Abstract Background Colorectal cancer (CRC) frequently exhibits hypoxic regions due to poor vascularization, leading to the stabilization of hypoxia-inducible factor 1 alpha (HIF-1α). Moreover, mutations in the tumour suppressor p53 occur in approximately half of all CRCs. While the individual roles of both transcription factors in tumour cell survival are well characterized, their interaction and its influence on the metabolic adaptation of CRC cells under hypoxic stress remain unclear. Methods Using HCT116 CRC cells with targeted deletions of TP53 and HIF1A, we examined the effects of p53 loss on HIF-1 signalling and the respective consequences for metabolic adaptation as well as the survival of CRC cells under moderate (1% O₂) and severe (0.1% O₂) hypoxia. Results Severe hypoxia stabilized p53 protein levels despite the transcriptional repression of TP53, possibly through posttranslational mechanisms and dependent on nutrient availability. In contrast to the assumption that p53 is transcriptionally inactive under hypoxia, we observed stable expression of p53 target genes (P21, BAX) under severe hypoxia, indicating functional transactivation. Loss of p53 impaired the early induction of HIF-1 target genes (VEGF, PHD2), although HIF-1α protein levels and DNA binding were unaffected, suggesting a coactivator role for p53. Furthermore, compared with wild-type cells, p53-deficient cells presented delayed but exaggerated expression of glycolytic genes, including Glucose Uptake Transporter 1 (GLUT1), Phosphofructokinase Liver-Type (PFKL) and Lactate Dehydrogenase A (LDHA), under hypoxia, with no impairment of glycolytic function or cell viability. Remarkably, even HIF1A knockout cells preserved glycolysis, whereas glycolytic genes were significantly downregulated, indicating HIF-1-independent metabolic compensation. Conclusion Our findings position p53 as a temporal gatekeeper and key regulator of hypoxic adaptation in CRC cells, coordinating early gene induction and metabolic responses. The ability of CRC cells to maintain glycolysis despite the loss of p53, respectively, HIF-1α underscores the existence of compensatory HIF-independent pathways. Targeting these alternative circuits may represent a promising strategy in hypoxic, p53-deficient CRC.
Tumour infiltrating lymphocytes (TILs) are a key component of the tumour microenvironment. To establish a clinically relevant TILs cut-off for patients with oesophago-gastric (OG) cancer, it is essential to know whether TILs density varies by patient and/or disease characteristics. TILs were quantified as TILs/mm2 (TILs density) by a deep-learning algorithm applied to digitised Haematoxylin/Eosin (H E)-stained biopsies and resection specimens from 4628 patients from nine phase III trials. 4533 patients with TILs density and matched clinicopathological data were included in the final analyses. Associations between TILs density, disease stage, geographical region (UK versus Asia), sex, age, and treatment were analysed. Median TILs density was higher in pre-treatment biopsies from patients with early-stage versus late-stage disease (962 vs 479 TILs/mm2, p < 0.001). Within the same geographical region and disease stage, TILs density was similar across different chemotherapy regimens. In UK-led trials of early-stage disease, post-chemotherapy resections showed higher TILs density than chemotherapy-naïve resections (618 vs 571 TILs/mm2, p = 0.003). TILs density was higher in Asian tumours compared to UK tumours (1419 vs 571 TILs/mm2, p < 0.001). No significant associations were observed with age or sex. This is the largest study to date evaluating TILs density in OG cancer. TILs density varied with stage and geographical region but not by age or sex. These findings may explain enhanced response to immunotherapy observed in published studies of patients with early-stage disease and highlight the need to account for baseline TILs heterogeneity when interpreting TILs as a possible biomarker in future studies.
Vascular allografts represent a potential alternative to autologous grafts when suitable vessels are unavailable. However, current preservation methods, particularly cryopreservation, lead to substantial endothelial damage and poor clinical outcomes. This study evaluated the long-term efficacy of an N-acetylhistidine-buffered, potassium-chloride-enriched and amino-acid-fortified solution augmented with iron chelators, as an advanced preservation solution (NTK-Chel), compared to histidine-tryptophan-ketoglutarate (HTK) and normal saline for vascular graft storage. Abdominal aortae from male Wistar rats (n = 75) were stored for 2 weeks at 4 °C in NTK-Chel (n = 24), HTK (n = 26), or normal saline (n = 25), then transplanted orthotopically. Grafts were harvested at 8, 16, and 26 weeks post-transplantation for vascular function testing and histological analysis. Upon explantation, smooth muscle contractility was severely impaired in all preservation solutions at 8, 16, and 26 weeks post-transplantation. At 8 weeks, only NTK-Chel-preserved grafts demonstrated detectable contractile responses to KCl, though these did not meet criteria for comprehensive vasomotor testing. By 16 and 26 weeks, all grafts had lost measurable contractility. The NTK-Chel group was associated with significantly reduced intimal hyperplasia compared to HTK and normal saline, particularly at 8 and 26 weeks. However, progressive smooth muscle cell loss occurred in all groups, with NTK-Chel showing only delayed rather than prevented degeneration. While NTK-Chel resulted in significantly less intimal hyperplasia compared to conventional solutions, prolonged cold storage combined with transplantation-associated remodeling ultimately compromises long-term vasomotor function regardless of preservation strategy.
To use gastric content analysis to estimate the postmortem interval, the last meal consumed, and its gastric emptying time must be known. However, previous studies have often investigated single foods, liquid meals, or applied heterogeneous protocols, limiting forensic applicability. Thus, this study aims to investigate gastric emptying times of commonly consumed solid mixed meals with different macronutrient compositions using non-invasive MRI-scans to measure gastric volumes. Fifteen healthy participants consumed high-fat, high-carbohydrate, and high-protein meals on separate days, with postprandial MRI-scans every two hours for up to eight hours (total of 232 scans). Gastric half-emptying-times (t₅₀) were calculated via linear interpolation. The high-carbohydrate meal (t₅₀ 132 ± 33 min) emptied significantly faster than the high-fat (t₅₀ 188 ± 41 min, p <0.001) and the high-protein (t₅₀ 158 ± 37 min, p=0.010) meals. Women showed on average greater t₅₀-values, significant only for the high-protein meal (p=0.003). This study provides reference values on nutrient-dependent gastric emptying times supporting forensic postmortem interval estimation as a supplementary method. Not applicable.
Adjuvant anti-PD-1 therapy improves outcomes in resected stage IIB/IIC melanoma at high risk of recurrence; however, no randomized head-to-head trial has directly compared pembrolizumab with nivolumab. The objective was to compare the efficacy of pembrolizumab and nivolumab as adjuvant therapies in resected stage IIB/IIC melanoma using an indirect treatment comparison (ITC). An anchored Bucher ITC was performed linking the results of KEYNOTE-716 and the CheckMate-76K trials. Primary analyses used the most recent follow-up hazard ratios (HRs), with secondary analyses based on the original trial publications. The ITC showed no statistically significant differences between pembrolizumab and nivolumab for recurrence-free survival (RFS; HR 0.97, 95
