosimeter utilizing 3D MEMS Fabrication Technology: Charge Collection Study and its application in mixed radiation fields

New 10-μm-thick silicon microdosimeters utilizing 3-D technology have been developed and investigated in this paper. The TCAD simulations were carried out to understand the electrical properties of the microdosimeters\u27 design. A charge collection study of the devices was performed using 5.5-MeV He2+ ions which were raster scanned over the surface of the detectors and the charge collection median energy maps were obtained and the detection yield was also evaluated. The devices were tested in a 290 MeV/u carbon ion beam at the Heavy Ion Medical Accelerator in Chiba (HIMAC) in Japan. Based on the microdosimetric measurements, the quality factor and dose equivalent out of field were obtained in a mixed radiation field mimicking the radiation environment for spacecraft in deep space

Real-time detection of Fe·EDTA/H2O2-induced DNA cleavage by linear dichroism

The conditions for the measurement of linear dichroism (LD) can be adjusted so as to solely reflect the length and the flexibility of DNA. The real-time detection of the EDTA·Fe2+-induced oxidative cleavage of double-stranded native and synthetic DNAs was performed using LD. The decrease in the magnitude of the LD at 260 nm, which reflects an increase in the flexibility and a decrease in the length of the DNA, can be described by the sum of two or three exponential curves in relation to the EDTA·Fe2+ concentration. The fast component was assigned to the cleavage of one of the double strands, inducing an increase in the flexibility, while the other slower component was assigned to the cleavage of the double strand, resulting in the shortening of DNA. The decrease in the magnitude of the LD of poly[d(A-T)2] was similar to that of poly[d(I-C)2], while that of poly[d(G-C)2] was found to be the slowest, indicating that the resistance of poly[d(G-C)2] against the Fenton-type reagent was the strongest. This observation suggests that the amine group in the minor groove of the double helix may play an important role in slowing the EDTA·Fe2+-induced oxidative cleavage

Computational modeling of beam-customization devices for heavy-charged-particle radiotherapy

A model for beam customization with collimators and a range-compensating filter based on the phase-space theory for beam transport is presented for dose distribution calculation in treatment planning of radiotherapy with protons and heavier ions. Independent handling of pencil beams in conventional pencil-beam algorithms causes unphysical collimator-height dependence in the middle of large fields, which is resolved by the framework comprised of generation, transport, collimation, regeneration, range-compensation, and edge-sharpening processes with a matrix of pencil beams. The model was verified to be consistent with measurement and analytic estimation at a submillimeter level in penumbra of individual collimators with a combinational-collimated carbon-ion beam. The model computation is fast, accurate, and readily applicable to pencil-beam algorithms in treatment planning with capability of combinational collimation to make best use of the beam-customization devices.Comment: 16 pages, 5 figure

Recoding of Translation in Turtle Mitochondrial Genomes: Programmed Frameshift Mutations and Evidence of a Modified Genetic Code

A +1 frameshift insertion has been documented in the mitochondrial gene nad3 in some birds and reptiles. By sequencing polyadenylated mRNA of the chicken (Gallus gallus), we have shown that the extra nucleotide is transcribed and is present in mature mRNA. Evidence from other animal mitochondrial genomes has led us to hypothesize that certain mitochondrial translation systems have the ability to tolerate frameshift insertions using programmed translational frameshifting. To investigate this, we sequenced the mitochondrial genome of the red-eared slider turtle (Trachemys scripta), where both the widespread nad3 frameshift insertion and a novel site in nad4l were found. Sequencing the region surrounding the insertion in nad3 in a number of other turtles and tortoises reveal general mitochondrial +1 programmed frameshift site features as well as the apparent redefinition of a stop codon in Parker’s snake-neck turtle (Chelodina parkeri), the first known example of this in vertebrate mitochondria

Identification of functional, endogenous programmed −1 ribosomal frameshift signals in the genome of Saccharomyces cerevisiae

In viruses, programmed −1 ribosomal frameshifting (−1 PRF) signals direct the translation of alternative proteins from a single mRNA. Given that many basic regulatory mechanisms were first discovered in viral systems, the current study endeavored to: (i) identify −1 PRF signals in genomic databases, (ii) apply the protocol to the yeast genome and (iii) test selected candidates at the bench. Computational analyses revealed the presence of 10 340 consensus −1 PRF signals in the yeast genome. Of the 6353 yeast ORFs, 1275 contain at least one strong and statistically significant −1 PRF signal. Eight out of nine selected sequences promoted efficient levels of PRF in vivo. These findings provide a robust platform for high throughput computational and laboratory studies and demonstrate that functional −1 PRF signals are widespread in the genome of Saccharomyces cerevisiae. The data generated by this study have been deposited into a publicly available database called the PRFdb. The presence of stable mRNA pseudoknot structures in these −1 PRF signals, and the observation that the predicted outcomes of nearly all of these genomic frameshift signals would direct ribosomes to premature termination codons, suggest two possible mRNA destabilization pathways through which −1 PRF signals could post-transcriptionally regulate mRNA abundance