Saliva screening of health care workers for SARS-CoV-2 detection.

Health care workers (HCWs) are at high risk for SARS-CoV-2. In addition, pre-symptomatic or asymptomatic transmission accounts for around half of the cases. Saliva testing is an option to detect SARS-CoV-2 infection. To determine the performance of saliva samples for screening, HCWs were tested for SARS-CoV-2 by RT-PCR. Those with a positive result in saliva were tested by nasopharyngeal swabbing for viral RNA detection and blood collection to search for the presence of specific antibodies. In September-October 2020, 100 HCWs were enrolled and followed up. Six subjects (6%) tested positive in saliva. Of them, 5/6 were positive in a subsequent nasopharyngeal swab and 4/6 developed signs and symptoms compatible with COVID-19. Among the latter, 3 seroconverted while asymptomatic HCWs remained seronegative. Saliva screening was helpful for identifying SARS-CoV-2 infection in HCWs. This screening permitted rapid personnel isolation avoiding further transmission of the virus in the hospital setting.

Self-collected saliva for SARS-CoV-2 detection: A prospective study in the emergency room.

Current diagnostic standards involve severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) detection in nasopharyngeal swabs (NPS), but saliva is an attractive and noninvasive option for diagnosis. The objectives were to determine the performance of saliva in comparison with NPS for detecting SARS-CoV-2 and to compare the optimized home brew reverse-transcription polymerase chain reaction (RT-PCR) with a commercial RT-PCR. Paired NPS and saliva specimens were prospectively collected and tested by RT-PCR from patients presenting at an emergency room with signs and symptoms compatible with coronavirus disease-2019. A total of 348 samples from 174 patients were tested by RT-PCR assays. Among 174 patients with symptoms, 63 (36%) were SARS-CoV-2 positive in NPS using the optimized home-brew PCR. Of these 63 patients, 61 (98%) were also positive in saliva. An additional positive SARS-CoV-2 saliva was detected in a patient with pneumonia. Kappa Cohen's coefficient agreement between NPS and saliva was 0.96 (95% confidence interval [CI], 0.90-0.99). Median Ct values in NPS versus saliva were 18.88 (interquartile range [IQR], 15.60-23.58; range, 11.97-38.10) versus 26.10 (IQR, 22.75-30.06; range, 13.78-39.22), respectively (p < .0001). The optimized home-brew RT-PCR demonstrated higher analytical and clinical sensitivity compared with the commercial RT-PCR assay. A high sensitivity (98%) and agreement (kappa 0.96) in saliva samples compared to NPS was demonstrated when using an optimized home-brew PCR even when the viral load in saliva was lower than in NPS. This noninvasive sample is easy to collect, requires less consumable and avoids discomfort to patients. Importantly, self-collection of saliva can diminish exposure to healthcare personnel.

Does ITV vaginal procedure ensure dosimetric coverage during IMRT of post-operative gynaecological tumours without instructions concerning rectal filling?

AIM: To find out whether the internal target volume (ITV) vaginal procedure ensures dosimetric coverage during intensity-modulated radiation therapy (IMRT) of post-operative gynaecological tumours without instructions on rectal filling. BACKGROUND: The ITV vaginal procedure does not necessarily include all movements of the bladder, and does not include changes in the rectal volume. We should know if the vaginal ITV is a useful tool in maintaining CTV coverage during treatment. MATERIALS AND METHODS: A retrospective analysis of 24 patients treated between July 2012 and July 2014 with adjuvant IMRT for gynaecological cancer. All patients underwent empty and full bladder CT on simulation (CT-planning) and three weeks later (CT-control). ITV displacement was measured and the 3D vector was calculated. ITV coverage was then evaluated by comparing the volume covered by the prescription isodose on both CT's. Patients were asked to have full bladder but they did not follow recommendations for the rectum. RESULTS: The mean 3D vector was 0.64 ± 0.32 cm (0.09-1.30). The mean ITV coverage loss was 5.8 ± 5.7% (0-20.2). We found a significant positive correlation between the 3D vector and the loss of coverage (Pearson correlation, r = 0.493, 95% CI: 0.111-0.748, p = 0.0144). We did not find any significant correlation between the bladder and rectal parameters with the 3D vector and loss of dosimetric coverage. We found a trend between the maximum rectal diameter in CT-planning and 3D vector (r = 0.400, 95% CI: -0.004 to 0.692, p = 0.0529). CONCLUSION: ITV vaginal procedure contributed to ensuring a good dose coverage without instructions on rectal filling.

Apoptosis for prediction of radiotherapy late toxicity: lymphocyte subset sensitivity and potential effect of TP53 Arg72Pro polymorphism.

We tested apoptosis levels in in vitro irradiated T-lymphocytes from breast cancer (BC) patients with radiotherapy-induced late effects. Previous results reported in the literature were revised. We also examined the effect of TP53 Arg72Pro polymorphism on irradiation-induced apoptosis (IA). Twenty BC patients, ten with fibrosis and/or telangiectasias and ten matched controls with no late reactions, were selected from those receiving radiotherapy between 1993 and 2007. All patients were followed-up at least 6 years after radiotherapy. Using the combination of both CD3 and CD8 antibodies the in vitro IA was measured in CD3, CD8 and CD4 T-lymphocytes, and CD8 natural killer lymphocytes (CD8 NK) by flow cytometry. The TP53 Arg72Pro genotype was determined by sequencing. Patients with late radiotherapy toxicity showed less IA for all T-lymphocytes except for the CD8 NK. CD8 NK showed the highest spontaneous apoptosis and the lowest IA. IA in patients with toxicity appears to be lower than the control patients only in TP53 Arg/Arg patients (P = 0.077). This difference was not present in patients carrying at least one Pro allele (P = 0.8266). Our data indicate that late side effects induced by radiotherapy of BC are associated to low levels of IA. CD8 NK cells have a different response to in vitro irradiation compared to CD8 T-lymphocytes. It would be advisable to distinguish the CD8 NK lymphocytes from the pool of CD8+ lymphocytes in IA assays using CD8+ cells. Our data suggest that the 72Pro TP53 allele may influence the IA of patients with radiotherapy toxicity.

Clinical effects of re-evaluating a lung SBRT failure mode and effects analysis in a radiotherapy department.

PURPOSE: The increasing complexity of radiation treatments can hinder its clinical success. This study aimed to better understand evolving risks by re-evaluating a Failure Mode and Effects Analysis (FMEA) in lung SBRT. METHODS: An experienced multidisciplinary team conducted an FMEA and made a reassessment 3 years later. A process map was developed with potential failure modes (FMs) identified. High-risk FMs and their possible causes and corrective actions were determined. The initial FMEA analysis was compared to gain a deeper perspective. RESULTS: We identified 232 FMs. The high-risk processes were plan approval, target contouring, and patient evaluation. The corrective measures were based on stricter standardization of plan approval, pre-planning peer review, and a supporting pretreatment checklist, which substantially reduced the risk priority number in the revised FMEA. In the FMEA reassessment, we observed that the increased complexity and number of patients receiving lung SBRT conditioned a more substantial presence of human factors and communication errors as causal conditions and a potential wrong dose as a final effect. CONCLUSIONS: Conducting a lung SBRT FMEA analysis has identified high-risk conditions that have been effectively mitigated in an FMEA reanalysis. Plan approval has shown to be a weak link in the process. The increasing complexity of treatments and patient numbers have shifted causal factors toward human failure and communication errors. The potential of a wrong dose as a final effect augments in this scenario. We propose that digital and artificial intelligence options are needed to mitigate potential errors in high-complexity and high-risk RT scenarios.

Contouring variation affects estimates of normal tissue complication probability for breast fibrosis after radiotherapy.

BACKGROUND: Normal tissue complication probability (NTCP) models can be useful to estimate the risk of fibrosis after breast-conserving surgery (BCS) and radiotherapy (RT) to the breast. However, they are subject to uncertainties. We present the impact of contouring variation on the prediction of fibrosis. MATERIALS AND METHODS: 280 breast cancer patients treated BCS-RT were included. Nine Clinical Target Volume (CTV) contours were created for each patient: i) CTV_crop (reference), cropped 5 mm from the skin and ii) CTV_skin, uncropped and including the skin, iii) segmenting the 95% isodose (Iso95%) and iv) 3 different auto-contouring atlases generating uncropped and cropped contours (Atlas_skin/Atlas_crop). To illustrate the impact of contour variation on NTCP estimates, we applied two equations predicting fibrosis grade ≥ 2 at 5 years, based on Lyman-Kutcher-Burman (LKB) and Relative Seriality (RS) models, respectively, to each contour. Differences were evaluated using repeated-measures ANOVA. For completeness, the association between observed fibrosis events and NTCP estimates was also evaluated using logistic regression. RESULTS: There were minimal differences between contours when the same contouring approach was followed (cropped and uncropped). CTV_skin and Atlas_skin contours had lower NTCP estimates (-3.92%, IQR 4.00, p < 0.05) compared to CTV_crop. No significant difference was observed for Atlas_crop and Iso95% contours compared to CTV_crop. For the whole cohort, NTCP estimates varied between 5.3% and 49.5% (LKB) or 2.2% and 49.6% (RS) depending on the choice of contours. NTCP estimates for individual patients varied by up to a factor of 4. Estimates from "skin" contours showed higher agreement with observed events. CONCLUSION: Contour variations can lead to significantly different NTCP estimates for breast fibrosis, highlighting the importance of standardising breast contours before developing and/or applying NTCP models.

Cell Senescence-Related Pathways Are Enriched in Breast Cancer Patients With Late Toxicity After Radiotherapy and Low Radiation-Induced Lymphocyte Apoptosis.

Background: Radiation-induced late effects are a common cause of morbidity among cancer survivors. The biomarker with the best evidence as a predictive test of late reactions is the radiation-induced lymphocyte apoptosis (RILA) assay. We aimed to investigate the molecular basis underlying the distinctive RILA levels by using gene expression analysis in patients with and without late effects and in whom we had also first identified differences in RILA levels. Patients and Methods: Peripheral blood mononuclear cells of 10 patients with late severe skin complications and 10 patients without symptoms, selected from those receiving radiotherapy from 1993 to 2007, were mock-irradiated or irradiated with 8 Gy. The 48-h response was analyzed in parallel by RILA assay and gene expression profiling with Affymetrix microarrays. Irradiated and non-irradiated gene expression profiles were compared between both groups. Gene set enrichment analysis was performed to identify differentially expressed biological processes. Results: Although differentially expressed mRNAs did not reach a significant adjusted p-value between patients suffering and not suffering clinical toxicity, the enriched pathways indicated significant differences between the two groups, either in irradiated or non-irradiated cells. In basal conditions, the main differentially expressed pathways between the toxicity and non-toxicity groups were the transport of small molecules, interferon signaling, and transcription. After 8 Gy, the differences lay in pathways highly related to cell senescence like cell cycle/NF-κB, G-protein-coupled receptors, and interferon signaling. Conclusion: Patients at risk of developing late toxicity have a distinctive pathway signature driven by deregulation of immune and cell cycle pathways related to senescence, which in turn may underlie their low RILA phenotype.

Treatment time and circadian genotype interact to influence radiotherapy side-effects. A prospective European validation study using the REQUITE cohort.

BACKGROUND: Circadian rhythm impacts broad biological processes, including response to cancer treatment. Evidence conflicts on whether treatment time affects risk of radiotherapy side-effects, likely because of differing time analyses and target tissues. We previously showed interactive effects of time and genotypes of circadian genes on late toxicity after breast radiotherapy and aimed to validate those results in a multi-centre cohort. METHODS: Clinical and genotype data from 1690 REQUITE breast cancer patients were used with erythema (acute; n=340) and breast atrophy (two years post-radiotherapy; n=514) as primary endpoints. Local datetimes per fraction were converted into solar times as predictors. Genetic chronotype markers were included in logistic regressions to identify primary endpoint predictors. FINDINGS: Significant predictors for erythema included BMI, radiation dose and PER3 genotype (OR 1.27(95%CI 1.03-1.56); P < 0.03). Effect of treatment time effect on acute toxicity was inconclusive, with no interaction between time and genotype. For late toxicity (breast atrophy), predictors included BMI, radiation dose, surgery type, treatment time and SNPs in CLOCK (OR 0.62 (95%CI 0.4-0.9); P < 0.01), PER3 (OR 0.65 (95%CI 0.44-0.97); P < 0.04) and RASD1 (OR 0.56 (95%CI 0.35-0.89); P < 0.02). There was a statistically significant interaction between time and genotypes of circadian rhythm genes (CLOCK OR 1.13 (95%CI 1.03-1.23), P < 0.01; PER3 OR 1.1 (95%CI 1.01-1.2), P < 0.04; RASD1 OR 1.15 (95%CI 1.04-1.28), P < 0.008), with peak time for toxicity determined by genotype. INTERPRETATION: Late atrophy can be mitigated by selecting optimal treatment time according to circadian genotypes (e.g. treat PER3 rs2087947C/C genotypes in mornings; T/T in afternoons). We predict triple-homozygous patients (14%) reduce chance of atrophy from 70% to 33% by treating in mornings as opposed to mid-afternoon. Future clinical trials could stratify patients treated at optimal times compared to those scheduled normally. FUNDING: EU-FP7.

Breast cancer subtype discrimination using standardized 4-IHC and digital image analysis.

Breast cancer is a heterogeneous disease. Surrogate classification of intrinsic subtypes of invasive carcinomas by combined immunohistochemistry for estrogen receptor (ER), progesterone receptor (PR), HER2, and Ki67 (4-IHC) has increased steadily since the 2011 St Gallen symposium, due to its rapid subtyping of tumors at a reasonable cost. An important step in improving 4-IHC reproducibility and reliability will be to provide reference values from the routine use of standardized 4-IHC followed by image analysis. The aims of the current study were (1) to analyze invasive breast carcinomas using standardized 4-IHC and quantitative image analysis and (2) to compare the results obtained in the classification of biological subtypes using current Ki67 and PR threshold values proposed by different authors to sub-classifying the luminal A-like and the luminal B-like (HER2-negative) subtypes. Five hundred twenty-one tumors were analyzed by standardized immunohistochemistry, with automatic image analysis, and HER2 FISH technique. Positivity for ER was found in 82.7% and for PR in 70.1% of cases. Using the Allred scoring system, hormone receptor results showed a bimodal distribution, particularly for ER. HER2 positivity was found in 15.7% of cases, and the mean Ki67 score was 32.3%. Using the most recently proposed surrogate definitions for the classification of luminal breast cancer subtypes, the percentages of different subtypes that we found were similar to those published with genomic platforms: 40.7% luminal A-like, 32.4% luminal B-like/HER2-negative, 9.8% luminal B-like/HER2-positive, 6.0% HER2-positive, and 11.1% triple negative. Standardized 4-IHC with automatic image analysis constitutes a low-cost method for surrogate definitions of biological subtypes of breast cancer that delivers accurate results in a day.

Saliva screening of health care workers for SARS-CoV-2 detection / Análisis del personal de salud a partir de saliva para detección de SARS-CoV-2

Abstract Health care workers (HCWs) are at high risk for SARS-CoV-2. In addition, pre-symptomatic or asymptomatic transmission accounts for around half of the cases. Saliva testingis an option to detect SARS-CoV-2 infection. To determine the performance of saliva samplesfor screening, HCWs were tested for SARS-CoV-2 by RT-PCR. Those with a positive result insaliva were tested by nasopharyngeal swabbing for viral RNA detection and blood collectionto search for the presence of specific antibodies. In September---October 2020, 100 HCWs wereenrolled and followed up. Six subjects (6%) tested positive in saliva. Of them, 5/6 were positivein a subsequent nasopharyngeal swab and 4/6 developed signs and symptoms compatible withCOVID-19. Among the latter, 3 seroconverted while asymptomatic HCWs remained seronega-tive. Saliva screening was helpful for identifying SARS-CoV-2 infection in HCWs. This screeningpermitted rapid personnel isolation avoiding further transmission of the virus in the hospitalsetting.

Resumen El personal de salud (PS) tiene un alto riesgo de contraer SARS-CoV-2. La transmisión presintomática/asintomática representa alrededor de la mitad de los casos y el análisis a partir de muestras de saliva puede ser una opción para detectar la infección. Para determinar el rendimiento de estas muestras, 100 voluntarios del PS se sometieron a la detección de SARS-CoV-2 por RT-PCR en muestras de saliva en el período septiembre-octubre de 2020. De aquellos con resultado positivo en saliva, se tomaron hisopados nasofaríngeos para detectar ARN viral y muestras de suero para evaluar anticuerpos específicos. Se detectó ARN viral en la saliva de seis individuos (6%). De ellos, 5/6 fueron SARS-CoV-2 positivos en hisopado nasofaríngeo y 4/6 desarrollaron signos y síntomas compatibles con COVID-19. Entre estos últimos, tres serocon-virtieron, en tanto que los voluntarios asintomáticos permanecieron seronegativos. La muestra de saliva fue útil para identificar la infección por SARS-CoV-2 en esta cohorte del personal de salud y así proceder al rápido aislamiento de los individuos infectados, lo que evitó una mayor transmisión del virus en el ámbito hospitalario.

Hypofractionated 3D radiotherapy for inoperable T1-3 N0-1 non-small-cell lung cancer.

OBJECTIVE: This study assessed the toxicity and clinical outcomes of three-dimensional (3D) hypofractionated radiotherapy (HFRT) for medically inoperable T1-3 N0-1 non-small-cell lung cancer (NSCLC). METHODS: 34 patients with inoperable early-stage NSCLC were treated from August 2008 to April 2013. Prior to enrolment, patients were required to be evaluated by an experienced thoracic surgeon to determine the "operability". All received 57 Gy in 19 fractions followed by escalated doses of 3-Gy fractions, up to a total dose of 66 Gy using a 3D conformal technique. Toxicities were measured using the Common Terminology Criteria for Adverse Effects v. 4.0. RESULTS: The median follow-up was 33 months (7-74 months). Toxicity grades ≥3 were not observed. Local control (LC) was 80.4% at 2 years, whereas regional control (RC) was 78%. The overall survival (OS), time to progression (TTP) and time to distant metastasis (TTM) at 2 years were 60%, 59% and 80%, respectively. For patients with T1-2 N0 and a tumour size <45 mm (n = 19), rates of OS, TTP and TTM at 2 years were 71%, 75% and 94%, respectively. LC and RC at 2 years were 85% and 94%, respectively. CONCLUSION: HFRT using 3.0-Gy fractions amounting to a total dose of 66 Gy is the recommended dose. A Phase 2 trial is warranted in order to assess the safety and efficacy of this fractionation scheme. ADVANCES IN KNOWLEDGE: HFRT results in a favourable outcome in early-stage lung cancer without the usual restrictions in tumour size and/or location associated with previous treatment methods. No special equipment is required, therefore permitting its application in any centre.