Enhanced Diaphragm Muscle Function upon Satellite Cell Transplantation in Dystrophic Mice.

The diaphragm muscle is essential for breathing, and its dysfunctions can be fatal. Many disorders affect the diaphragm, including muscular dystrophies. Despite the clinical relevance of targeting the diaphragm, there have been few studies evaluating diaphragm function following a given experimental treatment, with most of these involving anti-inflammatory drugs or gene therapy. Cell-based therapeutic approaches have shown success promoting muscle regeneration in several mouse models of muscular dystrophy, but these have focused mainly on limb muscles. Here we show that transplantation of as few as 5000 satellite cells directly into the diaphragm results in consistent and robust myofiber engraftment in dystrophin- and fukutin-related protein-mutant dystrophic mice. Transplanted cells also seed the stem cell reservoir, as shown by the presence of donor-derived satellite cells. Force measurements showed enhanced diaphragm strength in engrafted muscles. These findings demonstrate the feasibility of cell transplantation to target the diseased diaphragm and improve its contractility.

The adult environment promotes the transcriptional maturation of human iPSC-derived muscle grafts.

Pluripotent stem cell (PSC)-based cell therapy is an attractive option for the treatment of multiple human disorders, including muscular dystrophies. While in vitro differentiating PSCs can generate large numbers of human lineage-specific tissue, multiple studies evidenced that these cell populations mostly display embryonic/fetal features. We previously demonstrated that transplantation of PSC-derived myogenic progenitors provides long-term engraftment and functional improvement in several dystrophic mouse models, but it remained unknown whether donor-derived myofibers mature to match adult tissue. Here, we transplanted iPAX7 myogenic progenitors into muscles of non-dystrophic and dystrophic mice and compared the transcriptional landscape of human grafts with respective in vitro-differentiated iPAX7 myotubes as well as human skeletal muscle biospecimens. Pairing bulk RNA sequencing with computational deconvolution of human reads, we were able to pinpoint key myogenic changes that occur during the in vitro-to-in vivo transition, confirm developmental maturity, and consequently evaluate their applicability for cell-based therapies.

The HH-GLI2-CKS1B network regulates the proliferation-to-maturation transition of cardiomyocytes.

Cardiomyocyte (CM) proliferation and maturation are highly linked processes, however, the extent to which these processes are controlled by a single signaling axis is unclear. Here, we show the previously undescribed role of Hedgehog (HH)-GLI2-CKS1B cascade in regulation of the toggle between CM proliferation and maturation. Here we show downregulation of GLI-signaling in adult human CM, adult murine CM, and in late-stage hiPSC-CM leading to their maturation. In early-stage hiPSC-CM, inhibition of HH- or GLI-proteins enhanced CM maturation with increased maturation indices, increased calcium handling, and transcriptome. Mechanistically, we identified CKS1B, as a new effector of GLI2 in CMs. GLI2 binds the CKS1B promoter to regulate its expression. CKS1B overexpression in late-stage hiPSC-CMs led to increased proliferation with loss of maturation in CMs. Next, analysis of datasets of patients with heart disease showed a significant enrichment of GLI2-signaling in patients with ischemic heart failure (HF) or dilated-cardiomyopathy (DCM) disease, indicating operational GLI2-signaling in the stressed heart. Thus, the Hh-GLI2-CKS1B axis regulates the proliferation-maturation transition and provides targets to enhance cardiac tissue engineering and regenerative therapies.

Toxicity profile and Patient-Reported outcomes following salvage Stereotactic Ablative Radiation Therapy to the prostate Bed: The POPART multicentric prospective study.

Background: While SBRT to the prostate has become a valuable option as a radical treatment, limited data support its use in the postoperative setting. Here, we report the updated results of the multicentric Post-Prostatectomy Ablative Radiation Therapy (POPART) trial, investigating possible predictors of toxicities and patient-reported outcomes. Methods: Patients with PSA levels between 0.1-2.0 ng/mL after radical prostatectomy received Linac-based SBRT to the prostate bed in five fractions every other day for a total dose of 32.5 Gy (EQD21.5 = 74.3 Gy). Late toxicity was assessed using CTCAE v.5 scale, while EPIC-CP, ICIQ-SF, IIEF 5 questionnaires and PSA levels measured quality of life and biochemical control. Pre- and post-treatment scores were compared using a paired t-test, with MID established at > 0.5 pooled SD from the baseline. A logistic regression analysis was performed to evaluate potential associations between specific patient/tumor/treatment factors and outcome deterioration. Results: From April 2021 to April 2023 a total of 50 pts were enrolled and treated. Median follow-up was 12.2 (3-27) months. No late ≥ G2 GI or GU toxicity was registered. Late G1 urinary and rectal toxicities occurred in 46 % and 4 % of patients, respectively. Among 47 patients completing all EPIC-CP domains, four (9 %) showed worsened QoL, and eleven (26 %) developed erectile dysfunction correlating with PTV D2% (P = 0.032). At Multivariate analysis bladder wall D10cc independently correlated with late G1 GU toxicity (P = 0.034). Median post-treatment PSA nadir was 0.04 ng/mL (0.00 - 0.84). At the last follow-up, six patients presented with biochemical failure, including two nodal relapses. Conclusions: Our findings show that post-prostatectomy SBRT did not result in increased toxicity nor a significant decline in QoL measures, thus showing that it can be safely extended to the postoperative setting. Long-term follow-up and randomized comparisons with different RT schedules are needed to validate this approach.

Quality of Life Longitudinal Evaluation in Prostate Cancer Patients from Radiotherapy Start to 5 Years after IMRT-IGRT.

PURPOSE: The purpose of this study is to study the evolution of quality of life (QoL) in the first 5 years following Intensity-modulated radiation therapy (IMRT) for prostate cancer (PCa) and to determine possible associations with clinical/treatment data. MATERIAL AND METHODS: Patients were enrolled in a prospective multicentre observational trial in 2010-2014 and treated with conventional (74-80 Gy, 1.8-2 Gy/fr) or moderately hypofractionated IMRT (65-75.2 Gy, 2.2-2.7 Gy/fr). QoL was evaluated by means of EORTC QLQ-C30 at baseline, at radiation therapy (RT) end, and every 6 months up to 5 years after IMRT end. Fourteen QoL dimensions were investigated separately. The longitudinal evaluation of QoL was analysed by means of Analysis of variances (ANOVA) for multiple measures. RESULTS: A total of 391 patients with complete sets of questionnaires across 5 years were available. The longitudinal analysis showed a trend toward the significant worsening of QoL at RT end for global health, physical and role functioning, fatigue, appetite loss, diarrhoea, and pain. QoL worsening was recovered within 6 months from RT end, with the only exception being physical functioning. Based on ANOVA, the most impaired time point was RT end. QoL dimension analysis at this time indicated that acute Grade ≥ 2 gastrointestinal (GI) toxicity significantly impacted global health, physical and role functioning, fatigue, appetite loss, diarrhoea, and pain. Acute Grade ≥ 2 genitourinary (GU) toxicity resulted in lower role functioning and higher pain. Prophylactic lymph-nodal irradiation (WPRT) resulted in significantly lower QoL for global health, fatigue, appetite loss, and diarrhoea; lower pain with the use of neoadjuvant/concomitant hormonal therapy; and lower fatigue with the use of an anti-androgen. CONCLUSIONS: In this prospective, longitudinal, observational study, high radiation IMRT doses delivered for PCa led to a temporary worsening of QoL, which tended to be completely resolved at six months. Such transient worsening was mostly associated with acute GI/GU toxicity, WPRT, and higher prescription doses.

Pelvic bone marrow dose-volume predictors of late lymphopenia following pelvic lymph node radiation therapy for prostate cancer.

BACKGROUND AND PURPOSE: Given the substantial lack of knowledge, we aimed to assess clinical/dosimetry predictors of late hematological toxicity on patients undergoing pelvic-nodes irradiation (PNI) for prostate cancer (PCa) within a prospective multi-institute study. MATERIALS AND METHODS: Clinical/dosimetry/blood test data were prospectively collected including lymphocytes count (ALC) at baseline, mid/end-PNI, 3/6 months and every 6 months up to 5-year after PNI. DVHs of the Body, ileum (BMILEUM), lumbosacral spine (BMLS), lower pelvis (BMPELVIS), and whole pelvis (BMTOT) were extracted. Current analysis focused on 2-year CTCAEv4.03 Grade ≥ 2 (G2+) lymphopenia (ALC < 800/µL). DVH parameters that better discriminate patients with/without toxicity were first identified. After data pre-processing to limit overfitting, a multi-variable logistic regression model combining DVH and clinical information was identified and internally validated by bootstrap. RESULTS: Complete data of 499 patients were available: 46 patients (9.2 %) experienced late G2+ lymphopenia. DVH parameters of BMLS/BMPELVIS/BMTOT and Body were associated to increased G2+ lymphopenia. The variables retained in the resulting model were ALC at baseline [HR = 0.997, 95 %CI 0.996-0.998, p < 0.0001], smoke (yes/no) [HR = 2.9, 95 %CI 1.25-6.76, p = 0.013] and BMLS-V ≥ 24 Gy (cc) [HR = 1.006, 95 %CI 1.002-1.011, p = 0.003]. When acute G3+ lymphopenia (yes/no) was considered, it was retained in the model [HR = 4.517, 95 %CI 1.954-10.441, p = 0.0004]. Performances of the models were relatively high (AUC = 0.87/0.88) and confirmed by validation. CONCLUSIONS: Two-year lymphopenia after PNI for PCa is largely modulated by baseline ALC, with an independent role of acute G3+ lymphopenia. BMLS-V24 was the best dosimetry predictor: constraints for BMTOT (V10Gy < 1520 cc, V20Gy < 1250 cc, V30Gy < 850 cc), and BMLS (V24y < 307 cc) were suggested to potentially reduce the risk.

Worsening of 2-year patient-reported intestinal functionality after radiotherapy for prostate cancer including pelvic node irradiation.

BACKGROUND AND PURPOSE: To quantify patient-reported 2-year intestinal toxicity (IT) from pelvic nodal irradiation (PNI) for prostate cancer. The association between baseline/acute symptoms and 2-year worsening was investigated. MATERIALS AND METHODS: Patient-reported IT was prospectively assessed through the Inflammatory Bowel Disease Questionnaire (IBDQ), filled in at baseline, radiotherapy mid-point and end, at 3 and 6 months and every 6 months until 5 years. Two-year deterioration of IBDQ scores relative to the Bowel Domain was investigated for 400 patients with no severe baseline symptoms and with questionnaires available at baseline, 2 years, RT mid-point and/or end and at least three follow-ups between 3 and 18 months. The significance of the 2-year differences from baseline was tested. The association between baseline values and ΔAcute (the worst decline between baseline and RT mid-point/end) was investigated. RESULTS: In the IBDQ lower scores indicate worse symptoms. A significant (p < 0.0001) 2-year mean worsening, mostly in the range of -0.2/-0.4 points on a 1-7 scale, emerged excepting one question (IBDQ29, "nausea/feeling sick"). This decline was independent of treatment intent while baseline values were associated with 2-year absolute scores. The ΔAcute largely modulated 2-year worsening: patients with ΔAcute greater than the first quartile (Q1) and ΔAcute less or equal than Q1 showed no/minimal and highly significant (p < 0.0001) deterioration, respectively. Rectal incontinence, urgency, frequency and abdominal pain showed the largest mean changes (-0.5/-1): risk of severe worsening (deemed to be of clinical significance if ≤ 2) was 3-5 fold higher in the ΔAcute ≤ Q1 vs ΔAcute > Q1 group (p < 0.0001). CONCLUSION: A modest but significant deterioration of two-year patient-reported intestinal symptoms from PNI compared to baseline was found. Patients experiencing more severe acute symptoms are at higher risk of symptom persistence at 2 years, with a much larger prevalence of clinically significant symptoms.

The Prognostic Role of Preoperative PSMA PET/CT in cN0M0 pN+ Prostate Cancer: A Multicenter Study.

CONTEXT: Despite negative preoperative conventional imaging, up to 10% of patients with prostate cancer (PCa) harbor lymph-node involvement (LNI) at radical prostatectomy (RP). The advent of more accurate imaging modalities such as PET/CT improved the detection of LNI. However, their clinical impact and prognostic value are still unclear. We aimed to investigate the prognostic value of preoperative PET/CT in patients node positive (pN+) at RP. EVIDENCE SYNTHESIS: We retrospectively identified cN0M0 patients at conventional imaging (CT and/or MRI, and bone scan) who had pN+ PCa at RP at 17 referral centers. Patients with cN+ at PSMA/Choline PET/CT but cN0M0 at conventional imaging were also included. Systemic progression/recurrence was the primary outcome; Cox proportional hazards models were used for multivariate analysis. EVIDENCE ACQUISITION: We included 1163 pN+ men out of whom 95 and 100 had preoperative PSMA and/or Choline PET/CT, respectively. ISUP grade ≥4 was detected in 66.6%. Overall, 42% of patients had postoperative PSA persistence (≥0.1 ng/mL). Postoperative management included initial observation (34%), ADT (22.7%) and adjuvant RT+/-ADT (42.8%). Median follow-up was 42 months. Patients with cN+ on PSMA PET/CT had an increased risk of systemic progression (52.9% vs. 13.6% cN0 PSMA PET/CT vs. 21.5% cN0 at conventional imaging; P < .01). This held true at multivariable analysis: (HR 6.184, 95% CI: 3.386-11-295; P < .001) whilst no significant results were highlighted for Choline PET/CT. No significant associations for both PET types were found for local progression, BCR, and overall mortality (all P > .05). Observation as an initial management strategy instead of adjuvant treatments was related with an increased risk of metastases (HR 1.808; 95% CI: 1.069-3.058; P < .05). CONCLUSIONS: PSMA PET/CT cN+ patients with negative conventional imaging have an increased risk of systemic progression after RP compared to their counterparts with cN0M0 disease both at conventional and/or molecular imaging.

Stereotactic prostate radiotherapy with or without androgen deprivation therapy, study protocol for a phase III, multi-institutional randomized-controlled trial.

Objective: The therapeutic landscape for localized prostate cancer (PC) is evolving. Stereotactic radiotherapy (SRT) has been reported to be at least not inferior to standard radiotherapy, but the effect of androgen deprivation therapy (ADT) in this setting is still unknown and its use is left to clinical judgment. There is therefore the need to clarify the role of ADT in association with SRT, which is the aim of the present study. Methods: We present a study protocol for a randomized, multi-institutional, Phase III clinical trial, designed to study SRT in unfavorable intermediate and a subclass of high-risk localized PC. Patients (pts) will be randomized 1:1 to SRT + ADT or SRT alone. SRT will consists in 36.25 Gy in 5 fractions, ADT will be a single administration of Triptorelin 22.5 mg concurrent to SRT. Primary end point will be biochemical disease-free survival. Secondary end points will be disease-free survival, freedom from local recurrence, freedom from regional recurrence, freedom from distant metastasis and overall survival (OS); quality of life QoL and patient reported outcomes will be an exploratory end point and will be scored with EPIC-26, EORTC PR 25, IPSS, IIEF questionnaires in SRT + ADT and SRT alone arms. Moreover, clinician reported acute and late toxicity, assessed with CTCAE v. 5.0 scales will be safety end points. Results: Sample size is estimated of 310 pts. For acute toxicity and quality of life results are awaited after 6 months since last patient in, whereas, for efficacy end points and late toxicity mature results will be available 3-5 years after last patient in. Conclusion: Evidence is insufficient to guide decision making concerning ADT administration in the new scenario of prostate ultra-hypofractionation. Hence, the need to investigate the ADT role in SRT specific setting. Advances in knowledge: The stereotactic prostate radiotherapy with or without ADT trial (SPA Trial) has been designed to establish a new standard of care for SRT in localized unfavorable intermediate and a subclass of localized high risk PC.