Introduction

The detection of microsatellite instability (MSI) in tumor tissue is of high relevance for (1) identifying patients eligible for immune checkpoint blockade and (2) Lynch syndrome carriers. Gold standard for MSI analysis are DNA-based methods that are encompassing PCR-based amplification of short repetitive sequence stretches (microsatellites) and assessing length variations in the respective amplicons.
However, diagnostic complications can arise due to potential artifacts known as stutter bands, which are erroneous insertions or deletions occurring in PCR amplification that make it difficult to detect the true length of the microsatellite fragment. Correctly distinguishing these artifacts from true genetic insertion/deletion (indel) mutations in microsatellites in vivo is crucial for accurate MSI fragment length assessment. Factors such as PCR cycle number, laboratory conditions, and DNA quality can influence the occurrence of stutter bands. A thorough understanding of these factors is vital for preventing diagnostic errors. Moreover, improving our knowledge of changes during PCR amplification can advance our understanding of the evolvement of these mutations in microsatellites in vivo.

Aims & Methods

Fluorescent multiplex PCR for the BAT25, BAT26, BAT40, and CAT25 microsatellite fragments was performed on multiple microsatellite stable and microsatellite instable samples across different PCR cycle counts. Peak distribution was assessed for each reaction, and peak profiles were evaluated in relation to cycle numbers for different template DNA samples analyzed. In addition, we analyzed peak profiles to infer the potential fragment lengths prior to amplification, i.e. closest to the original sample.

Results

We analyzed the formation of stutter bands during PCR analysis in microsatellite fragments and were able to highlight a shift in the distributions of PCR peaks with increasing cycle count. Amplified fragment sizes tended to be of shorter length with higher numbers of cycles. This was paralleled by an increase of stutter bands in high compared to low cycle reactions. Additionally, in silico modeling of our peak data demonstrates how the length of microsatellite markers, the read depth and the number of PCR cycles affect the reliability and interpretability of MSI detection PCR.

Conclusion

Our results indicate that optimizing reaction conditions is a critical step to standardizing MSI detection by PCR. This may have important implications in MSI diagnostic procedures, particularly when assessing small samples with low DNA template concentration or tumors with borderline MSI status. Our observations suggest that a comprehensive understanding of the peak data produced in fluorescent multiplex PCR can significantly improve diagnostic capability in detecting MSI status in tumor samples and can further insight into the development of MSI mutations in vivo. Developing mathematical models that describe cycle-wise changes of peak distributions is the next step to improve the accuracy of PCR-based MSI testing.

Introduction

Resection of large non-pedunculated colorectal polyps (LNPCP) located in the rectum varies by institution and region with some, particularly colorectal surgeons, favouring TEM and others endoscopic resection (ER). There is a paucity of data directly comparing these strategies and the procedure of choice remains under debate. Our unit is a tertiary referral centre for complex polyps and early colorectal cancer. All rectal LNPCP were treated by TEM until a change in practice from 2011 when the first-line therapeutic strategy for rectal LNPCP changed to ER.

Aims & Methods

We aimed to report the outcomes of TEM and ER of rectal LNPCP from a large tertiary cohort. Large (≥20mm) rectal adenomas resected by TEM or ER (2009 – 2023) were analysed. From 2009-2011 all rectal adenomas were treated by TEM, after which the primary resection strategy changed to ER. Outcomes were compared between techniques. Given the difference in resection strategy, a composite outcome of septic complications included clinically significant perforation for ER (early or delayed perforation requiring any form of unplanned medical or surgical treatment) and septic complications for TEM (peritonitis or pelvic/anorectal sepsis).

Results

704 rectal adenomas were resected by TEM (n=71), EMR (n=145) and ESD/Hybrid ESD (n=488) with a mean size of 45mm for TEM vs 55mm for ER (p<0.001). 100% of patients undergoing TEM required hospital admission vs 15% for ER (p<0.001), and 100% required general anaesthesia (GA) for TEM vs 26%% for ER (GA or propofol sedation, p<0.001). En bloc resection was achieved for 96% of ESD resections, 54% of TEMS, 28% of EMR and 14% of hybrid ESD (p<0.001). The incidence of complications were similar (9.9% for TEM versus 5.0% for ER, p=0.10). Recurrence occurred in 2.7% after ESD, 21.1% after EMR, 23.6% after Hybrid ESD and 36.4% after TEM (p<0.001). No patients undergoing ER or TEM required surgery as a result of a complication.

Conclusion

Despite significantly larger lesion sizes, ER in our institution provides far superior results for patients than TEM with relatively fewer patients requiring hospital admission and lower recurrence rates, especially with ESD. En bloc resection was more likely with ER as a result of the difficulty achieving en bloc resection for extremely large lesions with TEM, which can invariably be resected en bloc by ESD. These results have led to a policy of ER for all apparently benign rectal tumours regardless of size and, in recent years, ESD as the primary approach for all rectal lesions.

Introduction

The United Kingdom National Institute for Health and Care Excellence recommends Faecal Immunochemical Testing (FIT) for all patients presenting to primary care with symptoms suggestive of colorectal cancer. How use of this test varies across England is unknown, which this study aimed to quantify.

Aims & Methods

Routinely-collected primary care records (Clinical Practice Research Datalink Aurum) were used to assess variations in symptomatic FIT use in adults (≥18 years) between regions and over time in England from 2019 to 2023. Rates were calculated using the population registered to each primary care practice as the denominator, with analysis split by calendar year.
The first coded FIT test with a numerical result per patient each year was extracted from the primary care records. Rates of testing were calculated using Poisson regression, adjusted for other confounders. Results were described between groups, regions and years at the recommended threshold of ≥10 µgHb/g faeces (results above this threshold henceforth referred to as FIT10), describing the proportions likely to have a secondary care referral based on national guidance. Logistic regression was used to compare the impact of year, region, gender and age on the differences in FIT10.

Results

Between 01/01/2019 and 05/06/2023 there were 531,735 FIT results among 495,121 patients and a denominator population of 17,910,681. The number of completed FITs increased from 8,395 in 2019 to 237,708 in 2022 (the last complete year within the study), with a corresponding ~thirty-fold increase from 0.69 tests per 1000 person-years in 2019 (95%CI 0.68-0.71) to 19.57 per thousand person-years in 2022 (95%CI 19.50-19.65). There was large variation in testing between regions, with rates 3.6 times higher in the North East compared to the West Midlands (17.05 per thousand person-years, 95%CI 16.87-17.23 versus 4.72 per thousand person-years, 95%CI 4.67-4.76). These differences persisted to 2022, with less than half the rate in the West Midlands (11.32) compared to the North East (31.85 per thousand person-years).
20.4% of results were FIT10. Despite increased testing, the overall proportion FIT10 did not change over time. 51.7% of all tests FIT10 were in those over 70 years of age. The percentage FIT10 was lower in regions with higher rates of FIT testing, for example FIT10 ranged from 16.7% in the South West (rate of testing 14.62 per thousand person-years) to 25.3% in the South East (rate of testing 8.00 per thousand person-years). After adjusting for year, region, gender and age, this difference persisted (adjusted OR 0.57, 95%CI 0.55-0.58) comparing FIT10 in the South West to the South East. The difference in proportions FIT10 between regions showed some convergence over time, from a range of 16.6%-26.7% (10.1%) in 2021 to 15.8%-23.7% (7.9%) in 2023.

Conclusion

There has been a marked increase over time in the use of FIT for patients with symptoms in primary care, with large and persistent variations between regions of England. The overall proportion FIT10 has not decreased over time with increasing use, however there were large differences in FIT10 between regions that correlated with rates of testing. These regional differences persisted but reduced over time as testing rates increased nationally. It is likely that clinical pathway differences in the implementation of FIT over time and between regions explain the variations we have observed. If used as a guide to referral, such variation could lead to inequalities in colorectal investigation and diagnosis.

Clinical Case Summary

Gastrointestinal tuberculosis (GITB) is an uncommon form of extrapulmonary tuberculosis that poses a diagnostic challenge due to its non-specific clinical presentation. Among its varied manifestations, gastrointestinal bleeding as an initial presentation is exceedingly rare. This case report aims to highlight the diagnostic difficulties, treatment modalities, and clinical outcomes in a patient with GITB presenting as gastrointestinal bleeding.
This is a case of a 24/M, no known comorbidities, immunocompetent host, no maintenance medications who came in for sudden onset of hematochezia with progressive drop in hemoglobin levels. Work up revealed a circumferential wall thickening involving the ileocecal region. Bleeding from mass was difficult to control despite medications and colonoscopy attempts hence patient underwent exploratory Laparotomy, right segmental colectomy. Histopathology results revealed caseating chronic granulomatous inflammation and lymph nodes showing caseating granulomas. Patient was then started on anti-Kochs treatment with note of improvement of symptoms post treatment.

References

1. Sharma, V., Mandavdhare, H. S., Lamoria, S., & Dutta, U. (2019). Gastrointestinal tuberculosis: A systematic review and meta-analysis. Scandinavian Journal of Gastroenterology, 54(5), 567-577. doi:10.1080/00365521.2019.1598343
2. Kapoor, A., & Schewitz, I. (2020). Gastrointestinal tuberculosis: Understanding the molecular mechanisms for better diagnosis and treatment. Journal of Digestive Diseases, 21(4), 195-204. doi:10.1111/1751-2980.12866
3. Amarapurkar, D. N., Patel, N. D., & Amarapurkar, A. D. (2018). Gastrointestinal tuberculosis. Indian Journal of Medical Research, 147(5), 401-416. doi:10.4103/ijmr.IJMR_1004_17
4. Sharma, V., Mandavdhare, H. S., & Lamoria, S. (2020). Update on gastrointestinal tuberculosis: Newer techniques and treatment strategies. World Journal of Gastrointestinal Pharmacology and Therapeutics, 11(3), 10-26. doi:10.4292/wjgpt.v11.i3.10

Disclosure

None

Clinical Case Summary

Leiomyomas are benign soft tissue neoplasms that arise from smooth muscle. They are uncommon in the colon. We report two cases of colonic sessile polyps that underwent polypectomy and histology showed leiomyoma.

The first patient previously had a history of polyps with tubular adenoma with low grade dysplasia. A repeat colonoscopy for polyp surveillance was done which found polyps in the ascending colon, and sigmoid. The sigmoid colon polyp was 4mm in size and removed via cold snare polypectomy (Figure 1).
The second patient had underlying non-alcoholic fatty liver disease cirrhosis and iron deficiency anaemia. Colonoscopy showed colonic polyps which were biopsied (Figure 2).

The histology of the first patient’s sigmoid colon polyp showed nodular spindle cell lesion centered in the submucosa/muscularis mucosa. The lesion was composed of spindle cells in fascicles with bland spindled nuclei and eosinophilic cytoplasm. Desmin and SMA stains were done and they were diffusely positive. CD117 stains were negative. DOG1 showed focal nonspecific staining in lesional cells. The overlying lamina propria also showed aggregates of foamy histiocytes and lymphoid aggregates. These were consistent with a leiomyoma.
The second patient’s histology also showed fascicles of spindle cells and immunohistochemistry stain showed positivity for SMA and desmin but negative for CD117 and DOG-1, consistent with leiomyoma.

As the polyps showed smooth margins endoscopically and has similar appearance to hyperplastic polyps, it was likely thought to have a hyperplastic component on top of leiomyoma, hence the patients are to be monitored and followed up for routine surveillance.

References

Miettinen M, Sobin LH, Am J Surg Pathol. 2001;25(11):1433

Introduction

The depth of early gastric cancer is an important factor in determining the treatment plan such as endoscopic resection or surgery. We developed an artificial intelligence (AI) program using deep neural networks to diagnose mucosal and submucosal cancers using endoscopic images. Performance evaluation tests were conducted for clinical applications.

Aims & Methods

A validation dataset was established comprising 204 patients with early gastric cancer who underwent endoscopic resection or surgery at our hospital during 2018–2021. The lesion sites were the upper part in 16 (8%), middle part in 106 (52%), and lower part in 82 (40%) patients. The median lesion size (range) was 14 (2–90) mm; the lesion depth was mucosal cancer in 136 (67%) and submucosal cancer in 68 (33%) patients. Five white-light images per case at a minimum were used (median: 8, range: 5–22). The primary endpoint was sensitivity of mucosal cancer, defined as the lower limit of the 95% confidence interval (CI) > 75%, based on previous reports on the depth diagnosis of early gastric cancer. The secondary endpoints were specificity and accuracy.

Results

Sensitivity of this program for the diagnosis of mucosal cancer was 91.2% (95% CI: 85.2%–94.9%), indicating the usefulness of this program. Specificity was 66.2% (95% CI: 54.3%–76.3%), and accuracy was 82.8% (95% CI: 77.1%–87.4%).

Conclusion

The AI program developed for diagnosing the depth of early gastric cancer is useful and expected to be applied clinically in the near future.

Introduction

Potential celiac disease (PCD) is a clinical condition characterized by the presence of a positive CD-specific serology and a normal intestinal architecture. Asymptomatic PCD patients are generally addressed to continue on a gluten containing diet (GCD), but long-term risks of this approach have never been explored.

Aims & Methods

In the present study we aim at investigating nutritional and autoimmune complications possibly developing overtime in a cohort of asymptomatic PCD children left on a GCD. We compared parameters of growth, nutritional status, and autoimmunity between time of diagnosis and in occasion of last medical check, after a long-term gluten containing diet.

Results

Altogether we collected data from 171 PCD children with a mean follow-up time of 3 years (range 0.35-15.3 years). During follow-up, despite patients did not reduce the amount of daily gluten intake , anti-tissue transglutaminase (anti-TG2) antibodies spontaneously and significantly decreased. Most of parameters analysed did not change over follow-up (height centile, ferritin, albumin., cholesterol, calcium, alkaline phosphatase , parathormone, vitamin D, p>0.05) or even improved (weight and BMI centile, haemoglobin, blood iron, HDL, glycaemia and HbA1C, p<0.05), always remaining in the limit of normality. Equally autoantibodies for other concomitant autoimmune disorders did not increase overtime. Similar results were obtained excluding from the analysis patients that stopped producing anti-TG2 and those with a follow up < 3 years.

Conclusion

Our pilot study provides reassuring results on the maintenance of a gluten containing diet in asymptomatic PCD children, even when a long term follow-up is considered.

Disclosure

no conflict of interest to declare