Characterisation of the effect of FBXW7 mutations in normal and malignant colonic tissue

Colorectal cancer harbours significant intratumoral heterogeneity. Yet, while intratumoural heterogeneity may be readily observed, events which give rise to heterogeneity, as well as an understanding of the molecular effects arising from interactions between clonal populations in each cancer’s unique milieu of heterogeneity, remain elusive. I use FBXW7 as the starting point in two separate but related themes. In the first theme, colorectal cancer cell lines are used to model the effect of FBXW7 mutations on neighbouring FBXW7 wildtype cells. I demonstrate that wildtype cells may acquire phenotypic characteristics present in mutant cells owing from proteins arising from the secretome of mutant cells. In the second theme, I investigate the role of FBXW7 present in phenotypically normal tissue using gene editing in patient derived human colon organoids. Here, I demonstrate that early FBXW7 mutations result in a more restricted whole genome chromosome accessibility, abrogating the effects of later-acquired cancer driver mutations such as APC. In addition, I demonstrate that an early FBXW7 mutation can result in fetal reprogramming. In this way, FBXW7 mutations acquired in phenotypically normal tissue appear to be protective in a human colonic model of early colorectal cancer. Taken together, my experiments bear significant implications on understanding colorectal cancer intratumoural heterogeneity. These results suggest that characterising molecular interactions in intra-cancer clonal populations may lead to a deeper understanding of phenotypic characteristics relevant to that cancer. The results also suggest that early events could forestall the generation of heterogeneity and be potentially protective of cancer development. Here, I use FBXW7 as an exemplar mutation to generate implications which may be validated using other genes, and in other types of cancer

Unsupervised High-Dimensional Analysis Aligns Dendritic Cells across Tissues and Species.

Dendritic cells (DCs) are professional antigen-presenting cells that hold great therapeutic potential. Multiple DC subsets have been described, and it remains challenging to align them across tissues and species to analyze their function in the absence of macrophage contamination. Here, we provide and validate a universal toolbox for the automated identification of DCs through unsupervised analysis of conventional flow cytometry and mass cytometry data obtained from multiple mouse, macaque, and human tissues. The use of a minimal set of lineage-imprinted markers was sufficient to subdivide DCs into conventional type 1 (cDC1s), conventional type 2 (cDC2s), and plasmacytoid DCs (pDCs) across tissues and species. This way, a large number of additional markers can still be used to further characterize the heterogeneity of DCs across tissues and during inflammation. This framework represents the way forward to a universal, high-throughput, and standardized analysis of DC populations from mutant mice and human patients

Human fetal dendritic cells promote prenatal T-cell immune suppression through arginase-2.

During gestation the developing human fetus is exposed to a diverse range of potentially immune-stimulatory molecules including semi-allogeneic antigens from maternal cells, substances from ingested amniotic fluid, food antigens, and microbes. Yet the capacity of the fetal immune system, including antigen-presenting cells, to detect and respond to such stimuli remains unclear. In particular, dendritic cells, which are crucial for effective immunity and tolerance, remain poorly characterized in the developing fetus. Here we show that subsets of antigen-presenting cells can be identified in fetal tissues and are related to adult populations of antigen-presenting cells. Similar to adult dendritic cells, fetal dendritic cells migrate to lymph nodes and respond to toll-like receptor ligation; however, they differ markedly in their response to allogeneic antigens, strongly promoting regulatory T-cell induction and inhibiting T-cell tumour-necrosis factor-α production through arginase-2 activity. Our results reveal a previously unappreciated role of dendritic cells within the developing fetus and indicate that they mediate homeostatic immune-suppressive responses during gestation

Successful gelfoam angioembolisation in anastomotic pseudoaneurysm: A case report

Introduction: The use of angioembolisation in patients with lower gastrointestinal tract haemorrhage has become well established, especially in cases of diverticular bleeding, or in bleeding from arterio-venous malformations. Pseudoaneurysms are rare and the evaluation of selective gelfoam angioembolisation amongst patients with lower gastrointestinal tract bleeding secondary to this etiology has not been extensively studied. The friable nature of pseudoaneurysms may lead to a greater risk of rupture during an attempted angioembolisation procedure. Presentation of case: We describe the successful treatment and outcome of a lady who initially presented with perforation of the colon at the rectosigmoid junction, for which she underwent resection and anastomosis. A few days later, she was noted to have persistent hematochezia, which was secondary to bleeding pseudoaneurysms at the rectosigmoid branches of the inferior mesenteric artery. She successfully underwent selective angioembolisation of these pseudoaneurysms with gelfoam. She did not suffer any complications from the procedure. Discussion: Although there have been significant advances in the armamentarium associated with percutaneous interventional radiology procedures for hemostasis in gastrointestinal bleeding, the use of selective angioembolisation for bleeding pseudoaneurysms have not been readily adopted due to the friable nature of the wall of the pseudoaneurysm, and its risk for rupture. Our case report illustrates that angioembolisation in such cases is feasible, and should be a consideration especially when the risk of surgical intervention is high. Conclusion: Selective gelfoam angioembolisation should be considered in the management of patients with bleeding from the gastrointestinal tract secondary to pseudoaneurysms

Commonalities and differences in the mutational signature and somatic driver mutation landscape across solid and hollow viscus organs

10.1038/s41388-023-02802-7Oncogen

Tumour heterogeneity and evolutionary dynamics in colorectal cancer

10.1038/s41389-021-00342-xONCOGENESIS10

Generation and immunofluorescent validation of gene knockouts in adult human colonic organoids using multi-guide RNA CRISPR-Cas9

Summary: While readily achieved in cell lines, the application of CRISPR-Cas9 gene editing in human-derived organoids suffers from limited efficacy and complex protocols. Here, we describe a multi-guide RNA CRISPR-Cas9 gene-editing protocol which efficiently achieves complete gene knockout in adult human colonic organoids. This protocol also describes crucial steps including how to harvest patient tissue to maximize gene-editing efficacy and a technique to validate gene knockout following editing with immunofluorescent staining of the organoids against the target protein. : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics

Enhanced recovery program versus conventional care after colorectal surgery in the geriatric population: a systematic review and meta-analysis

10.1007/s00464-020-07673-7Surgical Endoscop

Laparoscopic Resection for Rectal Cancer: What Is the Evidence?

Laparoscopic colectomy for colon cancer is a well-established procedure supported by several well-conducted large-scale randomised controlled trials. Patients could now be conferred the benefits of the minimally invasive approach while retaining comparable oncologic outcomes to the open approach. However, the benefits of laparoscopic proctectomy for rectal cancer remained controversial. While the laparoscopic approach is more technically demanding, results from randomised controlled trials regarding long term oncologic outcomes are only beginning to be reported. The impacts of bladder and sexual functions following proctectomy are considerable and are important contributing factors to the patients’ quality of life in the long-term. These issues present a delicate dilemma to the surgeon in his choice of operative approach in tackling rectal cancer. This is compounded further by the rapid proliferation of various laparoscopic techniques including the hand assisted, robotic assisted, and single port laparoscopy. This review article aims to draw on the significant studies which have been conducted to highlight the short- and long-term outcomes and evidence for laparoscopic resection for rectal cancer