Search

To evaluate a knowledge-based (KB) planning model for RapidPlan, generated using a five-field intensity-modulated radiotherapy (IMRT) class solution beam strategy and rigorous dosimetric constraints for accelerated partial breast irradiation (APBI).The RapidPlan model was configured using 64 APBI treatment plans and validated for 120 APBI patients who were not included in the training dataset. KB plan dosimetry was compared to clinical plan dosimetry, the clinical planning constraints, and the constraints used in phase III APBI trials. Dosimetric differences between clinical and KB plans were evaluated using paired two-tailed Wilcoxon signed-rank tests.KB planning was able to produce IMRT-based APBI plans in a single optimization without manual intervention that are comparable or better than the conventionally optimized, clinical plans. Comparing KB plans to clinical plans, differences in PTV, heart, contralateral breast, and ipsilateral lung dose-volume metrics were not clinically significant. The ipsilateral breast volume receiving at least 50% of the prescription dose was statistically and clinically significantly lower in the KB plans.KB planning for IMRT-based APBI provides equivalent or better dosimetry compared to conventional inverse planning. This model may be reliably applied in clinical practice and could be used to transfer planning expertise to ensure consistency in APBI plan quality.

Retinitis pigmentosa type 45 (RP45) is an autosomal-recessively inherited blinding disease caused by mutations in the cyclic nucleotide-gated channel subunit beta 1 (CNGB1) gene. In this study, we developed and tested a novel gene supplementation therapy suitable for clinical translation. To this end, we designed a recombinant adeno-associated virus (rAAV) vector carrying a genome that features a novel human rhodopsin promoter (hRHO194) driving rod-specific expression of full-length human CNGB1 (rAAV5.hCNGB1). rAAV5.hCNGB1 was evaluated for efficacy in the Cngb1 knockout (Cngb1(−/−)) mouse model of RP45. In particular, increasing doses of rAAV5.hCNGB1 were delivered through single subretinal injection in 4-week-old Cngb1(−/−) mice and the treatment effect was assessed over a follow-up period of 9 months at the level of (1) retinal morphology, (2) retinal function, (3) vision-guided behavior, and (4) transgene expression. We found that subretinal treatment with rAAV5.hCNGB1 resulted in efficient expression of the human CNGB1 protein in mouse rods and was able to normalize the expression of the endogenous mouse CNGA1 subunit, which together with CNGB1 forms the native heterotetrameric cyclic guanosine monophosphate-gated cation channel in rod photoreceptors. The treatment led to a dose-dependent recovery of rod photoreceptor-driven function and preservation of retinal morphology in Cngb1(−/−) mice. In summary, these results demonstrate the efficacy of hCNGB1 gene supplementation therapy in the Cngb1(−/−) mouse model of RP45 and support the translation of this approach toward future clinical application.

Adeno-associated viral (AAV) vectors represent an ideal vehicle for human gene transfer. One advantage to the AAV vector system is the availability of multiple naturally occurring serotypes that provide selective tropisms for various target cells. Strategies to enhance the properties of the natural AAV isolates have been developed and can be divided into two approaches, rational design or directed evolution. The rational design approach utilizes knowledge of AAV capsids to make targeted changes to the capsid to alter transduction efficiency or specificity, while the directed evolution approach does not require a priori knowledge of capsid structure and includes random mutagenesis, capsid shuffling, or random peptide insertion. In this study, we describe the generation of novel variants for both AAV2 and AAV5 using a rational design approach and knowledge of AAV receptor binding, surface charge, and AAV capsid protein posttranslational modifications. The novel AAV2 and AAV5 variants demonstrate improved transduction properties in both the mouse retina and cornea. The translational fidelity of the novel AAV2 variant was confirmed in the context of the nonhuman primate (NHP) retina, whereas a NHP tissue explant model was established to allow the rapid assessment of translational fidelity between species for the AAV5 variants. The capsid-modified AAV2 and AAV5 variants described in this study have novel attributes that will add to the efficacy and specificity of their potential use in gene therapy for a range of human ocular diseases.

Objective. To demonstrate an updated approach for deriving planning target volume (PTV) margins for a patient population treated with volumetric image-guided radiotherapy. Approach. The approach uses a semi-automated workflow within commercial radiotherapy applications that combines dose accumulation with the bidirectional local distance (BLD) metric. The patient cohort is divided into derivation and validation datasets. For each patient in the derivation dataset, a treatment plan is generated with a 0 mm PTV margin (the idealized treatment scenario without the influence of the standard margin). Deformable image registration enabled dose accumulation of these zero-margin plans. PTV margins are derived by using the BLD to calculate the geometric extent of underdosed regions of the clinical target volume (CTV). The PTV margin is validated by ensuring the specified CTV coverage criterion is met when the margin is applied to the validation dataset. Main results. The methodology was applied to two cohorts: 40 oropharyngeal cancer patients and 50 early-stage breast cancer patients. Ten patients from each cohort were used for validation. PTV margins derived for the oropharyngeal and early-stage breast cancer patient cohorts were 3 and 5 mm, respectively, and ensure that 95% of the prescription dose is delivered to 98% of the CTV for 90% of patients. Dose accumulation showed that the CTV coverage criterion was achieved for at least 90% of patients when the margins were applied. Significance. This methodology can be used to derive appropriate PTV margins for realistic treatment scenarios and any disease site, which will improve our understanding of patient outcomes.

The generation of clinical good manufacturing practices (GMP)-grade adeno-associated virus (AAV) vectors requires purification strategies that support the generation of vectors of high purity, and that exhibit a good safety and efficacy profile. To date, most reported purification schemas are serotype dependent, requiring method development for each AAV gene therapy product. Here, we describe a platform purification process that is compatible with the purification of multiple AAV serotypes. The method generates vector preparations of high purity that are enriched for capsids with full vector genomes, and that minimizes the fractional content of empty capsids. The two-column purification method, a combination of affinity and ion exchange chromatographies, is compatible with a range of AAV serotypes generated by either the transient triple transfection method or the more scalable producer cell line platform. In summary, the adaptable purification method described can be used for the production of a variety of high-quality AAV vectors suitable for preclinical testing in animal models of diseases. Keywords: AAV purification, epitope swapping, analytical ultracentrifugation

Purpose The purpose of the study was to establish a quantitative method for implant quality evaluation in permanent seed implant brachytherapy for credentialing. Delivery-based credentialing will promote consistency in brachytherapy seed delivery and improve patient outcomes. Methods A workflow for delivery-based credentialing was outlined and applied to permanent breast seed implant brachytherapy. Delivery simulations were performed on implantable anthropomorphic breast phantoms. Two institutions experienced in permanent seed implant brachytherapy demonstrated the peer credentialing process. Each delivery was evaluated for seed placement accuracy as the measure of implant quality, both for implant accuracy and across five simulations to assess implant variation. Initial credentialing criteria are set based on two factors; the mean seed placement accuracy (implant accuracy) and the mean standard deviation (seed variation) with the threshold for each set with the addition of two standard deviations. Results Across two institutions, seed placement accuracy (±standard deviation) was calculated for all five delivery simulations to yield 6.1 (±2.6) mm. To set credentialing criteria, the implant accuracy (6.1 mm) plus two standard deviations (2.0 mm) and the seed variation (2.6 mm) plus two standard deviations (0.8) mm yield a threshold of 8.1 ± 3.4 mm. It is expected that 95% of experienced institutions would perform the phantom simulation within this threshold. Conclusion Brachytherapy programs should validate delivery accuracy by formal credentialing, which is standard in external beam programs. This quantitative implant evaluation should be combined with current credentialing standards for permanent seed brachytherapy to form a comprehensive validation of institutional brachytherapy program quality.

Several recent studies have demonstrated the potential for quantitative imaging features to classify non-small cell lung cancer (NSCLC) patients as high or low risk. However applying the results from one institution to another has been difficult because of the variations in imaging techniques and feature measurement. Our study was designed to determine the effect of some of these sources of uncertainty on image features extracted from computed tomography (CT) images of non-small cell lung cancer (NSCLC) tumors. CT images from 20 NSCLC patients were obtained for investigating the impact of four sources of uncertainty: Two region of interest (ROI) selection conditions (breathing phase and single-slice vs. whole volume) and two imaging protocol parameters (peak tube voltage and current). Texture values did not vary substantially with the choice of breathing phase; however, almost half (12 out of 28) of the measured textures did change significantly when measured from the average images compared to the end-of-exhale phase. Of the 28 features, 8 showed a significant variation when measured from the largest cross sectional slice compared to the entire tumor, but 14 were correlated to the entire tumor value. While simulating a decrease in tube voltage had a negligible impact on texture features, simulating a decrease in mA resulted in significant changes for 13 of the 23 texture values. Our results suggest that substantial variation exists when textures are measured under different conditions, and thus the development of a texture analysis standard would be beneficial for comparing features between patients and institutions.