Commissioning and implementing a Quality Assurance program for dedicated radiation oncology MRI scanners.

PURPOSE: ACR and AAPM task group's guidelines addressing commissioning for dedicated MR simulators were recently published. The goal of the current paper is to present the authors' 2-year experience regarding the commissioning and introduction of a QA program based on these guidelines and an associated automated workflow. METHODS: All mandatory commissioning tests suggested by AAPM report 284 were performed and results are reported for two MRI scanners (MAGNETOM Sola and Aera). Visual inspection, vendor clinical or service platform, third-party software, or in-house python-based code were used. Automated QA and data analysis was performed via vendor, in-house or third-party software. QATrack+ was used for QA data logging and storage. 3D geometric distortion, B0 inhomogeneity, EPI, and parallel imaging performance were evaluated. RESULTS: Contrasting with AAPM report 284 recommendations, homogeneity and RF tests were performed monthly. The QA program allowed us to detect major failures over time (shimming, gradient calibration and RF interference). Automated QA, data analysis, and logging allowed fast ACR analysis daily and monthly QA to be performed in 3 h. On the Sola, the average distortion is 1 mm for imaging radii of 250 mm or less. For radii of up to 200 mm, the maximum, average (standard deviation) distortion is 1.2  and 0.4 mm (0.3 mm). Aera values are roughly double the Sola for radii up to 200 mm. EPI geometric distortion, ghosting ratio, and long-term stability were found to be under the maximum recommended values. Parallel imaging SNR ratio was stable and close to the theoretical value (ideal g-factor). No major failures were detected during commissioning. CONCLUSION: An automated workflow and enhanced QA program allowed to automatically track machine and environmental changes over time and to detect periodic failures and errors that might otherwise have gone unnoticed. The Sola is more geometrically accurate, with a more homogenous B0 field than the Aera.

Paternal age negatively affects sperm production of the progeny.

Parental age has profound consequences for offspring's phenotype. However, whether patrilineal age affects offspring sperm production remains unknown, despite the importance of sperm production for male reproductive success in species facing post-copulatory sexual selection. Using a longitudinal dataset on ejaculate attributes of the houbara bustard, we showed that offspring sired by old fathers had different age-dependent trajectories of sperm production compared to offspring sired by young fathers. Specifically, they produced less sperm (-48%) in their first year of life, and 14% less during their lifetime. Paternal age had the strongest effect, with weak evidence for grandpaternal or great grandpaternal age effects. These results show that paternal age can affect offspring reproductive success by reducing sperm production, establishing an intergenerational link between ageing and sexual selection.

Genetic assessment of a conservation breeding program of the houbara bustard (Chlamydotis undulata undulata) in Morocco, based on pedigree and molecular analyses.

Protection and restoration of species in the wild may require conservation breeding programs under genetic management to minimize deleterious effects of genetic changes that occur in captivity, while preserving populations' genetic diversity and evolutionary resilience. Here, through interannual pedigree analyses, we first assessed the efficiency of a 21-year genetic management, including minimization of mean kinship, inbreeding avoidance, and regular addition of founders, of a conservation breeding program targeting on Houbara bustard (Chlamydotis undulata undulata) in Morocco. Secondly, we compared pedigree analyses, the classical way of assessing and managing genetic diversity in captivity, to molecular analyses based on seven microsatellites. Pedigree-based results indicated an efficient maintenance of the genetic diversity (99% of the initial genetic diversity retained) while molecular-based results indicated an increase in allelic richness and an increase in unbiased expected heterozygosity across time. The pedigree-based average inbreeding coefficient F remained low (between 0.0004 and 0.003 in 2017) while the proportion of highly inbred individuals (F > .1) decreased over time and reached 0.2% in 2017. Furthermore, pedigree-based F and molecular-based individual multilocus heterozygosity were weakly negatively correlated, (Pearson's r = -.061 when considering all genotyped individuals), suggesting that they cannot be considered as alternatives, but rather as complementary sources of information. These findings suggest that a strict genetic monitoring and management, based on both pedigree and molecular tools can help mitigate genetic changes and allow to preserve genetic diversity and evolutionary resilience in conservation breeding programs.

Post-copulatory sexual selection allows females to alleviate the fitness costs incurred when mating with senescing males.

Male senescence has detrimental effects on reproductive success and offspring fitness. When females mate with multiple males during the same reproductive bout, post-copulatory sexual selection that operates either through sperm competition or cryptic female choice might allow females to skew fertilization success towards young males and as such limit the fitness costs incurred when eggs are fertilized by senescing males. Here, we experimentally tested this hypothesis. We artificially inseminated female North African houbara bustards with sperm from dyads of males of different (young and old) or similar ages (either young or old). Then, we assessed whether siring success was biased towards young males and we measured several life-history traits of the progeny to evaluate the fitness costs due to advanced paternal age. In agreement with the prediction, we found that siring success was biased towards young males, and offspring sired by old males had impaired hatching success, growth and post-release survival (in females). Overall, our results support the hypothesis that post-copulatory sexual selection might represent an effective mechanism allowing females to avoid the fitness costs of fertilization by senescing partners.

Sperm competition accentuates selection on ejaculate attributes.

Ejaculate attributes are important factors driving the probability of fertilizing eggs. When females mate with several males, competition between sperm to fertilize eggs should accentuate selection on ejaculate attributes. We tested this hypothesis in the North African houbara bustard ( Chlamydotis undulata undulata) by comparing the strength of selection acting on two ejaculate attributes when sperm from single males or sperm from different males were used for insemination. In agreement with the prediction, we found that selection on ejaculate attributes was stronger when sperm of different males competed for egg fertilization. These findings provide the first direct comparison of the strength of selection acting on ejaculate attributes under competitive and non-competitive fertilizations, confirming that sperm competition is a major selective force driving the evolution of ejaculate characteristics.

No evidence for prezygotic postcopulatory avoidance of kin despite high inbreeding depression.

Offspring resulting from mating among close relatives can suffer from impaired fitness through the expression of recessive alleles with deleterious effects. Postcopulatory sperm selection (a prezygotic mechanism of cryptic female choice) has been suggested to be an effective way to avoid inbreeding. To investigate whether postcopulatory female choice allows the avoidance of fertilization by close kin, we performed artificial inseminations in a promiscuous bird, the houbara bustard (Chlamydotis undulata undulata). Females were inseminated with a mix of sperm from triads of males, each constituted of a male genetically unrelated to the female, a first cousin and a half-sibling. When counting the number of eggs sired by unrelated males, cousins or half-siblings, we found a significant deviation from the expected random distribution, with more eggs sired by unrelated males. However, offspring sired by cousins, and especially by half-siblings suffered from high prehatching mortality, suggesting that the observed paternity skew towards unrelated males might reflect differential offspring mortality rather than sperm selection. In agreement with this hypothesis, within-triad siring success was similar for the three parental relatedness categories, but the relationship between siring and hatching success differed across categories. In clutches with high hatching success, unrelated males had the highest success while in clutches with high failure rate, half-siblings had the highest success. Offspring sired by half-siblings also suffered from reduced growth rate during the first three months and higher posthatching mortality. Hence, despite substantial fitness costs associated with fertilization by close relatives, females do not seem to select sperm of unrelated males.

Pathological investigation of high pathogenicity avian influenza H5N8 in captive houbara bustards (Chlamydotis undulata), the United Arab Emirates 2020.

At the end of 2020, an outbreak of HPAI H5N8 was registered in captive African houbara bustards (Chlamydotis undulata) in the United Arab Emirates. In order to better understand the pathobiology of this viral infection in bustards, a comprehensive pathological characterization was performed. A total of six birds were selected for necropsy, histopathology, immunohistochemistry, RNAscope in situ hybridization and RT-qPCR and nanopore sequencing on formalin-fixed and paraffin-embedded (FFPE) tissue blocks. Gross lesions included mottled and/or hemorrhagic pancreas, spleen and liver and fibrinous deposits on air sacs and intestine. Necrotizing pancreatitis, splenitis and concurrent vasculitis, hepatitis and fibrino-heterophilic peritonitis were identified, microscopically. Viral antigens (nucleoprotein) and RNAs (matrix gene) were both detected within necro-inflammatory foci, parenchymal cells, stromal cells and endothelial cells of affected organs, including the myenteric plexus. Molecular analysis of FFPE blocks successfully detected HPAI H5N8, further confirming its involvement in the lesions observed. In conclusion, HPAI H5N8 in African houbara bustards results in hyperacute/acute forms exhibiting marked pantropism, endotheliotropism and neurotropism. In addition, our findings support the use of FFPE tissues for molecular studies of poorly characterized pathogens in exotic and endangered species, when availability of samples is limited.

Quantitative genetics of sexual display, ejaculate quality and size in a lekking species.

The investment into extravagant sexual display and competitive sperm are two essential components of pre- and post-copulatory sexual selection. Even though the selective forces acting on sexual display and sperm characteristics have been extensively studied in recent years, the genetic architecture underlying the expression of these traits has been rarely explored. Here, we estimated the genetic variances and covariances of traits linked with ejaculate size and quality, and sexual display in the houbara bustard (Chlamydotis undulata undulata, Jacquin 1784). Using a very large pedigree-based data set, we show that sexual signalling and ejaculate size (but not ejaculate quality) are heritable and genetically positively correlated. The matrix of genetic covariances also provided support for some across-sex correlations: male and female gamete numbers are positively correlated, and more surprisingly, male display and female gamete numbers are also positively correlated. These results can have important implications for the understanding of the evolution of sperm traits and sexual display in animals.

Ranking parameters driving siring success during sperm competition in the North African houbara bustard.

Sperm competition is a powerful force driving the evolution of ejaculate and sperm traits. However, the outcome of sperm competition depends on many traits that extend beyond ejaculate quality. Here, we study male North African houbara bustards (Chlamydotis undulata undulata) competing for egg fertilization, after artificial insemination, with the aim to rank the importance of 14 parameters as drivers of siring success. Using a machine learning approach, we show that traits independent of male quality (i.e., insemination order, delay between insemination and egg laying) are the most important predictors of siring success. Traits describing intrinsic male quality (i.e., number of sperm in the ejaculate, mass motility index) are also positively associated with siring success, but their contribution to explaining the outcome of sperm competition is much lower than for insemination order. Overall, this analysis shows that males mating at the last position in the mating sequence have the best chance to win the competition for egg fertilization. This raises the question of the importance of female behavior as determinant of mating order.

Genome-wide CRISPR Screen Reveals RAB10 as a Synthetic Lethal Gene in Colorectal and Pancreatic Cancers Carrying SMAD4 Loss.

The TGFß signaling mediator SMAD4 is frequently mutated or deleted in colorectal and pancreatic cancers. SMAD4 acts as a tumor suppressor and its loss is associated with poorer patient outcomes. The purpose of this study was to find synthetic lethal interactions with SMAD4 deficiency to find novel therapeutic strategies for the treatment of patients with SMAD4-deficient colorectal or pancreatic cancers. Using pooled lentiviral single-guide RNA libraries, we conducted genome-wide loss-of-function screens in Cas9-expressing colorectal and pancreatic cancer cells harboring altered or wild-type SMAD4. The small GTPase protein RAB10 was identified and validated as a susceptibility gene in SMAD4-altered colorectal and pancreatic cancer cells. Rescue assays showed that RAB10 reintroduction reversed the antiproliferative effects of RAB10 knockout in SMAD4-negative cell lines. Further investigation is necessary to shed light on the mechanism by which RAB10 inhibition decreases cell proliferation of SMAD4-negative cells. Significance: This study identified and validated RAB10 as new synthetic lethal gene with SMAD4. This was achieved by conducting a whole-genome CRISPR screens in different colorectal and pancreatic cell lines. A future RAB10 inhibitors could correspond to a new therapeutic solution for patients with cancer with SMAD4 deletion.

A Monte Carlo based formalism to identify potential locations at high risk of tumor recurrence with a numerical model for glioblastoma multiforme.

PURPOSE: The strategy currently used to treat glioblastoma multiforme (GBM) patients, which mostly relies on population-based failure patterns, does not consider the important variability in such patterns reported in the literature. As part of the multidisciplinary efforts being made to develop personalized therapeutic approaches, numerical models of tumor growth and treatment are increasingly being used by different groups around the world. In this study, a new formalism relying on the proliferation-invasion model is developed to identify potential locations of GBM recurrences. The authors assess the sensitivity of the location of potential tumor recurrences to the input parameter values predicted for a given patient by varying those values using a Monte-Carlo based approach. Our approach is designed to be prospective in the sense that it relies on patient-specific imaging data that can be gathered in one single preradiotherapy imaging session. METHODS: The authors modeled the infiltration paths of glial cells using patient-specific diffusion tensor imaging (DTI) data. Nine GBM patients with preradiotherapy DTI data are considered in this study. The possible locations of tumor recurrences are determined by randomly selecting many ensembles of values for each of the growth and radiobiological parameters in the GBM growth model. A novel concept, the occurrence probability (OP), is introduced to assess the sensitivity of potential tumor recurrence locations to the input parameter values. For a given patient, the OP map is derived from a superposition of all potential tumor recurrence locations obtained with all sets of parameter values. RESULTS: For eight out of nine of patients, the authors have identified a statistically significant region where the OP is above 50%. For two patients, these high risk regions are found to be located at a distance greater than 3.9 cm from the border of the gross tumor volume highlighting the inaccuracy of current margins for some patients. The exact location and size of these volumes with OP > 50 % are, however, sensitive to the number N of ensembles of parameter values for N ≲ 400. On the other hand, the authors have identified for each patient a threshold OP, the OP(T), which defines a volume that converges more rapidly with increasing N. The OP(T) for each patient varies between 20% and 40%. The volume defined by OP > OP(T) may be an adequate candidate to define a personalized margin for radiotherapy treatment planning of GBM patients. CONCLUSIONS: A new Monte-Carlo based formalism was described and used to assess the variability of sites of potential recurrence predicted by the proliferation-invasion model to input parameter values. The authors have shown that high risk areas could be consistently identified with a limited number of sets (N ≲ 400) of randomly chosen parameter values. A major strength of this formalism is its potential prospective nature. Although a validation of the accuracy of the model-predicted tumor recurrence location still remains to be done, our method is potentially applicable to orient patient-specific definition of margins.

Reference dosimetry using radiochromic film.

The objectives of this study are to identify and quantify factors that influence radiochromic film dose response and to determine whether such films are suitable for reference dosimetry. The influence of several parameters that may introduce systematic dose errors when performing reference dose measurements were investigated. The effect of the film storage temperature was determined by comparing the performance of three lots of GAFCHROMIC EBT2 films stored at either 4ºC or room temperature. The effect of high (> 80%) or low (< 20%) relative humidity was also determined. Doses measured in optimal conditions with EBT and EBT2 films were then compared with an A12 ionization chamber measurement. Intensity-modulated radiation therapy quality controls using EBT2 films were also performed in reference dose. The results obtained using reference dose measurements were compared with those obtained using relative dose measurements. Storing the film at 4ºC improves the stability of the film over time, but does not eliminate the noncatalytic film development, seen as a rise in optical density over time in the absence of radiation. Relative humidity variations ranging from 80% to 20% have a strong impact on the optical density and could introduce dose errors of up to 15% if the humidity were not controlled during the film storage period. During the scanning procedure, the film temperature influences the optical density that is measured. When controlling for these three parameters, the dose differences between EBT or EBT2 and the A12 chamber are found to be within ± 4% (2σ level) over a dose range of 20-350 cGy. Our results also demonstrate the limitation of the Anisotropic Analytical Algorithm for dose calculation of highly modulated treatment plans.

Assisted Reproduction Techniques to Improve Reproduction in a Non-Model Species: The Case of the Arabian Bustard (Ardeotis arabs) Conservation Breeding Program.

Artificial reproductive technologies are highly valuable for ex situ conservation. While Arabian bustard populations are declining and extinct in some parts of the range, the International Fund for Houbara Conservation in the United Arab Emirates implemented a conservation breeding program. Since 2012, a total of 1253 eggs were laid through natural reproduction, 1090 were incubated and 379 of these were fertile (fertility rate of 34.8%), leading to the production of 251 chicks. To improve fertility and acquire crucial knowledge for other endangered large birds, artificial reproduction was implemented in 2018 using fresh, refrigerated, and frozen sperm. A total of 720 ejaculates were collected from 12 birds. We analysed these samples for concentration, volume, motility score (0 to 5), viability (eosin/nigrosine), length, and morphology. The first age at collection was 35.7 ± 18.8 months, mean volume was 89.2 ± 65.3 µL, mean concentration was 928 ± 731 sptz/mL and mean motility score was 2.61 ± 0.95. Morphology analyses revealed a bimodal distribution of sperm length. Five hundred and thirty-five ejaculates were cryopreserved and the initial motility score was 3.4 ± 0.7 and 2.0 ± 0.6 after thawing, while the percentage of normal and intact membrane sperm cells decreased from 88.8 ± 7.5% to 52.9 ± 1%. Sixty-five artificial inseminations were performed, leading to a global fertility rate of 84.3%-more precisely, 85.2% and 83.3%, respectively, for fresh and cryopreserved semen. All methods successfully produced fertile eggs, indicating that artificial insemination is an efficient tool for the conservation and genetic management of the species.

Advantages of TRUS-based delineation for high-dose-rate prostate brachytherapy planning.

PURPOSE: To evaluate the variability of prostate contours delineated on computed tomography (CT) and transrectal ultrasound (TRUS). MATERIAL AND METHODS: A TRUS-based high-dose-rate (HDR) brachytherapy procedure was introduced in 2016 in our center. The first thirty patients were additionally imaged with CT immediately after the treatment. In 2018, four different radiation oncologists (ROs: 1, 2, 3, 4) contoured the prostate on both modalities. A volume comparison was performed between CT and TRUS imaging. Using prostate gold fiducial makers, a rigid registration between CT and TRUS was done in 20 of the 30 patients studied. Jaccard index (JI) was computed to evaluate the inter-observer volume delineation agreement. RESULTS: The ratio of TRUS/CT volumes was 0.82 (95% CI: 0.79-0.87%). The mean JI was 87% for CT and 92% for TRUS, when comparing all four ROs; CT and TRUS JIs were significantly different (p < 0.001). The mean JI for the prostate on CT was significantly more consistent (p < 0.001) when comparing RO1, 2, and 3 together (RO1-2, RO1-3, and RO2-3; mean = 89%) than when comparing RO4 (newest to clinical practice) to others (RO1-4, RO2-4, and RO3-4; mean = 85%). For TRUS planning, the mean JI was not significantly different (p > 0.05) when comparing all ROs. CONCLUSIONS: The inter-observer and intra-observer variability were statistically significantly smaller on TRUS compared to CT-based planning, despite varying ROs clinical experiences. The superior soft tissue contrast offered by TRUS obviates the effect of the ROs experience on prostate contour volumes and enables more reproducible prostate delineation.

Sexually extravagant males age more rapidly.

Evolutionary theories of ageing posit that increased reproductive investment occurs at the expense of physiological declines in later life. Males typically invest heavily in costly sexual ornaments and behaviour, but evidence that the expression of these traits can cause senescence is lacking. Long-lived houbara bustards (Chlamydotis undulata) engage in extravagant sexual displays to attract mates and here we show that males investing most in these displays experience a rapid senescent deterioration of spermatogenic function at a younger age. This effect is sufficiently large that the expected links between male 'showiness' and fertility reverse in later life, despite 'showy' males continuing to display at near maximal levels. We show that our results cannot be explained by the selective disappearance of competitive phenotypes and that they are instead consistent with an early vs. late life trade-off in male reproductive competence, highlighting the potential significance of sexual selection in explaining rates of ageing.

Extraction of depth-dependent perturbation factors for silicon diodes using a plastic scintillation detector.

PURPOSE: This work presents the experimental extraction of the perturbation factor in megavoltage electron beams for three models of silicon diodes (IBA Dosimetry, EFD and SFD, and the PTW 60012 unshielded) using a plastic scintillation detector (PSD). METHODS: The authors used a single scanning PSD mounted on a high-precision scanning tank to measure depth-dose curves in 6-, 12-, and 18-MeV clinical electron beams. They also measured depth-dose curves using the IBA Dosimetry, EFD and SFD, and the PTW 60012 unshielded diodes. The authors used the depth-dose curves measured with the PSD as a perturbation-free reference to extract the perturbation factors of the diodes. RESULTS: The authors found that the perturbation factors for the diodes increased substantially with depth, especially for low-energy electron beams. The experimental results show the same trend as published Monte Carlo simulation results for the EFD diode; however, the perturbations measured experimentally were greater. They found that using an effective point of measurement (EPOM) placed slightly away from the source reduced the variation of perturbation factors with depth and that the optimal EPOM appears to be energy dependent. CONCLUSIONS: The manufacturer recommended EPOM appears to be incorrect at low electron energy (6 MeV). In addition, the perturbation factors for diodes may be greater than predicted by Monte Carlo simulations.

Validation of an electron Monte Carlo dose calculation algorithm in the presence of heterogeneities using EGSnrc and radiochromic film measurements.

The purpose of this study is to validate Eclipse's electron Monte Carlo algorithm (eMC) in heterogeneous phantoms using radiochromic films and EGSnrc as a reference Monte Carlo algorithm. Four heterogeneous phantoms are used in this study. Radiochromic films are inserted in these phantoms, including in heterogeneous media, and the measured relative dose distributions are compared to eMC calculations. Phantoms A, B, and C contain 1D heterogeneities, built with layers of lung- (phantom A) and bone- (phantoms B and C) equivalent materials sandwiched in Plastic Water. Phantom D is a thorax anthropomorphic phantom with 2D lung heterogeneities. Electron beams of 6, 9, 12 and 18 MeV from a Varian Clinac 2100 are delivered to these phantoms with a 10 × 10 cm2 applicator. Monte Carlo simulations with an independent algorithm (EGSnrc) are also used as a reference tool for two purposes: (1) as a second validation of the eMC dose calculations, and (2) to calculate the stopping power ratio between radiochromic films and bone medium, when dose is measured inside the heterogeneity. Percent depth dose (PDD) film measurements and eMC calculations agree within 2% or 3 mm for phantom A, and within 3% or 3 mm for phantoms B and C for almost all beam energies. One exception is observed with phantom B and the 6 MeV, where measured PDDs and those calculated with eMC differ by up to 4 mm. Gamma analysis of the measured and calculated 2D dose distributions in phantom D agree with criteria of 3%, 3mm for 9, 12, and 18 MeV beams, and criteria of 5%, 3 mm for the 6 MeV beam. Dose calculations in heterogeneous media with eMC agree within 3% or 3 mm with radiochromic film measurements. Six (6) MeV beams are not modeled as accurately as other beam energies. The eMC algorithm is suitable for clinical dose calculations involving lung and bone.

Enforced monoandry over generations induces a reduction of female investment into reproduction in a promiscuous bird.

While uncovering the costs and benefits of polyandry has attracted considerable attention, assessing the net effect of sexual selection on population fitness requires the experimental manipulation of female mating over generations, which is usually only achievable in laboratory populations of arthropods. However, knowing if sexual selection improves or impairs the expression of life-history traits is key for the management of captive populations of endangered species, which are mostly long-lived birds and mammals. It might therefore be questionable to extrapolate the results gathered on laboratory populations of insects to infer the net effect of sexual selection on populations of endangered species. Here, we used a longitudinal dataset that has been collected on a long-lived bird, the houbara bustard, kept in a conservation breeding program, to investigate the effect of enforced monoandry on female investment into reproduction. In captivity, female houbara bustards are artificially inseminated with sperm collected from a single male (enforced monoandry), or sequentially inseminated with semen of different males (polyandry), allowing postcopulatory sexual selection to operate. We identified female lines that were produced either by monoandrous or polyandrous inseminations over three generations, and we compared reproductive investment of females from the two mating system groups. We found that females in the polyandrous lines had higher investment into reproduction as they laid more eggs per season and produced heavier hatchlings. Higher reproductive investment into reproduction in the polyandrous lines did not result from inherited differences from females initially included in the two mating system groups. These results show that removal of sexual selection can alter reproductive investment after only few generations, potentially hindering population fitness and the success of conservation breeding programs.

PSA outcomes and late toxicity of single-fraction HDR brachytherapy and LDR brachytherapy as monotherapy in localized prostate cancer: A phase 2 randomized pilot study.

PURPOSE: To evaluate the PSA outcomes and the late patient's reported health related quality of life (HRQOL) and toxicity after single-fraction High-Dose-Rate brachytherapy (HDRB) and Low-Dose-Rate brachytherapy (LDRB) for prostate cancer. METHODS: Men with low and favorable intermediate-risk prostate cancer across 3 centres were randomized between monotherapy brachytherapy with either Iodine-125 LDRB or 19 Gy single-fraction HDRB. Biochemical outcomes were evaluated using the Phoenix definition, PSA nadir and absolute PSA value <0.4 ng/mL. Toxicities and HRQOL were recorded at 24 and 36 months. RESULTS: A total of 31 patients were randomized, 15 in the LDRB arm and 16 patients in the HDRB arm. After a median follow-up of 45(36-53) months, 3 patients in the HDRB arm experienced biochemical failure (p = 0.092). Nineteen Gy single-fraction HDRB was associated with significantly higher PSA nadir compared to LDRB (1.02 ± 0.66vs 0.25 ± 0.39, p < 0.0001). Moreover, a significantly larger proportion of patients in the LDRB group had a PSA <0.4 ng/mL (13/15 vs 2/16, p < 0.0001). For late Genito-Urinary, Gastro-Intestinal, and sexual toxicities at 24 and 36 months, no significant differences were found between the 2 arms. As for HRQOL, the IPSS and EPIC-26 urinary irritative score were significantly better for patients treated with HDRB over the first 36 months post-treatment (p = 0.001 and p = 0.01, respectively), reflecting superior HRQOL. CONCLUSION: HDRB resulted in superior HRQOL in the irritative urinary domain compared to LDRB. PSA nadir was significantly lower in the LDRB group and a higher proportion of patients in the LDRB group reached PSA <0.4 ng/mL.

Sensitivity of Oncogenic KRAS-Expressing Cells to CDK9 Inhibition.

Oncogenic forms of KRAS proteins are known to be drivers of pancreatic, colorectal, and lung cancers. The goal of this study is to identify chemical leads that inhibit oncogenic KRAS signaling. We first developed an isogenic panel of mouse embryonic fibroblast (MEF) cell lines that carry wild-type RAS, oncogenic KRAS, and oncogenic BRAF. We validated these cell lines by screening against a tool compound library of 1402 annotated inhibitors in an adenosine triphosphate (ATP)-based cell viability assay. Subsequently, this MEF panel was used to conduct a high-throughput phenotypic screen in a cell viability assay with a proprietary compound library. All 126 compounds that exhibited a selective activity against mutant KRAS were selected and prioritized based on their activities in secondary assays. Finally, five chemical clusters were chosen. They had specific activity against SW620 and LS513 over Colo320 colorectal cancer cell lines. In addition, they had no effects on BRAFV600E, MEK1, extracellular signal-regulated kinase 2 (ERK2), phosphoinositide 3-kinase alpha (PI3Kα), AKT1, or mammalian target of rapamycin (mTOR) as tested in in vitro enzymatic activity assays. Biophysical assays demonstrated that these compounds did not bind directly to KRAS. We further identified the mechanism of action and showed that three of them have CDK9 inhibitory activity. In conclusion, we have developed and validated an isogenic MEF panel that was used successfully to identify RAS oncogenic or wild-type allele-specific vulnerabilities. Furthermore, we identified sensitivity of oncogenic KRAS-expressing cells to CDK9 inhibitors, which warrants future studies of treating KRAS-driven cancers with CDK9 inhibitors.