Rapamycin Regulates Autophagy and Cell Adhesion in Induced Pluripotent Stem Cells

Background Cellular reprogramming is a stressful process, which requires cells to engulf somatic features and produce and maintain stemness machineries. Autophagy is a process to degrade unwanted proteins and is required for the derivation of induced pluripotent stem cells (iPSCs). However, the role of autophagy during iPSC maintenance remains undefined. Methods Human iPSCs were investigated by microscopy, immunofluorescence, and immunoblotting to detect autophagy machinery. Cells were treated with rapamycin to activate autophagy and with bafilomycin to block autophagy during iPSC maintenance. High concentrations of rapamycin treatment unexpectedly resulted in spontaneous formation of round floating spheres of uniform size, which were analyzed for differentiation into three germ layers. Mass spectrometry was deployed to reveal altered protein expression and pathways associated with rapamycin treatment. Results We demonstrate that human iPSCs express high basal levels of autophagy, including key components of APMKα, ULK1/2, BECLIN-1, ATG13, ATG101, ATG12, ATG3, ATG5, and LC3B. Block of autophagy by bafilomycin induces iPSC death and rapamycin attenuates the bafilomycin effect. Rapamycin treatment upregulates autophagy in iPSCs in a dose/time-dependent manner. High concentration of rapamycin reduces NANOG expression and induces spontaneous formation of round and uniformly sized embryoid bodies (EBs) with accelerated differentiation into three germ layers. Mass spectrometry analysis identifies actin cytoskeleton and adherens junctions as the major targets of rapamycin in mediating iPSC detachment and differentiation. Conclusions High levels of basal autophagy activity are present during iPSC derivation and maintenance. Rapamycin alters expression of actin cytoskeleton and adherens junctions, induces uniform EB formation, and accelerates differentiation. IPSCs are sensitive to enzyme dissociation and require a lengthy differentiation time. The shape and size of EBs also play a role in the heterogeneity of end cell products. This research therefore highlights the potential of rapamycin in producing uniform EBs and in shortening iPSC differentiation duration

Adding 6 months of androgen deprivation therapy to postoperative radiotherapy for prostate cancer: a comparison of short-course versus no androgen deprivation therapy in the RADICALS-HD randomised controlled trial

Background Previous evidence indicates that adjuvant, short-course androgen deprivation therapy (ADT) improves metastasis-free survival when given with primary radiotherapy for intermediate-risk and high-risk localised prostate cancer. However, the value of ADT with postoperative radiotherapy after radical prostatectomy is unclear. Methods RADICALS-HD was an international randomised controlled trial to test the efficacy of ADT used in combination with postoperative radiotherapy for prostate cancer. Key eligibility criteria were indication for radiotherapy after radical prostatectomy for prostate cancer, prostate-specific antigen less than 5 ng/mL, absence of metastatic disease, and written consent. Participants were randomly assigned (1:1) to radiotherapy alone (no ADT) or radiotherapy with 6 months of ADT (short-course ADT), using monthly subcutaneous gonadotropin-releasing hormone analogue injections, daily oral bicalutamide monotherapy 150 mg, or monthly subcutaneous degarelix. Randomisation was done centrally through minimisation with a random element, stratified by Gleason score, positive margins, radiotherapy timing, planned radiotherapy schedule, and planned type of ADT, in a computerised system. The allocated treatment was not masked. The primary outcome measure was metastasis-free survival, defined as distant metastasis arising from prostate cancer or death from any cause. Standard survival analysis methods were used, accounting for randomisation stratification factors. The trial had 80% power with two-sided α of 5% to detect an absolute increase in 10-year metastasis-free survival from 80% to 86% (hazard ratio [HR] 0·67). Analyses followed the intention-to-treat principle. The trial is registered with the ISRCTN registry, ISRCTN40814031, and ClinicalTrials.gov, NCT00541047. Findings Between Nov 22, 2007, and June 29, 2015, 1480 patients (median age 66 years [IQR 61–69]) were randomly assigned to receive no ADT (n=737) or short-course ADT (n=743) in addition to postoperative radiotherapy at 121 centres in Canada, Denmark, Ireland, and the UK. With a median follow-up of 9·0 years (IQR 7·1–10·1), metastasis-free survival events were reported for 268 participants (142 in the no ADT group and 126 in the short-course ADT group; HR 0·886 [95% CI 0·688–1·140], p=0·35). 10-year metastasis-free survival was 79·2% (95% CI 75·4–82·5) in the no ADT group and 80·4% (76·6–83·6) in the short-course ADT group. Toxicity of grade 3 or higher was reported for 121 (17%) of 737 participants in the no ADT group and 100 (14%) of 743 in the short-course ADT group (p=0·15), with no treatment-related deaths. Interpretation Metastatic disease is uncommon following postoperative bed radiotherapy after radical prostatectomy. Adding 6 months of ADT to this radiotherapy did not improve metastasis-free survival compared with no ADT. These findings do not support the use of short-course ADT with postoperative radiotherapy in this patient population

Duration of androgen deprivation therapy with postoperative radiotherapy for prostate cancer: a comparison of long-course versus short-course androgen deprivation therapy in the RADICALS-HD randomised trial

Background Previous evidence supports androgen deprivation therapy (ADT) with primary radiotherapy as initial treatment for intermediate-risk and high-risk localised prostate cancer. However, the use and optimal duration of ADT with postoperative radiotherapy after radical prostatectomy remains uncertain. Methods RADICALS-HD was a randomised controlled trial of ADT duration within the RADICALS protocol. Here, we report on the comparison of short-course versus long-course ADT. Key eligibility criteria were indication for radiotherapy after previous radical prostatectomy for prostate cancer, prostate-specific antigen less than 5 ng/mL, absence of metastatic disease, and written consent. Participants were randomly assigned (1:1) to add 6 months of ADT (short-course ADT) or 24 months of ADT (long-course ADT) to radiotherapy, using subcutaneous gonadotrophin-releasing hormone analogue (monthly in the short-course ADT group and 3-monthly in the long-course ADT group), daily oral bicalutamide monotherapy 150 mg, or monthly subcutaneous degarelix. Randomisation was done centrally through minimisation with a random element, stratified by Gleason score, positive margins, radiotherapy timing, planned radiotherapy schedule, and planned type of ADT, in a computerised system. The allocated treatment was not masked. The primary outcome measure was metastasis-free survival, defined as metastasis arising from prostate cancer or death from any cause. The comparison had more than 80% power with two-sided α of 5% to detect an absolute increase in 10-year metastasis-free survival from 75% to 81% (hazard ratio [HR] 0·72). Standard time-to-event analyses were used. Analyses followed intention-to-treat principle. The trial is registered with the ISRCTN registry, ISRCTN40814031, and ClinicalTrials.gov , NCT00541047 . Findings Between Jan 30, 2008, and July 7, 2015, 1523 patients (median age 65 years, IQR 60–69) were randomly assigned to receive short-course ADT (n=761) or long-course ADT (n=762) in addition to postoperative radiotherapy at 138 centres in Canada, Denmark, Ireland, and the UK. With a median follow-up of 8·9 years (7·0–10·0), 313 metastasis-free survival events were reported overall (174 in the short-course ADT group and 139 in the long-course ADT group; HR 0·773 [95% CI 0·612–0·975]; p=0·029). 10-year metastasis-free survival was 71·9% (95% CI 67·6–75·7) in the short-course ADT group and 78·1% (74·2–81·5) in the long-course ADT group. Toxicity of grade 3 or higher was reported for 105 (14%) of 753 participants in the short-course ADT group and 142 (19%) of 757 participants in the long-course ADT group (p=0·025), with no treatment-related deaths. Interpretation Compared with adding 6 months of ADT, adding 24 months of ADT improved metastasis-free survival in people receiving postoperative radiotherapy. For individuals who can accept the additional duration of adverse effects, long-course ADT should be offered with postoperative radiotherapy. Funding Cancer Research UK, UK Research and Innovation (formerly Medical Research Council), and Canadian Cancer Society

An intervention using quality of life and symptom information as a clinical tool in patients with advanced cancer

The Schedule for Evaluation of Individual Quality of Life-Direct Weighting (SEIQoL-DW) recognizes the individuals’ values with no pre-judgement. The objective of this study was to determine if clinician awareness and understanding of individual patients’ subjective quality of life (SQoL), bothersome symptoms and symptom interference on SQoL, would lead to improvement over time in the SQoL of patients with advanced cancer. Methods: SEIQoL-DW and symptom bother/interference with SQoL was measured up to 4 time points on Sixty-five (n=65) patients receiving chemotherapy for advanced cancer. Measurements from the intervention group (n=33) was reported to the clinical team at each time point via an info-graph placed in the clinical notes. Measurements from the control group (n=32) was not reported to the clinical team. Results: SEIQoL-DW of all patients improved from baseline to time point 4 (p >0.05).A significant improvement in patient’s perception of health functioning was seen in the intervention group compared to the control group (51% v 19%, p=0.014). Symptom interference in SQoL reduced by 9% in the intervention group versus 37% increase in the control group. In totality, a 46% difference was observed between groups (p < 0.05). Interpretation: Translating SEIQoL-DW and symptom bother/interference into a format for use as a ‘Clinical Tool’ in routine practice improves patient reported health functioning and symptom interference on their SQoL. This patient centered approach could lead to a paradigm shift in our understanding of patient’s needs and in harmony with other clinical interventions, should be considered a valuable step towards personalized medicine

De drie gezichten van Pieter Bruegel de Oude (ca. 1525-1569)

Modern approaches to patient treatment and personalized medicine increasingly recognize the importance of the patient’s quality of life outcome. The art of assessing and using quality of life information in routine practice remains a challenge and poses a stumbling block for clinicians. This critical review of the literature identifies useful measures to minimize those challenges and guide nurses and researchers in selecting the correct tool for measuring quality of life in their clinical settingThis publication is supported by Mayo roscommon Hospice Foundation, Knock, Co. Mayo, Ireland and supported by an educational grant from the Irish Hospice Foundation, Dublin Ireland.peer-reviewe

The motivational factors and adverse events experienced by healthy volunteers donating bone marrow for research

Background: With the advancement of cell therapy research, there is an increasing need for healthy volunteers (HV) to donate small volumes (30 ml) of human bone marrow (BM). The BM procedure required to procure small volumes is invasive, although short-lived (25 seconds), is not without risk. To ensure a sustainable supply of BM for research and cell therapy, greater information of the risks and factors that motivate HV to donate small volumes of BM will help optimize the procedure and HV enrolment, ensuring donors are fully informed of the potential risks. Objective: To identify the adverse events (AE) experienced by HV during and after small volume BM procedure and understand the motivating factors that infl uence HV to donate BM for research. Method: HV (n = 55) who donated BM (30 ml) for scientifi c research and provided informed consent were administered a questionnaire to identify the type, duration and severity of AE experienced during and post-BM aspiration; and to determine the motivating factors that influenced their willingness to donate BM. Results: Pain was experienced by 89% of participants during the BM procedure with moderate grade reported by 40%. One/more of the following AE were experienced by 73% of the volunteers post-BM procedure: pain, fatigue, site reaction, nausea and transient hypotension. AE resolved within an average of three days. The reported motivational factors ranked in the following order:first, to advance research for the benefit of future patients; compensation for participation; free medical check-up; lastly, the research question was interesting. Conclusion: Young HV, motivated primarily by altruism and financial compensation, risk the occurrence of transient AE following donation of small-volume BM for research.Not peer reviewe

The motivational factors and adverse events experienced by healthy volunteers donating bone marrow for research

Background: With the advancement of cell therapy research, there is an increasing need for healthy volunteers (HV) to donate small volumes (30 ml) of human bone marrow (BM). The BM procedure required to procure small volumes is invasive, although short-lived (25 seconds), is not without risk. To ensure a sustainable supply of BM for research and cell therapy, greater information of the risks and factors that motivate HV to donate small volumes of BM will help optimize the procedure and HV enrolment, ensuring donors are fully informed of the potential risks. Objective: To identify the adverse events (AE) experienced by HV during and after small volume BM procedure and understand the motivating factors that influence HV to donate BM for research. Method: HV (n = 55) who donated BM (30 ml) for scientific research and provided informed consent were administered a questionnaire to identify the type, duration and severity of AE experienced during and post-BM aspiration; and to determine the motivating factors that influenced their willingness to donate BM. Results: Pain was experienced by 89% of participants during the BM procedure with moderate grade reported by 40%. One/more of the following AE were experienced by 73% of the volunteers post-BM procedure: pain, fatigue, site reaction, nausea and transient hypotension. AE resolved within an average of three days. The reported motivational factors ranked in the following order: first, to advance research for the benefit of future patients; compensation for participation; free medical check-up; lastly, the research question was interesting. Conclusion: Young HV, motivated primarily by altruism and financial compensation, risk the occurrence of transient AE following donation of small-volume BM for research