Clinical Impact of KRAS and GNAS Analysis Added to CEA and Cytology in Pancreatic Cystic Fluid Obtained by EUS-FNA.

BACKGROUND: Pancreatic cysts are common incidental findings with malignant potential, raising diagnostic and treatment dilemmas. AIMS: To determine the added value of KRAS and GNAS mutation analysis on cyst classification and decision making. METHODS: We analyzed 52 frozen samples of pancreatic cystic fluid obtained by EUS-FNA between 2008 and 2014. In addition to cytology and CEA, mutations of GNAS (exons 8 and 9) and KRAS (exons 2 and 3) genes were analyzed using Sanger sequencing. RESULTS: There were 52 patients, 67% females, with a mean age of 59 ± 15 years (29-91). Cysts were classified as mucinous in 21 patients (40%) (14 low-risk, seven malignant) and non-mucinous in 31 patients (60%). After EUS-FNA, 11 patients had surgery, six had chemotherapy or palliation, one had endoscopic drainage, and 34 are on follow-up after a mean of 57 months. KRAS mutation was detected in nine and GNAS in two samples. Patients harboring cysts with KRAS mutations were older (p = 0.01), cysts were more commonly mucinous (p = 0.001) and malignant (p = 0.01). KRAS mutations were present in both low-risk and malignant mucinous lesions. For identifying mucinous lesions, CEA > 192 ng/mL performed better (AUC ROC = 93%), whereas for malignant/high-risk mucinous lesions, EUS imaging had the best accuracy (AUC ROC = 88%). After molecular analysis, a modification in cyst classification occurred in ten patients, but was correct in only two, a pseudocyst re-classified as IPMN and a malignant cyst as a non-mucinous cyst. CONCLUSIONS: In this cohort of patients with pancreatic cysts, KRAS and GNAS mutations had no significant diagnostic benefit in comparison with conventional testing.

Incidence and risk factors for neoplasia in inflammatory bowel disease.

INTRODUCTION: Inflammatory Bowel Disease (IBD) patients may have an increased risk of neoplasia. The aim was to evaluate the incidence of malignant neoplasia in IBD patients, associated risk factors and therapy adjustments. METHODS: Unicentric retrospective cohort study. All patients followed for IBD in a tertiary portuguese hospital and oncological centre during 2015-2020 were included. RESULTS: 318 patients were included female 55.0%, age at diagnosis = 37.24(±15,28), Crohn's disease 52.5%, Primary Sclerosing Cholangitis n = 7, family history of cancer n = 12, previous diagnosis of neoplasia n = 23(7.2%). 42 cancers were diagnosed in 35 patients (11.0%) - median of 12.0(IQR = 7.5-21.0) years after IBD diagnosis. Most affected organs were the skin (n = 15 in 11 patients; melanoma = 1), colon/rectum (n = 8 in 6 patients), prostate (n = 4), breast (n = 3) and anal canal (n = 2). In those with non-melanoma skin cancer, 6 were under active treatment with azathioprine and 2 had stopped it for more than two years. In the univariate analysis, the occurrence of neoplasia was positively associated with tobacco exposure (p = 0.022), age at IBD diagnosis (p = 0.021), and negatively with infliximab exposure (p = 0.046). In 9 cases, cancer treatment was different because of the IBD, while IBD treatment was changed in 9 patients. In those affected by cancer, in the univariate analysis, its cure/remission was negatively associated with tobacco exposure (p = 0.004) and positively with salicylates use (p = 0.007). CONCLUSION: In IBD patients, cancer mostly affected the skin and the lower digestive system. As in the general population, tobacco exposure was a risk factor for the development of neoplasia.

Socio-economic and environmental factors associated with the occurrence of canine infection by Leishmania infantum in Teresina, Brazil.

American visceral leishmaniasis (AVL) is a zoonosis caused by protozoan parasites of the genus Leishmania. In Brazil, the disease is caused by Leishmania infantum and the main vector is the phlebotomine sandfly Lutzomyia longipalpis, found both in natural ecotopes and in the rural and urban environments, being very well adapted to the peridomestic environment. The domestic dog has been incriminated as the main reservoir of the parasite in the urban environment, but the control measures based on culling seropositive dogs have not shown to be effective to contain the spread of the disease throughout the country. Many studies evaluated risk factors for human visceral leishmaniasis but few focused on the socioeconomic and environmental factors associated with infection among dogs. Knowledge of these factors might help identify the conditions that contribute to the maintenance of transmission cycles in the urban environment and identify new targets for intervention. The objective of this study was to assess the association between socioeconomic and environmental factors and the occurrence of canine leishmaniasis at Teresina city, Brazil. This cross-sectional study was developed in ten districts of Teresina, involving 532 houses and 810 dogs. Peripheral blood samples were collected by vein punction using vacutainer tubes without anticoagulant for performing serological test (indirect immunofluorescence - IFI). Serum samples with IFI titers ≥1: 80 were considered positive. Owners of the selected dwellings were interviewed using a semi-structured questionnaire addressing socioeconomic and environment aspects. The association between variables and seropositivity was assessed through multilevel logistic regression models. Global seropositivity was 39%. There was no statistically significant difference between seropositivity and age and sex of animals, literacy of the household head, presence of other domestic animals or with household characteristic like water supply, inadequate sewage disposal system, type of floor and roof. Mixed-breed dogs and those living for a long time in houses with absence of masonry walls and presence of a kennel showed higher odds of seropositivity. These results suggest that some peridomestic characteristics, especially the absence of barriers that allow dogs to have free access to the street, in association with the presence of a kennel, might contribute to maintaining the infection cycle in urban areas. Intervention measures oriented to the management of the peridomestic environment and responsible dog possession could be useful tools for reducing disease burden in endemic area.

Colon Adenocarcinoma Stage IIA-Can We Predict Relapse?

PURPOSE: To determine prognostic factors for stage IIA colon cancer (CC) recurrence in patients undergoing curative intent surgery without adjuvant treatment. METHODS: Single-centre cohort study. All patients with stage IIA CC discussed in a multidisciplinary colorectal cancer clinic from January 2010 to December 2012 were evaluated. Clinical data, laboratory data and tumour features, including expression of DNA repair proteins (EDRP), were analysed. Assessment of overall and disease free survival, recurrence, recurrence site and recurrence's method of diagnosis was performed. The associations between variables were tested through the Fisher's exact test (SPSS 23). RESULTS: Fifty-five patients were included (55% male gender; mean age at diagnosis was 70.3 years (42-88)). CC was in the left colon in 62%, high grade in 7% and had lymphovascular invasion in 7% of the cases. Only one patient was submitted to emergent surgery for obstructive symptoms. In 55% of cases ≥ 12 lymph nodes were collected. There was EDRP loss in nine patients (MLH1/PMS2: six; MSH2/MSH6: three)-only two fulfilled revised Bethesda criteria. Recurrence occurred in five patients (8.9%), and it was diagnosed through surveillance in all of them. No variable showed a statistically significant association with recurrence; however, there were no recurrences in patients with EPRD loss (p = 0.209). Mean follow-up time was 43 months (2-70). In those with recurrence, mean disease-free survival was 23.4 months. CONCLUSIONS: The overall good prognosis and absence of recurrence predictive factors were confirmed, validating the decision of not to submit stage IIA CC patients to chemotherapy risks.

Detection of anti-infliximab antibodies is impacted by antibody titer, infliximab level and IgG4 antibodies: a systematic comparison of three different assays.

BACKGROUND: There is scant information on the accuracy of different assays used to measure anti-infliximab antibodies (ADAs), especially in the presence of detectable infliximab (IFX). We thus aimed to evaluate and compare three different assays for the detection of IFX and ADAs and to clarify the impact of the presence of circulating IFX on the accuracy of the ADA assays. METHODS: Blood samples from 79 ulcerative colitis (UC) patients treated with infliximab were assessed for IFX levels and ADAs using three different assays: an in-house assay and two commercial kits, Immundiagnostik and Theradiag. Sera samples with ADAs and undetectable levels of IFX were spiked with exogenous IFX and analyzed for ADAs. RESULTS: The three assays showed 81-96% agreement for the measured IFX level. However, the in-house assay and Immundiagnostik assays detected ADAs in 34 out of 79 samples, whereas Theradiag only detected ADAs in 24 samples. Samples negative for ADAs with Theradiag, but ADA-positive in both the in-house and Immundiagnostik assays, were positive for IFX or IgG4 ADAs. In spiking experiments, a low concentration of exogenous IFX (5 µg/ml) hampered ADA detection with Theradiag in sera samples with ADA levels of between 3 and 10 µg/ml. In the Immundiagnostik assay detection interference was only observed at concentrations of exogenous IFX higher than 30 µg/ml. However, in samples with high levels of ADAs (>25 µg/ml) interference was only observed at IFX concentrations higher than 100 µg/ml in all three assays. Binary (IFX/ADA) stratification of the results showed that IFX+/ADA- and IFX-/ADAs+ were less influenced by the assay results than the double-positive (IFX+/ADAs+) and double-negative (IFX-/ADAs-) combination. CONCLUSIONS: All three methodologies are equally suitable for measuring IFX levels. However, erroneous therapeutic decisions may occur when patients show double-negative (IFX-/ADAs-) or double-positive (IFX+/ADAs+) status, since agreement between assays is significantly lower in these circumstances.

Clinical and genetic factors predicting response to therapy in patients with Crohn's disease.

AIM: To identify clinical and/or genetic predictors of response to several therapies in Crohn's disease (CD) patients. METHODS: We included 242 patients with CD (133 females) aged (mean ± standard deviation) 39 ± 12 years and a disease duration of 12 ± 8 years. The single-nucleotide polymorphisms (SNPs) studied were ABCB1 C3435T and G2677T/A, IL23R G1142A, C2370A, and G9T, CASP9 C93T, Fas G670A and LgC844T, and ATG16L1 A898G. Genotyping was performed with real-time PCR with Taqman probes. RESULTS: Older patients responded better to 5-aminosalicylic acid (5-ASA) and to azathioprine (OR 1.07, p = 0.003 and OR 1.03, p = 0.01, respectively) while younger ones responded better to biologicals (OR 0.95, p = 0.06). Previous surgery negatively influenced response to 5-ASA compounds (OR 0.25, p = 0.05), but favoured response to azathioprine (OR 2.1, p = 0.04). In respect to genetic predictors, we observed that heterozygotes for ATGL16L1 SNP had a significantly higher chance of responding to corticosteroids (OR 2.51, p = 0.04), while homozygotes for Casp9 C93T SNP had a lower chance of responding both to corticosteroids and to azathioprine (OR 0.23, p = 0.03 and OR 0.08, p = 0.02,). TT carriers of ABCB1 C3435T SNP had a higher chance of responding to azathioprine (OR 2.38, p = 0.01), while carriers of ABCB1 G2677T/A SNP, as well as responding better to azathioprine (OR 1.89, p = 0.07), had a lower chance of responding to biologicals (OR 0.31, p = 0.07), which became significant after adjusting for gender (OR 0.75, p = 0.005). CONCLUSIONS: In the present study, we were able to identify a number of clinical and genetic predictors of response to several therapies which may become of potential utility in clinical practice. These are preliminary results that need to be replicated in future pharmacogenomic studies.

Bone marrow-derived CD11b+Jagged2+ cells promote epithelial-to-mesenchymal transition and metastasization in colorectal cancer.

Timely detection of colorectal cancer metastases may permit improvements in their clinical management. Here, we investigated a putative role for bone marrow-derived cells in the induction of epithelial-to-mesenchymal transition (EMT) as a marker for onset of metastasis. In ectopic and orthotopic mouse models of colorectal cancer, bone marrow-derived CD11b(Itgam)(+)Jagged2 (Jag2)(+) cells infiltrated primary tumors and surrounded tumor cells that exhibited diminished expression of E-cadherin and increased expression of vimentin, 2 hallmarks of EMT. In vitro coculture experiments showed that the bone marrow-derived CD11b(+)Jag2(+) cells induced EMT through a Notch-dependent pathway. Using neutralizing antibodies, we imposed a blockade on CD11b(+) cells' recruitment to tumors, which decreased the tumor-infiltrating CD11b(+)Jag2(+) cell population of interest, decreasing tumor growth, restoring E-cadherin expression, and delaying EMT. In support of these results, we found that peripheral blood levels of CD11b(+)Jag2(+) cells in mouse models of colorectal cancer and in a cohort of untreated patients with colorectal cancer were indicative of metastatic disease. In patients with colorectal cancer, the presence of circulating CD11b(+)Jag2(+) cells was accompanied by loss of E-cadherin in the corresponding patient tumors. Taken together, our results show that bone marrow-derived CD11b(+)Jag2(+) cells, which infiltrate primary colorectal tumors, are sufficient to induce EMT in tumor cells, thereby triggering onset of metastasis. Furthermore, they argue that quantifying circulating CD11b(+)Jag2(+) cells in patients may offer an indicator of colorectal cancer progression to metastatic levels of the disease.

Factors associated with Leishmania chagasi infection in domestic dogs from Teresina, State of Piauí, Brazil.

INTRODUCTION: Many studies have evaluated risk factors for human visceral leishmaniasis, but few have focused on the infection among dogs. The objective of this study was to assess the association between peridomestic socioeconomic and environmental factors and the presence of dogs seropositive for Leishmania chagasi in the City of Teresina, Brazil. METHODS: This case-control study was based on the results of a routine seroepidemiological survey among domestic dogs carried out in 2007. Serological tests were performed by means of indirect immunofluorescence antibody test. All dwellings in which at least one seropositive dog was detected were considered cases, and controls were a random sample of dwellings in which only seronegative dogs were identified. Associations between variables were expressed as odds ratios (OR) and their respective 95% confidence intervals (95%CI) estimated using multivariate logistic regression. RESULTS: Dwellings with a history of dogs removed by the visceral leishmaniasis control program in the last 12 months had five-fold higher odds of having at least one seropositive dog as compared with dwellings having no history of dog removal (OR = 5.19; 95%CI = 3.20-8.42). Dwellings with cats had 58% increased odds of dog infection as compared with those having no cats (OR = 1.58; 95%CI = 1.01-2.47). CONCLUSIONS: Identification of factors associated with canine visceral leishmaniasis might be used for the delimitation of areas of higher risk for human visceral leishmaniasis, since infection in dogs generally precedes the appearance of human cases.