Optimizing early phase clinical trial washout periods: a report from the therapeutic advances in childhood leukemia and lymphoma (TACL) consortium.

PURPOSE: The National Cancer Institute (NCI) issued a 2021 memorandum adopting the American Society of Clinical Oncology (ASCO) and Friends of Cancer Research (Friends) task force recommendations to broaden clinical study eligibility criteria. They recommended that washout periods be eliminated for most prior cancer therapy and when required, to utilize evidence/rationale-based criteria. The Therapeutic Advances in Childhood Leukemia and Lymphoma (TACL) consortium responded to this guidance. PROCESS: A TACL task force reviewed the consortium's research portfolio, the relevant literature and guidance documents from ASCO-Friends, NCI, and US Food and Drug Administration (FDA) to make expert consensus and evidence-based recommendations for modernizing, broadening and codifying TACL-study washout periods while ensuring consistency with pediatric ethics and federal regulations. TACL's screening log was reviewed to estimate the impact that updated washout periods would have on patient inclusivity and recruitment. RESULTS: Over a 19-year period, 42 patients (14.6% of all screened ineligible (n = 287) patients), were identified as excluded from TACL early-phase studies exclusively due to not meeting washout criteria. An additional six (2.1%) did not meet washout and at least one other exclusion criterion. A new TACL washout guidance document was developed/adopted for use. Where washout criteria were not eliminated, rationale/evidenced-based criteria were established with citation. CONCLUSION: In an effort to reduce unnecessary exclusion from clinical trials, TACL created rationale/evidenced-based washout period standards largely following guidance from the NCI/ASCO-Friends recommendations. These new, expanded eligibility criteria are expected to increase access to TACL clinical trials while maintaining safety and scientific excellence.

Utilization of telemedicine to provide post-discharge care: A comparison of pre-pandemic vs. pandemic care.

BACKGROUND: Due to the COVID-19 pandemic, post-discharge virtual visits transitioned from a novel intervention to standard practice. Our aim was to evaluate participation in and outcomes of virtual post-discharge visits in the early-pandemic timeframe. METHODS: Pandemic cohort patients were compared to historical patients. Patient demographics, clinical information, and post-discharge 30-day hospital encounters were compared between groups. RESULTS: The historical cohort included 563 patients and the pandemic cohort had 823 patients. There was no difference in 30-day hospital encounters between patients who completed a video vs. telephone visit in the pandemic cohort (3.8% vs. 7.6%, p = 0.11). There was a lower 30-day hospital encounter rate in pandemic video and telephone visits compared to similar historical sub-groups. CONCLUSION: Expansion of virtual post-discharge visits to include all patients and telephone calls did not negatively impact rates of 30-day post-discharge hospital encounters. Offering telehealth options for post-discharge follow-up does not appear to have negative impact on healthcare utilization.

Impact of sample clarification by size exclusion on virus detection and diversity in wastewater-based epidemiology.

The use of wastewater-based epidemiology (WBE) for early detection of virus circulation and response during the SARS-CoV-2 pandemic increased interest in and use of virus concentration protocols that are quick, scalable, and efficient. One such protocol involves sample clarification by size fractionation using either low-speed centrifugation to produce a clarified supernatant or membrane filtration to produce an initial filtrate depleted of solids, eukaryotes and bacterial present in wastewater (WW), followed by concentration of virus particles by ultrafiltration of the above. While this approach has been successful in identifying viruses from WW, it assumes that majority of the viruses of interest should be present in the fraction obtained by ultrafiltration of the initial filtrate, with negligible loss of viral particles and viral diversity. We used WW samples collected in a population of ~700,000 in southwest USA between October 2019 and March 2021, targeting three non-enveloped viruses (enteroviruses [EV], canine picornaviruses [CanPV], and human adenovirus 41 [Ad41]), to evaluate whether size fractionation of WW prior to ultrafiltration leads to appreciable differences in the virus presence and diversity determined. We showed that virus presence or absence in WW samples in both portions (filter trapped solids [FTS] and filtrate) are not consistent with each other. We also found that in cases where virus was detected in both fractions, virus diversity (or types) captured either in FTS or filtrate were not consistent with each other. Hence, preferring one fraction of WW over the other can undermine the capacity of WBE to function as an early warning system and negatively impact the accurate representation of virus presence and diversity in a population.

RNA sequencing of glioblastoma tissue slice cultures reveals the effects of treatment at the transcriptional level.

One of the major challenges in cancer research is finding models that closely resemble tumors within patients. Human tissue slice cultures are a promising approach to provide a model of the patient's tumor biology ex vivo. Recently, it was shown that these slices can be successfully analyzed by whole transcriptome sequencing as well as automated histochemistry, increasing their usability as preclinical model. Glioblastoma multiforme (GBM) is a highly malignant brain tumor with poor prognosis and little is known about its genetic background and heterogeneity regarding therapy success. In this study, tissue from the tumors of 25 patients with primary GBM was processed into slice cultures and treated with standard therapy (irradiation and temozolomide). Total RNA sequencing and automated histochemistry were performed to enable analysis of treatment effects at a transcriptional and histological level. Slice cultures from long-term survivors (overall survival [OS] > 24 months) exhibited more apoptosis than cultures from patients with shorter OS. Proliferation within these slices was slightly increased in contrast to other groups, but not significantly. Among all samples, 58 protein-coding genes were upregulated and 32 downregulated in treated vs. untreated slice cultures. In general, an upregulation of DNA damage-related and cell cycle checkpoint genes as well as enrichment of genotoxicity pathways and p53-dependent signaling was found after treatment. Overall, the current study reproduces knowledge from former studies regarding the feasibility of transcriptomic analyses and automated histology in tissue slice cultures. We further demonstrate that the experimental data merge with the clinical follow-up of the patients, which improves the applicability of our model system.

Annuloaortic ectasia in a four-month-old male Newfoundland dog: long-term follow-up and immunofluorescent study.

A 4 month-old, 14.8 kg, male Newfoundland dog was presented for cardiovascular evaluation following detection of a heart murmur. Echocardiography revealed enlargement of the sinuses of Valsalva and marked, diffuse dilation of the ascending aorta (annuloaortic ectasia, AAE), with mild/equivocal subaortic stenosis (SAS). The dog was monitored over the duration of its lifetime, with serial echocardiograms performed at 5, 6, and 8 months and 1, 2, 3, 4, 8, and 10 years demonstrating persistent, diffuse dilation of the ascending aorta. The dog lived until it was 10 years old and died of metastatic carcinoma. Postmortem examination confirmed AAE and mild SAS. Hematoxylin and eosin and Weigert van Gieson stains were used to compare the ascending aorta to the descending aorta and left subclavian artery, and to compare aortic samples to those of three control dogs. Histopathologic evaluation revealed mild medial degeneration in the ascending aorta of all four dogs. Immunofluorescent microscopy was used for determining the deposition of proteins known to play a role in aortic aneurysms in humans: fibrillin-1 (FBN1), latent transforming growth factor beta binding protein 4 (LTBP4) and fibronectin. The ascending aorta of the AAE case demonstrated reduced deposition of FBN1, indicating that its loss may have contributed to aortic dilation. Diffuse, primary ascending aortic dilation is uncommonly reported in dogs; when it is, it carries a poor prognosis. This case provides an important example of marked dilation of the ascending aorta in a dog that lived with no associated adverse effects for 10 years.

Postdischarge Virtual Visits for Low-risk Surgeries: A Randomized Noninferiority Clinical Trial.

Importance: Postdischarge video-based virtual visits are a growing aspect of surgical care and have dramatically increased in the setting of the coronavirus disease 2019 (COVID-19) pandemic. Objective: To evaluate the outcomes of all-cause 30-day hospital encounter proportion among patients who have a postdischarge video-based virtual visit follow-up compared with in-person follow-up. Design, Setting, and Participants: Randomized, active, controlled noninferiority trial in an urban setting, including patients from a small community hospital and a large, tertiary care hospital. Patients who underwent minimally invasive appendectomy or cholecystectomy by a group of surgeons who cover emergency general surgery at these 2 hospitals were included. Patients undergoing elective and nonelective procedures were included. Interventions: Patients were randomized in a 2:1 fashion to video-based virtual visit or in-person visit. Main Outcomes and Measures: The primary outcome is the percentage of patients with 30-day hospital encounter, and we hypothesized that there would not be a significant increase in the 30-day hospital encounter proportion for patients who receive video-based virtual postdischarge care compared with patients who receive standard (in-person) care. Hospital encounter includes emergency department visit, observation, or inpatient admission. Results: A total of 1645 patients were screened; 289 patients were randomized to the virtual group and 143 to the in-person group. Fifty-three patients crossed over to the in-person follow-up group. The percentage of patients who had a hospital encounter was noninferior for virtual visits (12.8% vs 13.3% for in-person, Δ 0.5% with 1-sided 95% CI, -∞ to 5.2%). The amount of time patients spent with the clinician (mean of 8.4 minutes virtual vs 7.8 minutes in-person; P = .30) was not different, but the median overall postoperative visit time was 27.5 minutes shorter (95% CI, -33.5 to -24.0). Conclusions and Relevance: Postdischarge video-based virtual visits did not increase hospital encounter proportions and provided shorter overall time commitment but equal time with the surgical team member. This information will help surgeons and patients feel more confident in using video-based virtual visits. Trial Registration: ClinicalTrials.gov Identifier: NCT03258177.

Deep sequencing and automated histochemistry of human tissue slice cultures improve their usability as preclinical model for cancer research.

Cancer research requires models closely resembling the tumor in the patient. Human tissue cultures can overcome interspecies limitations of animal models or the loss of tissue architecture in in vitro models. However, analysis of tissue slices is often limited to histology. Here, we demonstrate that slices are also suitable for whole transcriptome sequencing and present a method for automated histochemistry of whole slices. Tumor and peritumoral tissue from a patient with glioblastoma was processed to slice cultures, which were treated with standard therapy including temozolomide and X-irradiation. Then, RNA sequencing and automated histochemistry were performed. RNA sequencing was successfully accomplished with a sequencing depth of 243 to 368 x 106 reads per sample. Comparing tumor and peritumoral tissue, we identified 1888 genes significantly downregulated and 2382 genes upregulated in tumor. Treatment significantly downregulated 2017 genes, whereas 1399 genes were upregulated. Pathway analysis revealed changes in the expression profile of treated glioblastoma tissue pointing towards downregulated proliferation. This was confirmed by automated analysis of whole tissue slices stained for Ki67. In conclusion, we demonstrate that RNA sequencing of tissue slices is possible and that histochemical analysis of whole tissue slices can be automated which increases the usability of this preclinical model.

Oral Methods of Urinary Alkalinization for High-dose Methotrexate Administration: Alternatives to Intravenous Sodium Bicarbonate During a Critical Drug Shortage.

A nationwide shortage of intravenous (IV) sodium bicarbonate required institutions to explore alternative options for urinary alkalinization for high-dose methotrexate (HDMTX). Children's Hospital Colorado implemented a protocol utilizing oral alkalinizing agents as alternatives to intravenous sodium bicarbonate during the shortage. The purpose of this study was to determine the safety and efficacy of oral alkalinization strategies for HDMTX administration. This retrospective study was conducted at a pediatric institution and evaluated cycles of HDMTX administered with at least one dose of oral sodium bicarbonate tablets or sodium citrate-citric acid oral solution. The time to achieve urine pH of ≥7 was 3.48 hours from the start of alkalinization. A median dose of 66.4 mEq/m/day of oral sodium bicarbonate was administered to maintain a urine pH of ≥7 until methotrexate was cleared. Gastrointestinal side effects occurred with 43% of HDMTX cycles and patients switched to IV sodium acetate in 25.5% of HDMTX cycles, primarily due to inadequate alkalinization or intolerance. During a shortage of IV sodium bicarbonate, oral alkalinization is an effective strategy for most patients to allow for administration of HDMTX.

"Are we there yet?"- factors affecting postoperative follow-up after general surgery procedures.

BACKGROUND: Surgical follow-up allows patients to discuss pathology and preventative maintenance. Multiple factors impact patients' compliance with surgical follow-up. We hypothesized that increased travel time would be associated with lack of post-discharge surgical follow-up. METHODS: Retrospective analysis identified patients undergoing laparoscopic appendectomy or laparoscopic cholecystectomy. Descriptive statistics and logistic regression assessed the relationship between patient characteristics and post-discharge follow-up. RESULTS: We identified 1830 patients from 2015-2016. 31% did not complete follow-up, were more likely to have had an appendectomy, be un- or underinsured, not married, and live outside North Carolina. Median round-trip travel time was not significantly different. After adjustment for patient factors, each additional 10 min of travel time increased the odds of not following up by 6% (p < 0.01). CONCLUSIONS: Travel time was the only modifiable factor associated with post-discharge follow-up. Novel methods of completing follow-up that minimize travel time, such as virtual visits, may increase compliance with recommended follow-up.

Impact of X-irradiation on microglia.

Irradiation is widely used to treat brain tumors, and also to create bone marrow (BM) chimeras. BM chimeras are widely used to dissect functions and origin of microglia and blood-derived mononuclear cells under homeostatic or pathological conditions. This is facilitated by the fact that microglia survive irradiation and are thus regarded radio-resistant. In this study, we tested whether microglia are indeed radio-resistant and looked for potential mechanisms that might explain this phenomenon. We analyzed the radio-resistance of microglia independently of their physiological brain environment compared to other mononuclear cells from spleen and brain after X-irradiation with 7 Gy or 30 Gy. Furthermore, we investigated long-term effects of X-irradiation on microglia using organotypic hippocampal slice cultures (OHSCs). We found a significant higher survival rate of isolated microglia 4 hr after X-irradiation with 30 Gy accompanied by a decreased proliferation rate. Investigations of apoptosis-related genes revealed no regulation of a specific antiapoptotic pathway but ataxia telangiectasia mutated (ATM), a DNA-repair-related gene, was significantly upregulated in isolated microglia 4 hr after 30 Gy. Irradiation of OHSCs with 7 and 30 Gy revealed a highly and significantly decreased cell number, morphological changes and an increase in migration velocity of microglia. Furthermore, cell loss, increased soma size and process length of microglia was also found in BM chimeras irradiated with 9.5 Gy 5 weeks after irradiation. Here, we present new evidence implying that microglia are not a homogeneous population of radio-resistant cells and report on long-term alterations of microglia that survived irradiation.

Association Between Baseline Corticothalamic-Mediated Inhibitory Control and Smoking Relapse Vulnerability.

Importance: Tobacco use disorder is associated with dysregulated neurocognitive function in the right inferior frontal gyrus (IFG)-one node in a corticothalamic inhibitory control (IC) network. Objective: To examine associations between IC neural circuitry structure and function and lapse/relapse vulnerability in 2 independent studies of adult smokers. Design, Setting, and Participants: In study 1, treatment-seeking smokers (n = 81) completed an IC task during functional magnetic resonance imaging (fMRI) before making a quit attempt and then were followed up for 10 weeks after their quit date. In study 2, a separate group of smokers (n = 26) performed the same IC task during fMRI, followed by completing a laboratory-based smoking relapse analog task. Study 1 was performed at Duke University Medical Center between 2008 and 2012; study 2 was conducted at the Medical University of South Carolina between 2013 and 2016. Main Outcomes and Measures: Associations between corticothalamic-mediated IC, gray-matter volume, and smoking lapse/relapse. Results: Of the 81 study participants in study 1 (cessation study), 45 were women (56%), with mean (SD) age, 38.4 (10.2) years. In study 1, smoking relapse was associated with less gray-matter volume (F1,74 = 28.32; familywise error P threshold = 0.03), greater IC task-related blood oxygenation level-dependent (BOLD) response in the right IFG (F1,78 = 14.87) and thalamus (F1,78 = 14.97) (P < .05), and weaker corticothalamic task-based functional connectivity (tbFC) (F1,77 = 5.87; P = .02). Of the 26 participants in study 2 (laboratory study), 15 were women (58%), with mean (SD) age, 34.9 (10.3). Similar to study 1, in study 2, greater IC-BOLD response in the right IFG (t23 = -2.49; ß = -0.47; P = .02), and weaker corticothalamic tbFC (t22 = 5.62; ß = 0.79; P < .001) were associated with smoking sooner during the smoking relapse-analog task. In both studies, corticothalamic tbFC mediated the association between IC performance and smoking outcomes. Conclusions and Relevance: In these 2 studies, baseline differences in corticothalamic circuitry function were associated with mediated IC and smoking relapse vulnerability. These findings warrant further examination of interventions for augmenting corticothalamic neurotransmission and enhancing IC during the course of tobacco use disorder treatment.

Cell Cycle Regulation and Apoptotic Responses of the Embryonic Chick Retina by Ionizing Radiation.

Ionizing radiation (IR) exerts deleterious effects on the developing brain, since proliferative neuronal progenitor cells are highly sensitive to IR-induced DNA damage. Assuming a radiation response that is comparable to mammals, the chick embryo would represent a lower vertebrate model system that allows analysis of the mechanisms underlying this sensitivity, thereby contributing to the reduction, refinement and replacement of animal experiments. Thus, this study aimed to elucidate the radiation response of the embryonic chick retina in three selected embryonic stages. Our studies reveal a lack in the radiation-induced activation of a G1/S checkpoint, but rapid abrogation of G2/M progression after IR in retinal progenitors throughout development. Unlike cell cycle control, radiation-induced apoptosis (RIA) showed strong variations between its extent, dose dependency and temporal occurrence. Whereas the general sensitivity towards RIA declined with ongoing differentiation, its dose dependency constantly increased with age. For all embryonic stages RIA occurred during comparable periods after irradiation, but in older animals its maximum shifted towards earlier post-irradiation time points. In summary, our results are in good agreement with data from the developing rodent retina, strengthening the suitability of the chick embryo for the analysis of the radiation response in the developing central nervous system.

Erwinia asparaginase achieves therapeutic activity after pegaspargase allergy: a report from the Children's Oncology Group.

AALL07P2 evaluated whether substitution of Erwinia asparaginase 25000 IU/m(2) for 6 doses given intramuscularly Monday/Wednesday/Friday (M/W/F) to children and young adults with acute lymphoblastic leukemia and clinical allergy to pegaspargase would provide a 48-hour nadir serum asparaginase activity (NSAA) ≥ 0.10 IU/mL. AALL07P2 enrolled 55 eligible/evaluable patients. NSAA ≥ 0.1 IU/mL was achieved in 38 of 41 patients (92.7%) with acceptable samples 48 hours and in 38 of 43 patients (88.4%) 72 hours after dosing during course 1. Among samples obtained during all courses, 95.8% (252 of 263) of 48-hour samples and 84.5% (125 of 148) of 72-hour samples had NSAA ≥ 0.10-IU/mL. Pharmacokinetic parameters were estimated by fitting the serum asparaginase activity-time course for all 6 doses given during course 1 to a 1-compartment open model with first order absorption. Erwinia asparaginase administered with this schedule achieved therapeutic NSAA at both 48 and 72 hours and was well tolerated with no reports of hemorrhage, thrombosis, or death, and few cases of grade 2 to 3 allergic reaction (n = 6), grade 1 to 3 hyperglycemia (n = 6), or grade 1 pancreatitis (n = 1). Following allergy to pegaspargase, Erwinia asparaginase 25000 IU/m(2) × 6 intramuscularly M/W/F can be substituted for a single dose of pegaspargase.

Tacrolimus as an alternative to cyclosporine in the maintenance phase of immunosuppressive therapy for severe aplastic anemia in children.

OBJECTIVE: Given the paucity of data on the use of agents other than cyclosporine (CsA) in the maintenance phase of immunosuppressive therapy (IST) for severe aplastic anemia (SAA) in children, we sought to describe our experience with tacrolimus in pediatric SAA, and to compare outcomes with a preceding series of patients who received CsA. METHODS: Eight patients with SAA diagnosed between 2003 and 2008 for whom no human leukocyte antigen (HLA)-matched sibling donor was identified underwent tacrolimus-based IST. These children were compared with a previously described series of 13 patients who had undergone CsA-based IST at our institution between 1990 and 2003. All patients initially received equine antithymocyte globulin (ATG) and corticosteroids. RESULTS: Complete response (CR) rate was 88% for tacrolimus and 85% for CsA. Median time to CR was approximately 7 months in both groups. Median follow-up duration was 2.4 years for tacrolimus and 8.4 years for CsA. Among responders with de novo SAA, relapse rate was 25% (n = 1) at 2 years for tacrolimus and 0% at 2 years and 23% (n = 3) at 5 years for CsA; no significant difference in relapse-free survival was detected between the two groups (P = 0.07). Paroxysmal nocturnal hemoglobinuria was seen in one patient on tacrolimus who had relapsed after CsA-based IST. Tacrolimus-based IST was well-tolerated. CONCLUSION: These data provide evidence that tacrolimus may be a suitable alternative to CsA as part of an IST regimen for SAA in children who lack an HLA-matched sibling and may have a more favorable profile of side effects than CsA.