The Ubiquitin-Proteasome Pathway (UPP) in colonic adenocarcinoma

The ubiquitin-proteasome pathway (UPP) facilitates intracellular protein degradation, and as such has become the target of scientific work as many of the substrates of the proteasome are mediators of cell cycle pathways which become dysregulated during tumourigenesis. Modulation of this pathway through proteasomal inhibition represents a novel approach to the targeted treatment of malignant disease. We investigated the expression of proteins involved in the UPP, using fresh frozen colonic tumour tissue and patient - matched normal adjacent tissue from a cohort of patients with Dukes\u27 C colonic adenocarcinoma. (n=18) Subsequently, expression levels of ubiquitin and proteasome subunits were assessed in a large tissue microarray (TMA) of patients with Dukes\u27 B and C colon cancer. (n=240) Immunoblotting analysis demonstrated increased expression of ubiquitinylated proteins in tumour specimens compared with normal colonic specimens. Furthermore, parallel up-regulation in expression of regulatory subunits of the proteasome, Rpt4 and 20s core were observed. Furthermore, we investigated treatment of colon cancer HCTl16 cells with theproteasome inhibitor, bonezomib and demonstrated caspase dependent apoptosis maximal at 24 hours. Moreover, flow cytometry analysis of a panel of five colon cancer cell lines treated with bortezomib, revealed that these cells were highly sensitive to proteasome inhibition.(p\u3c0.05) In addition, we demonstrated that siRNA silencing of Rpt4 induces apoptosis and prevents proliferation of colon cancer cells.(p\u3c0.05) Analysis of our TMA, yielded interesting results. Most notably, Rpt4 was determined to be an independent prognostic marker for reduced disease free and overall survival in Dukes B patients. (Disease free survival :*p=0.0217, hazard ratio 0.6) Overall survival: *p=0.0467, hazard ratio 0.65) Our data supports an overstressed UPP with increased levels of ubiquitinylatedproteins despite elevated proteasomal levels. Moreover, proteasomal inhibition induces caspase dependent apoptosis in vitro. Herein, proteasomal inhibition represents a novel chemotherapeutic entry point for the treatment of colon cancer

Targeting the 19S proteasomal subunit, Rpt4, for the treatment of colon cancer.

Deregulation of the ubiquitin-proteasome pathway has been frequently observed in a number of malignancies. Using quantitative Western blotting of normal and matched tumour tissue, we here identified a significant increase in the 19S proteasome subunit Rpt4 in response to chemoradiation in locally advanced rectal cancer patients with unfavourable outcome. We therefore explored the potential of Rpt4 reduction as a therapeutic strategy in colorectal cancer (CRC). Utilizing siRNA to down regulate Rpt4 expression, we show that silencing of Rpt4 reduced proteasomal activity and induced endoplasmic reticulum stress. Gene silencing of Rpt4 also inhibited cell proliferation, reduced clonogenic survival and induced apoptosis in HCT-116 colon cancer cells. We next developed a cell penetrating peptide-based nanoparticle delivery system to achieve in vivo gene silencing of Rpt4. Administration of Rpt4 siRNA nanoparticles reduced tumour growth and improved survival in a HCT-116 colon cancer xenograft tumour model in vivo. Collectively, our data suggest that inhibition of Rpt4 represents a novel strategy for the treatment of CRC

Calnexin, an ER-induced protein, is a prognostic marker and potential therapeutic target in colorectal cancer.

BACKGROUND: Colorectal cancer (CRC) is a leading cause of cancer mortality in the Western world and commonly treated with genotoxic chemotherapy. Stress in the endoplasmic reticulum (ER) was implicated to contribute to chemotherapeutic resistance. Hence, ER stress related protein may be of prognostic or therapeutic significance. METHODS: The expression levels of ER stress proteins calnexin, calreticulin, GRP78 and GRP94 were determined in n = 23 Stage II and III colon cancer fresh frozen tumour and matched normal tissue samples. Data were validated in a cohort of n = 11 rectal cancer patients treated with radiochemotherapy in the neoadjuvant setting. The calnexin gene was silenced using siRNA in HCT116 cells. RESULTS: There were no increased levels of ER stress proteins in tumour compared to matched normal tissue samples in Stage II or III CRC. However, increased calnexin protein levels were predictive of poor clinical outcome in the patient cohort. Data were validated in the rectal cancer cohort treated in the neoadjuvant setting. Calnexin gene-silencing significantly reduced cell survival and increased cancer cell susceptibility to 5FU chemotherapy. CONCLUSION: Increased tumour protein levels of calnexin may be of prognostic significance in CRC, and calnexin may represent a potential target for future therapies

Erratum to: Calnexin, an ER stress-induced protein, is a prognostic marker and potential therapeutic target in colorectal cancer

Erratum to: Calnexin, an ER stress-induced protein, is a prognostic marker and potential therapeutic target in colorectal cancer

Global disparities in surgeons’ workloads, academic engagement and rest periods: the on-calL shIft fOr geNEral SurgeonS (LIONESS) study

: The workload of general surgeons is multifaceted, encompassing not only surgical procedures but also a myriad of other responsibilities. From April to May 2023, we conducted a CHERRIES-compliant internet-based survey analyzing clinical practice, academic engagement, and post-on-call rest. The questionnaire featured six sections with 35 questions. Statistical analysis used Chi-square tests, ANOVA, and logistic regression (SPSS® v. 28). The survey received a total of 1.046 responses (65.4%). Over 78.0% of responders came from Europe, 65.1% came from a general surgery unit; 92.8% of European and 87.5% of North American respondents were involved in research, compared to 71.7% in Africa. Europe led in publishing research studies (6.6 ± 8.6 yearly). Teaching involvement was high in North America (100%) and Africa (91.7%). Surgeons reported an average of 6.7 ± 4.9 on-call shifts per month, with European and North American surgeons experiencing 6.5 ± 4.9 and 7.8 ± 4.1 on-calls monthly, respectively. African surgeons had the highest on-call frequency (8.7 ± 6.1). Post-on-call, only 35.1% of respondents received a day off. Europeans were most likely (40%) to have a day off, while African surgeons were least likely (6.7%). On the adjusted multivariable analysis HDI (Human Development Index) (aOR 1.993) hospital capacity > 400 beds (aOR 2.423), working in a specialty surgery unit (aOR 2.087), and making the on-call in-house (aOR 5.446), significantly predicted the likelihood of having a day off after an on-call shift. Our study revealed critical insights into the disparities in workload, access to research, and professional opportunities for surgeons across different continents, underscored by the HDI

Clinical application of a systems model of apoptosis execution for the prediction of colorectal cancer therapy responses and personalisation of therapy

Key to the clinical management of colorectal cancer is identifying tools which aid in assessing patient prognosis and determining more effective and personalised treatment strategies. We evaluated whether an experimental systems biology strategy which analyses the susceptibility of cancer cells to undergo caspase activation can be exploited to predict patient responses to 5-fluorouracil-based chemotherapy and to case-specifically identify potential alternative targeted treatments to reactivate apoptosis.status: publishe

Collating evidence to support the validation of a simulated laparotomy incision and closure-training model

Background: It is essential to evaluate the functionality of surgical simulation models, in order to determine whether they perform as intended. In this study, we assessed the use of a simulated laparotomy incision and closure-training model by collating validity evidence to determine its utility as well as pre and post-test interval data. Method: This was a quantitative study design, informed by Messick's unified validity framework. In total, 93 participants (surgical trainees ​= ​80, experts ​= ​13) participated in this study. Evaluation of content validity and the models' relationships with other variables was conducted, along with a pre and post-test confidence assessment. Results: The model was deemed realistic and useful as a teaching tool, providing strong content validity evidence. In assessment of relationships with other variables, the expert group out-performed the novice group conclusively. Pre and post-test evaluation reported a statistically significant increase in confidence levels. Conclusion: We present strong validity evidence of a novel laparotomy incision and closure simulation-training model.</p