Rectal Biopsy Research Articles

Image-based consensus molecular subtyping in rectal cancer biopsies and response to neoadjuvant chemoradiotherapy

The development of deep learning (DL) models to predict the consensus molecular subtypes (CMS) from histopathology images (imCMS) is a promising and cost-effective strategy to support patient stratification. Here, we investigate whether imCMS calls generated from whole slide histopathology images (WSIs) of rectal cancer (RC) pre-treatment biopsies are associated with pathological complete response (pCR) to neoadjuvant long course chemoradiotherapy (LCRT) with single agent fluoropyrimidine. DL models were trained to classify WSIs of colorectal cancers stained with hematoxylin and eosin into one of the four CMS classes using a multi-centric dataset of resection and biopsy specimens (n = 1057 WSIs) with paired transcriptional data. Classifiers were tested on a held out RC biopsy cohort (ARISTOTLE) and correlated with pCR to LCRT in an independent dataset merging two RC cohorts (ARISTOTLE, n = 114 and SALZBURG, n = 55 patients). DL models predicted CMS with high classification performance in multiple comparative analyses. In the independent cohorts (ARISTOTLE, SALZBURG), cases with WSIs classified as imCMS1 had a significantly higher likelihood of achieving pCR (OR = 2.69, 95% CI 1.01–7.17, p = 0.048). Conversely, imCMS4 was associated with lack of pCR (OR = 0.25, 95% CI 0.07–0.88, p = 0.031). Classification maps demonstrated pathologist-interpretable associations with high stromal content in imCMS4 cases, associated with poor outcome. No significant association was found in imCMS2 or imCMS3. imCMS classification of pre-treatment biopsies is a fast and inexpensive solution to identify patient groups that could benefit from neoadjuvant LCRT. The significant associations between imCMS1/imCMS4 with pCR suggest the existence of predictive morphological features that could enhance standard pathological assessment.

Clinical Relevance of Pathological Diagnosis of Hirschsprung's Disease with Acetylcholine-Esterase Histochemistry or Calretinin Immunohistochemistry.

Hirschsprung disease (HD) manifests as a developmental anomaly affecting the enteric nervous system, where there is an absence of ganglion cells in the lower part of the intestine. This deficiency leads to functional blockages within the intestines. HD is usually confirmed or ruled out through rectal biopsy. The identification of any ganglion cells through hematoxylin and eosin (H&E) staining rules out HD. If ganglion cells are absent, further staining with acetylcholine-esterase (AChE) histochemistry or calretinin immunohistochemistry (IHC) forms part of the standard procedure for determining a diagnosis of HD. In 2017, our Institute of Pathology at University Hospital of Heidelberg changed our HD diagnostic procedure from AChE histochemistry to calretinin IHC. In this paper, we report the impact of the diagnostic procedure change on surgical HD therapy procedures and on the clinical outcome of HD patients. We conducted a retrospective review of the diagnostic procedures, clinical data, and postoperative progress of 29 patients who underwent surgical treatment for HD in the Department of Pediatric Surgery, University of Heidelberg, between 2012 and 2021. The patient sample was divided into two groups, each covering a treatment period of 5 years. In 2012-2016, HD diagnosis was performed exclusively using AChE histochemistry (AChE group, n = 17). In 2017-2021, HD diagnosis was performed exclusively using calretinin IHC (CR group, n = 12). There were no significant differences between the groups in sex distribution, weeks of gestation, birth weight, length of the aganglionic segment, or associated congenital anomalies. Almost half of the children in the AChE group, twice as many as in the CR group, required an enterostomy before transanal endorectal pull-through procedure (TERPT). In the AChE group, 4 patients (23.5%) required repeat bowel sampling to confirm the diagnosis. Compared to the AChE group, more children in the CR group suffered from constipation post TERPT. Elevated AChE expression is linked to hypertrophied extrinsic cholinergic nerve fibers in the aganglionic segment in the majority of patients with HD. The manifestation of increased AChE expression develops over time. Therefore, in neonatal patients with HD, especially those in the first 3 weeks of life, an increase in AChE reaction is not detected. Calretinin IHC reliably identifies the presence or absence of ganglion cells and offers multiple benefits over AChE histochemistry. These include the ability to perform the test on paraffin-embedded tissue sections, a straightforward staining pattern, a clear binary interpretation (negative or positive), cost-effectiveness, and utility regardless of patient age. The ability of calretinin IHC to diagnose HD early and time-independently prevented repeated intestinal biopsies in our patient population and allowed us to perform a one-stage TERPT in the first months of life, reducing the number of enterostomies and restoring colonic continuity early. Patients undergoing transanal pull-through under the age of 3 months require a close follow-up to detect cases with bowel movement problems.

Cardiac amyloidosis – A rarely acknowledged cause of heart failure: A case report and review of literature

Amyloidosis is characterized by the precipitation of insoluble, misfolded, fibrillary proteins in the extracellular matrix in the form of beta-pleated sheets, leading to the loss of normal tissue architecture and ultimately organ dysfunction. The manifestations of the disease are due to the noxious effects of aggregated proteins (Amyloid) deposited in the tissues. Amyloidosis involves multiple organs, including the heart, kidneys, liver, soft tissues, peripheral and/or autonomic nervous system, and gastrointestinal tract. Cardiac amyloidosis is one of the myriad manifestations of systemic amyloidosis. It is characterized by extracellular deposition of amyloid fibrils, leading to progressive cardiac failure. We report a case of a 54-year-old male, who presented to us with a history of exertional dyspnea for the past 2 years with left ventricular systolic and diastolic dysfunction unexplained by the coronary anatomy. The findings on cardiac imaging led us to the suspicion of cardiac amyloidosis. Further, evaluation with an abdominal fat pad and rectal mucosal biopsy confirmed amyloidosis.

Frequency of loss of calretinin expression in clinically susceptible cases of Hirschprung disease in rectal biopsies

Objective: To determine the frequency of loss of calretinin expression in clinically susceptible cases of Hirschprung disease in rectal biopsies. Material and Methods: This descriptive, cross-sectional study was conducted at Department of Histopathology, Combined Military Hospital, Multan for a period of 6 months (17th Oct 2021 to 16th April 2022). A total 93 suspected cases of Hirschprung disease were taken. Specimens received in 10 % buffered formalin and fulfilling inclusion criteria were included in the study. The cases diagnosed as Hirschprung disease were taken and calretinin immunohistochemical stain was applied and result recorded after evaluation by consultant histopathologist. Data was analyzed using (Statistical Software for Social Sciences (SPSS) version 19). Results: Out of 93 cases, 67 (72.0%) were male patients, whereas 26 (28.0%) were female patients. The average age of the study participants was 7.95 months, with a minimum age of 1 month and a maximum age of 12 months. Twenty-eight (30.1%) of the 93 study cases were from rural areas, while 65 (69.9%) were from metropolitan areas. Twenty-five families (26.9%) reported monthly incomes of up to Rs. 35000, while 68 families (73.1%) reported incomes of over Rs. 35,000. Of these 93 study cases, failure to pass meconium was noted in 24 (25.8%) patients, constipation was noted in 75 (80.6%) and abdominal distension was noted in 62 (66.7%) patients. Loss of calretinin expression was noted in 34 (36.6%) of our study cases. Conclusion: According to the findings of our investigation, calretinin is very helpful in identifying suspicious cases of Hirschsprung disease. When acetylcholinesterase enzyme histochemistry is not accessible, it can be a helpful, cost-effective diagnostic assistance in centers with limited resources. It is a reliable and efficient diagnostic tool for early diagnosis of the illness. Loss of Calretinin expression was significantly associated with age and residential status. Keywords: Calretinin expression, Frequency, Hirschsprung disease, Immunohistochemistry

Клініко-патологічна характеристика хвороби Гіршпрунга в дітей шкільного віку

In modern literature, there are only isolated cases of diagnosis and treatment of Hirschsprung disease (HD) in school-aged children, and these cases are from pediatric surgical clinics in developing countries. Aim - to investigate the clinical and pathological symptoms of HD in school-aged children for early detection of this pathology. Materials and methods. We conducted a retrospective analysis of the clinical symptoms of HD in 196 school-aged children for the period from 1980 to 2021. There were 117 (59.69%) patients with the rectal form of HD, 76 (38.78%) with the rectosigmoid form, 3 (1.53%) with the subtotal form. To study the clinical course, we considered the data from the anamnesis, physical examination, general clinical laboratory parameters (blood, urine and feces), electrocardiography, echocardiography, ultrasonography, irrigography, manometry and rectal biopsy. Results. Among 196 school-aged patients with HD, we found enterocolitis in 86 (43.88%) patients: the 1st degree - in 16 (8.16%), the 2nd degree - in 29 (14.79%), and the 3rd degree - in 41 (20.92%) patients. Toxic megacolon was present in 9 (5.83%) patients. Anemia of various degrees was established in 24 (12.24%) patients. Associated malformations were found in 61 (31.12%) children, of which 18 (9.18%) patients had associated malformations of the intestine. The acute form of HD was established in 70 (35.71%) patients, subacute - in 64 (32.65%) children and chronic - in 62 (31.64%) patients. Conclusions. Clinical signs of HD in school-aged children are typical, which can be obtained by careful collection of anamnesis and evaluation of the patient’s objective examination. Irigography, manometry, and rectal biopsy are required to confirm the diagnosis of HD in school-aged children. The research was carried out in accordance with the principles of the Declaration of Helsinki. The research protocol was approved by the Local Ethics Committee of all institutions mentioned in the work. Informed consent of the children’s parents was obtained for the research. No conflict of interests was declared by the authors.

Abstract 2215: Effects of a high-fiber, high-fruit and -vegetable, low-fat dietary intervention on rectal tissue bacteria

Abstract Introduction. Emerging evidence suggests that the bacteria residing in colon and rectal tissue are plausibly associated with colorectal cancer (CRC). Prior studies investigated the effects of dietary interventions on the fecal microbiome but few assessed colorectal tissue microbiome endpoints. We investigated the effects of a high-fiber, high-fruit and -vegetable, low-fat dietary intervention on the rectal tissue microbiome in the Polyp Prevention Trial (PPT). Methods. PPT is a 4-year randomized clinical trial, with specific goals including consuming: 1) ≥ 18g of fiber per 1,000 kcal/day; 2) ≥ 3.5 servings of fruits and vegetables per 1,000 kcal/day; and 3) <20% of kcal/day from fat. We extracted DNA from rectal biopsies using the AutoGen Animal Tissue DNA Extraction Kit and sequenced the V4 region of the 16S rRNA gene to characterize the bacteria in biopsies collected at baseline and the end of years 1 and 4 (N=233 in intervention arm and N=222 in control arm). We estimated effects of the intervention on rarefied alpha diversity and a priori-selected presence and relative abundance (presented as percentages summing to 100) of 10 bacteria most strongly historically associated with CRC presence using repeated-measures linear mixed-effects models. We also explored intervention effects on presence/abundance of all bacteria (with Bonferroni correction for multiple testing) and stratified the analysis among intervention arm participants who adhered to 75% of the dietary goals (super compliers) over the 4 years (N=48) and comparable participants in the control arm (N=64). Results. The intervention did not statistically significantly modify rectal tissue alpha diversity. Compared to the control arm, relative abundance of a priori-selected Porphyromonas (absolute difference-in-difference (DiD) at T1 vs. T0 = -0.24 and T4 vs. T0 = -0.12; P=0.004) and Prevotella (absolute DiD at T1 vs. T0 = -0.40 and at T4 vs. T0 = -0.32; P=0.01) were more strongly decreased in the intervention arm. Other bacteria selected for exploratory analyses were not statistically significantly modified by the intervention after accounting for multiple testing. Among super compliers and comparable control arm participants, the findings were similarly weak. Conclusion. Overall, our findings did not strongly support that a high-fiber, -fruit, and -vegetable, and low-fat diet intervention influences rectal tissue microbiome diversity nor the abundance of most bacteria, except for two oral-originating bacteria that were previously associated with CRC presence. It is possible that dietary exposures over longer periods of time (than 4 years) may more strongly influence the tissue microbiome. Future intervention studies, including tissue samples collected throughout the colon and rectum among larger, diverse study populations for longer periods of time are warranted to inform potential primary prevention strategies for CRC. Citation Format: Doratha Armenthus Byrd, Maria Gomez, Stephanie Hogue, Ana Ortega-Villa, Yunhu Wan, Andrew Warner, Bin Zhu, Casey Dagnall, Kristine Jones, Belynda Hicks, Paul Albert, Gwen Murphy, Rashmi Sinha, Emily Vogtmann. Effects of a high-fiber, high-fruit and -vegetable, low-fat dietary intervention on rectal tissue bacteria [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2215.

Hirschsprung’s Disease Diagnosis: A Comparison of Calretinin and CD56 Immunohistochemistry

Background & objectives: Hirschsprung’s disease is attributable to the congenital lack of ganglion cells in the far intestine. Rectal biopsy is deemed critical for its testing. In some instances, regular techniques fail to detect it. This study aims to evaluate the testing role of calretinin and CD56 immunohistochemistry and correlate the results to routine hematoxylin & eosin stained samples. Methods: This retrospective study was conducted in Rizgary Teaching Hospital, Erbil, Kurdistan region, Iraq. Rectal biopsies and colonic resection specimens of the clinically suspected Hirschsprung’s disease patients were collected and stained with calretinin and CD56 then their findings were compared to the hematoxylin and eosin stained sections during the period between February 2016 to October 2021. Results: Fifty patients aged from 3 days to 8 years with a male-to-female ratio of 3.6:1 were examined for rectal and colonic biopsies. Forty out of 50 cases were detected as HD by H&E staining. The sensitivity, specificity, positive predictive value, negative predictive value, and accuracy rate of calretinin were coherent with that of H&E-stained samples (100%), the likelihood ratio was 50 and the kappa test was 1, were superior to CD56 results with sensitivity (100%), specificity (90%), positive predictive value (97.5%), negative predictive value (100%), accuracy rate (98%), likelihood ratio was 27.77 and kappa test was 0.805. Conclusions: Immunohistochemical expression of calretinin is more specific and accurate than CD56 in HD diagnosis. Calretinin is a trustworthy, additional diagnostic tool for better morphological evaluation of ganglion cells and thereby assists in making a reliable diagnosis of HD.

CFTR function is impaired in a subset of patients with pancreatitis carrying rare CFTR variants

BackgroundMany affected by pancreatitis harbor rare variants of the cystic fibrosis (CF) gene, CFTR, which encodes an epithelial chloride/bicarbonate channel. We investigated CFTR function and the effect of CFTR modulator drugs in pancreatitis patients carrying CFTR variants. MethodsNext-generation sequencing was performed to identify CFTR variants. Sweat tests and nasal potential difference (NPD) assays were performed to assess CFTR function in vivo. Intestinal current measurement (ICM) was performed on rectal biopsies. Patient-derived intestinal epithelial monolayers were used to evaluate chloride and bicarbonate transport and the effects of a CFTR modulator combination: elexacaftor, tezacaftor and ivacaftor (ETI). ResultsOf 32 pancreatitis patients carrying CFTR variants, three had CF-causing mutations on both alleles and yielded CF-typical sweat test, NPD and ICM results. Fourteen subjects showed a more modest elevation in sweat chloride levels, including three that were provisionally diagnosed with CF. ICM indicated impaired CFTR function in nine out of 17 non-CF subjects tested. This group of nine included five carrying a wild type CFTR allele. In epithelial monolayers, a reduction in CFTR-dependent chloride transport was found in six out of 14 subjects tested, whereas bicarbonate secretion was reduced in only one individual. In epithelial monolayers of four of these six subjects, ETI improved CFTR function. ConclusionsCFTR function is impaired in a subset of pancreatitis patients carrying CFTR variants. Mutations outside the CFTR locus may contribute to the anion transport defect. Bioassays on patient-derived intestinal tissue and organoids can be used to detect such defects and to assess the effect of CFTR modulators.

Mpox Case Presenting With Genital Lesions and Proctitis.

Monkeypox (Mpox) is a zoonotic Orthopoxvirus of the Poxviridae family, endemic to Africa. In August 2022, the US government declared it an emergency because of the worldwide spread. Traditionally, Mpox infection spreads through contact with infected animals. However, the 2022 outbreak Centers for Disease Control and Prevention (CDC) data note that 94% of cases had recent male-to-male sexual or close intimate contact, suggesting a novel sexual transmission. In this article, we report a 39-year-old HIV-positive man presenting with a diffuse cutaneous rash, perianal pain, and bloody stool of 2-week duration. A medical history includes intravenous drug use and multiple sexual partners. Physical examination revealed umbilicated, tan-colored, crusted cutaneous papules scattered across the face, trunk, and genital regions. Perianal lesion biopsy showed an acanthotic epidermis with spongiosis, ballooning degeneration of keratinocytes, and the formation of multinucleated syncytial keratinocytes. A dermal superficial/lichenoid mixed inflammatory cell infiltrate with multinucleated giant cells was noted. Perianal lesion polymerase chain reaction (PCR) was positive for Mpox. Colonoscopy revealed a 3-cm circumferential rectal ulcer with gray exudate and necrosis. A rectal ulcer biopsy showed an ulcerated mucosa with acute proctitis and necrosis. There were scattered macrophages with intranuclear inclusion and glassy vacuolization, and Mpox infection was confirmed by immunostaining with a Mpox-specific anti-Vaccinia virus antibody.

Delayed diagnosis in Hirschsprung disease.

Diagnostic delay in Hirschsprung disease is uncommon. Different definitions have been proposed but that of a diagnosis achieved after 12months of age seems to be the most reliable and resorted to. Some authors reported a worse outcome in case of delay. Our study aims at providing the most relevant features of a series of patients who received a delayed diagnosis of Hirschsprung disease. All consecutive patients admitted to our Center with a delayed diagnosis of Hirschsprung diseases between January 2017 and July 2023 have been retrospectively enrolled. Demographic data, phenotype, genotype, surgical complications, and outcome were assessed and compared to those of literature. A number of variables were also compared to those of a series of patients admitted during the same study period without a delayed diagnosis. A total of 45 patients were included (16.4% out of a series of 346 patients with data regarding age at diagnosis). Male to female ratio was 3.1:1. Median age at diagnosis was 41months with a wide variation (range between 17months and 58years). All patients but 2 suffered from classic rectosigmoid aganglionosis. Normal meconium passage (58%) was reported in a significantly higher number of patients compared to what observed in a series without diagnostic delay (p = 0.0140). All other variables (associated anomalies, preoperative enterocolitis, complications, and functional outcome) proved not to have statistically significant differences compared to a series of patients without a diagnostic delay. The results of our study underline that a significant percentage of patients are basically missed in the neonatal period mostly due to mild symptoms. Overall outcome does not differ from that of patients without diagnostic delay. Nonetheless, we underline the importance of a throughout investigation of all patients with meconium delay/failure and that of adopting a low threshold for performing rectal suction biopsies in constipated children to avoid misdiagnosis to serve the best for our patients.

Comparison of Immunohistochemical Expression of Calretinin, Map2, S-100 and Glut1 in Rectal Biopsies from Suspected Hirschsprung’s Disease

Background: This study aimed to compare the immunohistochemical expression of calretinin, Map2, S-100, and Glut1 in rectal biopsy samples from patients suspected of Hirschsprung’s disease (HD).Methods and Results: Rectal biopsy samples from 40 patients with suspected HD were analyzed using hematoxylin and eosin (H&E) and immunohistochemistry (IHC). Immunohistochemical stains were assessed after previous routine histology interpretation, which was classified as “Positive or in favor for HD” and “Equivocal or negative for HD.” The staining patterns for calretinin, Map2, S-100, and Glut1 were analyzed regarding the following structures: lamina propria small nerve fibrils, submucosal small nerve fibrils,submucosal nerve fibers, and submucosal ganglia. The IHC stains for calretinin and Map2 were score-ranked as 0 – negative and 1 – positive. The IHC stain for S-100 was score-ranked as 0 – normal and 1 – hypertrophic. The IHC stain for Glut1 was ranked as 0 – normal perineural and 1 – conspicuous perineural accentuation Calretinin had 92% accuracy, the highest sensitivity (80%) and specificity (92.00%).Map2 also had the same accuracy as calretinin but lower sensitivity (46.67%). Regarding S-100 and Glut1, these two markers did not support a conclusive diagnosis. The accuracy for S-100 and Glut1 was 66.7% and 60%, respectively. Conclusion: Calretinin remains the currently most valuable single IHC marker in diagnosing difficult cases of HD.

Pharmacokinetics of single dose doxycycline in the rectum, vagina, and urethra: implications for prevention of bacterial sexually transmitted infections

Clinical trials showed a single oral dose of doxycycline taken after sex protects against STIs among men who have sex with men (MSM) but not women. Pharmacokinetic data at vaginal, rectal and penile sites of STI exposure are lacking. We examined vaginal, rectal and urethral doxycycline concentrations in men and women to better inform STI prevention. Doxycycline pharmacokinetics in male and female participants 18-59 years of age were evaluated in blood and urine and on rectal and vaginal swabs collected at 1, 2, 4, 8, 24, 48, 72, 96 and 168h after receiving a 200mg oral doxycycline dose in a non-randomised single dose open label single centre study in Atlanta, Georgia. Rectal, vaginal, and cervical biopsies and male urethral swabs were collected 24h after dosing (Trial registration: NCT04860505). Doxycycline was measured by liquid chromatography-mass spectrometry. Eleven male and nine female participants participated in the study. Doxycycline concentrations on rectal and vaginal swabs collected up to 96h after dosing were approximately twice those of plasma and remained above minimum inhibitory concentrations (MICs) for at least four, three, and two days for Chlamydia trachomatis, Treponema pallidum, and tetracycline-sensitive Neisseria gonorrhoeae, respectively. Geometric mean doxycycline concentrations in male urethral secretions (1.166μg/mL; 95% CI 0.568-2.394μg/mL), male rectal (0.596μg/g; 0.442-0.803μg/g), vaginal (0.261μg/g; 0.098-0.696μg/g) and cervical tissue (0.410μg/g; 0.193-0.870μg/g) in biopsies collected 24h after dosing exceeded MICs. Plasma and urine doxycycline levels defined adherence markers up to four and seven days postdosing, respectively. No adverse events were reported in this study. Doxycycline efficiently distributes to the rectum, vagina and urethra. Findings can help explain efficacy of STI prevention by doxycycline. Funded by CDC intramural funds, CDC contract HCVJCG-2020-45044 (to CFK).

High-Resolution Anorectal Manometry as a Screening Tool for Hirschsprung's Disease: A Comprehensive Retrospective Analysis.

Hirschsprung's disease (HD) is characterized by a congenital absence of enteric ganglion cells in the intestine, posing challenges in diagnosis, particularly in pediatric patients. The gold standard, rectal suction biopsy (RSB), carries risks, prompting an exploration of non-invasive alternatives such as high-resolution anorectal manometry (HR-ARM) for HD screening. We conducted a retrospective analysis of 136 patients suspected of HD between 2018 and 2022, which were stratified into three age groups: ≤12 months, ≤24 months, and >24 months. Criteria for suspicion included delayed meconium passage, unresponsive chronic constipation, and abnormal prior test results. HR-ARM, supplemented by additional tests, confirmed 16 HD cases. HR-ARM exhibited 93.75% sensitivity, 89.47% specificity, 99.03% negative predictive value (NPV), and 55.56% positive predictive value (PPV). Notably, HR-ARM consistently performed well in patients ≤ 2 years old but demonstrated reduced efficacy in older children, which was likely due to complications from chronic constipation. This study underscores HR-ARM's promise as a non-invasive HD screening tool, especially in younger patients. However, its limitations in older children warrant consideration. Establishing standardized protocols, particularly for assessing the recto-anal inhibitory reflex, is crucial. Further research is imperative to optimize HR-ARM's diagnostic role across varied age groups in HD assessment.

Mucosal Genes Encoding Clock, Inflammation and Their Mutual Regulators Are Disrupted in Pediatric Patients with Active Ulcerative Colitis.

Patients with active ulcerative colitis (UC) display a misalignment of the circadian clock, which plays a vital role in various immune functions. Our aim was to characterize the expression of clock and inflammation genes, and their mutual regulatory genes in treatment-naïve pediatric patients with UC. Using the Inflammatory Bowel Disease Transcriptome and Metatranscriptome Meta-Analysis (IBD TaMMA) platform and R algorithms, we analyzed rectal biopsy transcriptomic data from two cohorts (206 patients with UC vs. 20 healthy controls from the GSE-109142 study, and 43 patients with UC vs. 55 healthy controls from the GSE-117993 study). We compared gene expression levels and correlation of clock genes (BMAL1, CLOCK, PER1, PER2, CRY1, CRY2), inflammatory genes (IκB, IL10, NFκB1, NFκB2, IL6, TNFα) and their mutual regulatory genes (RORα, RORγ, REV-ERBα, PGC1α, PPARα, PPARγ, AMPK, SIRT1) in patients with active UC and healthy controls. The clock genes BMAL1, CLOCK, PER1 and CRY1 and the inflammatory genes IκB, IL10, NFκB1, NFκB2, IL6 and TNFα were significantly upregulated in patients with active UC. The genes encoding the mutual regulators RORα, RORγ, PGC1α, PPARα and PPARγ were significantly downregulated in patients with UC. A uniform pattern of gene expression was found in healthy controls compared to the highly variable expression pattern in patients with UC. Among the healthy controls, inflammatory genes were positively correlated with clock genes and they all showed reduced expression. The difference in gene expression levels was associated with disease severity and endoscopic score but not with histological score. In patients with active UC, clock gene disruption is associated with abnormal mucosal immune response. Disrupted expression of genes encoding clock, inflammation and their mutual regulators together may play a role in active UC.

P626 JNJ-77242113, an oral peptide selectively targeting the IL-23 receptor, demonstrates pharmacodynamic activity in rat and human colon tissue explants following oral dosing

Abstract Background The interleukin (IL)-23 pathway is a pathogenic driver in psoriasis, psoriatic arthritis, and inflammatory bowel disease (IBD). There are currently no approved oral therapies selectively targeting the IL-23 pathway. JNJ-77242113 (JNJ-2113) is a targeted oral peptide that binds the IL-23 receptor with high affinity, and potently and selectively inhibits IL-23 proximal signaling and downstream cytokine production. Oral JNJ-2113 showed dose-dependent inhibition of IL-23–induced IL-17A production in rat blood and attenuated signs of disease in a rat colitis model. A Phase 1 study showed inhibition of IL-23–induced IFN-γ production in blood from participants dosed with JNJ-2113 (Fourie et al. ISID. 2023). Methods We evaluated IL-23–induced IL-22 expression in colon tissue after exposure to JNJ-2113 or JNJ-2100 (a tool molecule with similar affinity and potency). IL-22 mRNA was measured after IL-23 stimulation in rat colon biopsies treated with JNJ-2100 in vitro or in colon tissue from rats 6 h after oral JNJ-2100 dosing. Human colon explants from donors were treated in vitro with JNJ-2100 or JNJ-2113, stimulated with IL-23, and IL-22 mRNA was measured. Colon and rectum biopsies from healthy participants dosed for 10 days with oral JNJ-2113 or placebo in a Phase 1 study were obtained, stimulated with IL-23 ex vivo, and IL-22 mRNA was evaluated. Results Overall, inhibition of IL-23–induced IL-22 expression was observed in colon tissue exposed to JNJ-2100 or JNJ-2113. Dose-dependent inhibition of IL-22 expression was observed in rat colon biopsies exposed to JNJ-2100 in vitro. Colon explants from rats dosed with 0.3, 3, and 30 mg/kg oral JNJ-2100 showed dose-dependent inhibition of IL-22 expression (52%, 88% and 96%, respectively) compared with explants from rats treated with vehicle. Human colon explants exposed to JNJ-2100 or JNJ-2113 in vitro also showed dose-dependent inhibition of IL-23–induced IL-22 expression, with >90% inhibition at the highest doses. Importantly, in colon biopsies from Phase 1 participants dosed with oral JNJ-2113 versus placebo, significant inhibition of IL-23–induced IL-22 expression was seen (P<0.05), with an average of 65% and 66% inhibition observed in sigmoid colon and rectum biopsies, respectively. Conclusion These results demonstrate pharmacodynamic activity of JNJ-2100 in vitro and ex vivo in rat colon tissue, of JNJ-2100 and JNJ-2113 in vitro in human colon explants, and of JNJ-2113 in colon biopsies from healthy human volunteers. These data support development of JNJ-2113 for the treatment of IBD. Of note, a Phase 2b dose-ranging study has demonstrated clinical activity of JNJ-2113 in moderate-to-severe plaque psoriasis (NCT05223868).

P173 Characterization of the gut microbiota and the colonic immune response in acute severe ulcerative colitis : the multi-omics ITAC project

Abstract Background Limited understanding exists regarding the pathogenesis of acute severe ulcerative colitis (UC). Microorganisms are proposed as potential triggers due to the resemblances to infectious colitis and the essential role played by gut microbiota in UC-related inflammation. Our aim was to identify microbiome elements and host factors associated with acute severe UC. Methods This was a prospective study across three referral centers comparing two UC patient groups: acute severe hospitalized UC (as per Truelove and Witts criteria) and non-severe UC. We analyzed gut microbiota using 16S rRNA gene sequencing and conducted single-cell RNA-Seq on rectal biopsies in a subgroup of patients to uncover cellular subtypes and pathways involved in mucosal inflammation. We utilized whole blood RNA-Seq to investigate the host pathways involved in mediating systemic inflammation. Results Forty-one patients (23 (56%) female, median (interquartile range IQR) age 42 (34 – 57) years were included: 19 with acute severe UC and 22 with non-severe UC. Compared to patients with non-severe UC, those with acute severe UC displayed distinct gut microbiota with reduced diversity, increased Proteobacteria (Escherichia/Shigella genus), and decreased Lachnospiraceae and Ruminococcaceae. In severe cases (n=4 - 13,047 cells), single-cell RNA-Seq analysis of rectal biopsies revealed distinctive patterns compared to non-severe cases (n=5 - 18,433 cells): plasmablasts showed an altered transcriptomic profile with heightened IgG expression, and there was an expanded cluster of interleukin (IL) 26 expressing T cell population. Innate immune cells exhibited a pro-inflammatory profile marked by increased expression of the IL1B gene. In blood samples, we observed no distinct differentiation of transcriptomic profiles between severe and non-severe cases. Conclusion Our multi-omics study reveals key cellular and bacterial components in acute severe UC pathogenesis. We identified Proteobacteria, especially Escherichia coli as a potential pathobiont in acute severe UC. At the colonic level, we observed an increased IgG/IgA ratio, while both T cells and innate immune cells indicated a pro-Th17 mucosal environment. Enhanced systemic inflammation in acute severe UC was not reflected by specific changes in immune circulating cells. These insights may pave the way for future research focusing on microbiome modulation, interventions targeting plasmablasts, or nuanced inhibition of the Th17/IL-23 axis in acute severe UC.

P267 Prospective validation of the IBD - Distribution, Chronicity, Activity (DCA) histological score for Ulcerative Colitis and correlation with UCEIS and MES

Abstract Background Histological assessment is not currently recognised as an independent parameter to guide personalised treatment for patients with ulcerative colitis (UC). The development of the Inflammatory Bowel Disease – Distribution, Chronicity, Activity (IBD-DCA) score allows a pragmatic approach to quantitatively evaluate mucosal injury and healing in UC, compared with other pre-existing histological scores (Lang-Schwarz et al., J Crohns Colitis, 2021). This scoring system facilitates a template-based reporting to produce consistent reports with information representing UC severity. Each domain of the IBD-DCA is scored as one of 0 (normal), 1 (mild), or 2 (moderate to severe) based on the presence of established histological UC findings (Figure 1). This project aims to prospectively validate the IBD-DCA score for routine clinical deployment and correlate with endoscopic scores – UC Endoscopic Index of Severity (UCEIS) and Mayo Endoscopic Score (MES). Methods Colonoscopic biopsies from patients with a previously confirmed diagnosis of UC attending one of four hospitals in a single healthcare group from August 2022 to January 2023 were reported using the IBD-DCA histological score, and the corresponding, clinician-reported UCEIS and MES scores were recorded. The histopathology reports were assessed based on the completeness of information in their respective D, C, and A domains. The proportion of reports with information fully represented by the IBD-DCA reporting template and the distribution of biopsy sites were described. The correlation between IBD-DCA with UCEIS and MES was quantified via Kendall’s tau (τ) using the R statistical package v4.3.2. Results Of the 151 cases reported, 70 (46.4%) patients were female and 81 (53.6%) were male. 12 patients (7.9%) had full colonic biopsies, while the remaining cases were assessed with selective biopsies. Rectal biopsy was most often performed (83.4%). The IBD-DCA components were reported for every biopsied sites in 133 (88.1%) cases. Notable correlations included UCEIS vascular pattern with histological chronicity (τ = 0.3891), UCEIS erosions/ulceration with histological activity (τ = 0.3252), and histological activity with the total UCEIS score (τ = 0.3450). MES showed the strongest correlation with histological disease distribution (τ = 0.3450)(Table 1). All p-values < 0.001. Conclusion These data show encouraging uptake of this simple histological reporting template in UC in daily routine use. The findings suggest that the IBD-DCA score may be a useful addition in the clinical management in patients with UC.

Light chain (AL) amyloidosis following gastrointestinal symptoms that involve multiple organs: a case report

Light chain (AL) amyloidosis is a complex and rare disease characterized by low incidence and diverse clinical manifestations. At the time of diagnosis, most patients exhibit involvement of multiple organs or tissues, leading to severe illness and a poor prognosis. Therefore, early diagnosis, active treatment, and a comprehensive assessment of the disease hold paramount importance. The initial presentation of this rare condition often manifests as gastrointestinal symptoms, posing challenges in clinical identification and differential diagnosis. In this case report, we describe a 70-year-old man with AL amyloidosis, initially misdiagnosed as irritable bowel syndrome and colon polyps. Subsequently, he experienced a series of complications including renal function impairment, pulmonary nodule, pleural effusion, mediastinal lymph node enlargement, spleen enlargement, reduction of white blood cells, red blood cells and platelets, and small intestinal obstruction. Despite multiple pulmonary nodule biopsy and lymph node biopsy, as well as concurrent splenectomy and partial resection of the small intestine, a clear diagnosis remained elusive. Before admission, diarrhea was aggravated, accompanied by emaciation and fatigue. Following the completion of serum immunofixed protein electrophoresis, renal biopsy, bone marrow and rectal biopsy, a conclusive diagnosis of AL amyloidosis involving multiple organs (Mayo 2012 revision stage III) was finally confirmed after a 12-year period. Treatment with proteasome inhibitors, immunomodulators, and glucocorticoids was recommended. The patient underwent methylprednisolone treatment and was discharged after symptom improvement. However, eight months into the follow-up, the patient succumbed to multiple organ failure. The repeated misdiagnoses over the past 12 years were attributed to limited perspectives among some specialists who did not conduct a systematic and detailed medical history analysis or comprehensive physical examinations. Additionally, they did not strictly follow the principle of "monism" in diagnosis. On the contrary, early diagnosis and active treatment of the disease are of great significance to improve the prognosis.

A recall-optimised machine learning framework for small data improves risk stratification for Hirschsprung's disease

ObjectiveTo improve the selection of patients with suspected Hirschsprung's disease for rectal biopsy with a recall-optimised machine learning framework that strongly penalises missed diagnoses, identify the most important clinical features, and investigate if this methodology is superior to the current practice of conventional logistic regression analysis. Study designThe study data comprised the medical records of 178 children older than one month who underwent a rectal biopsy for the evaluation of Hirschsprung's disease. Each medical record contained the clinical features recorded before the rectal biopsy and the biopsy result. However, only 20 of the 178 children were diagnosed with Hirschsprung's disease. Thus, as is frequently the case for rare diseases, the dataset was small and imbalanced. We present a machine learning framework that, based on these data, produces a champion machine learning model for predicting the presence of Hirschsprung's disease in patients. Notably, the machine learning framework is recall-optimised, i.e., it strongly penalises missed diagnoses. This is achieved using a combination of synthetic minority over-sampling techniques, Bayesian hyper-parameter optimisation, and bootstrapping-based prediction thresholding for a competing set of models comprised of a logistic regression model, a random forest classifier, and a gradient-boosted classifier. Finally, the machine learning framework evaluates the performance of these three competing models and conventional logistic regression with 5-fold stratified cross-validation using an 80:20 train-test split. Model performance is ranked by the average recall across the cross-validation folds. The model with the highest recall is selected as the champion. The model with the highest accuracy is chosen in case of a tie. ResultsThis study revealed that about 35% of our cohort's children without Hirschsprung's disease could have been spared a rectal biopsy while incurring no missed diagnoses using our machine learning framework. Given our aim of avoiding missed diagnosis, the champion model was vastly superior to conventional logistic regression, i.e., the status quo, which missed 40% of HD-positive cases. Moreover, the champion model pointed to a new hierarchy for the importance of clinical features associated with Hirschsprung's disease. Among all features, “gross abdominal distention” was the most important. ConclusionThere is considerable scope for a stricter selection when referring constipated children for a rectal biopsy to diagnose or exclude Hirschsprung's disease. This study demonstrates that a recall-optimised pipeline based on classical supervised machine learning is superior to the conventional statistical and heuristic approaches used today, also for small and imbalanced datasets. This finding opens a path to better care for patients with rare diseases while alleviating pressures on healthcare systems.

Unexpected outcome of dyspeptic syndrome.

A 57-year-old black male patient was admitted with flatulence and post-prandial fullness a month ago. No other gastrointestinal complaints. He had history of arterial hypertension, medicated with nifedipine 30mg/day and hydrochlorthiazide 50mg/day, and had been diagnosed and treated for malaria a week before admission. The patient reported frequent bathing in rivers and smoking habits since the age of 18, around a pack and a half a day. Abdominal ultrasound showed periportal fibrosis, without findings of portal hypertension. A rectal biopsy was performed with findings of Schistosome mansoni eggs on microscopy. This case illustrates a patient who was admitted due to dyspeptic syndrome, and during the study of his condition was found to have periportal fibrosis, which in a country endemic for Schistosomiasis such as Mozambique, this pathology must always be investigated. A history of bathing in rivers increases the chances of it being Schistosomiasis, confirmed by the demonstration of parasite eggs in a rectal biopsy. Treatment with prazinquantel was done at a dose of 40 mg/kg and outpatient follow-up was scheduled for six months.