The specific seroreactivity to ∆Np73 isoforms shows higher diagnostic ability in colorectal cancer patients than the canonical p73 protein

The p53-family is tightly regulated at transcriptional level. Due to alternative splicing, up to 40 different theoretical proteoforms have been described for p73 and at least 20 and 10 for p53 and p63, respectively. However, only the canonical proteins have been evaluated as autoantibody targets in cancer patients for diagnosis. In this study, we have cloned and expressed in vitro the most upregulated proteoforms of p73, ΔNp73α and ΔNp73β, for the analysis of their seroreactivity by a developed luminescence based immunoassay test using 145 individual plasma from colorectal cancer, premalignant individuals and healthy controls. ∆Np73α seroreactivity showed the highest diagnostic ability to discriminate between groups. The combination of ∆Np73α, ∆Np73β and p73 proteoforms seroreactivity were able to improve their individual diagnostic ability. Competitive inhibition experiments further demonstrated the presence of unique specific epitopes in ΔNp73 isoforms not present in p73, with several colorectal patients showing unique and specific seroreactivity to the ΔNp73 proteoforms. Overall, we have increased the complexity of the humoral immune response to the p53-family in cancer patients, showing that the proteoforms derived from the alternative splicing of p73 possess a higher diagnostic ability than the canonical protein, which might be extensive for p53 and p63 proteins.This work was supported by the Ramon y Cajal programme of the MINECO and the financial support of the PI17CIII/00045 grant from the AES-ISCIII program to R.B., cofounded by FEDER funds. G.D. acknowledges the financial support of PI15/00246 grant of the FIS and Cátedra UAM-Roche en Medicina de Innovación. M.G-A. was supported by a contract of the Programa Operativo de Empleo Juvenil y la Iniciativa de Empleo Juvenil (YEI) with the participation of the Consejería de Educación, Juventud y Deporte de la Comunidad de Madrid y del Fondo Social Europeo. We thank the excellent technical support of Maricruz Sánchez. A.M-C. is a recipient of a FPU fellowship from the Ministerio de Educación, Cultura y Deporte.S

Evaluation of different bowel preparations for small bowel capsule endoscopy: a prospective, randomized, controlled study

To obtain an adequate view of the whole small intestine during capsule endoscopy (CE) a clear liquid diet and overnight fasting is recommended. However, intestinal content can hamper vision in spite of these measures. Our aim was to evaluate tolerance and degree of intestinal cleanliness during CE following three types of bowel preparation. PATIENTS AND METHODS: This was a prospective, multicenter, randomized, controlled study. Two-hundred ninety-one patients underwent one of the following preparations: 4 L of clear liquids (CL) (group A; 92 patients); 90 mL of aqueous sodium phosphate (group B; 89 patients); or 4 L of a polyethylene glycol electrolyte solution (group C; 92 patients). The degree of cleanliness of the small bowel was classified by blinded examiners according to four categories (excellent, good, fair or poor). The degree of patient satisfaction, gastric and small bowel transit times, and diagnostic yield were measured. RESULTS: The degree of cleanliness did not differ significantly between the groups (P = 0.496). Interobserver concordance was fair (k = 0.38). No significant differences were detected between the diagnostic yields of the CE (P = 0.601). Gastric transit time was 35.7 +/- 3.7 min (group A), 46.1 +/- 8.6 min (group B) and 34.6 +/- 5.0 min (group C) (P = 0.417). Small-intestinal transit time was 276.9 +/- 10.7 min (group A), 249.7 +/- 13.1 min (group B) and 245.6 +/- 11.6 min (group C) (P = 0.120). CL was the best tolerated preparation. Compliance with the bowel preparation regimen was lowest in group C (P = 0.008). CONCLUSIONS: A clear liquid diet and overnight fasting is sufficient to achieve an adequate level of cleanliness and is better tolerated by patients than other forms of preparation

Multiplexed monitoring of a novel autoantibody diagnostic signature of colorectal cancer using HaloTag technology-based electrochemical immunosensing platform

Background and Purpose: The humoral immune response in cancer patients can be used for early detection of the disease. Autoantibodies raised against tumor-associated antigens (TAAs) are promising clinical biomarkers for reliable cancer diagnosis, prognosis, and therapy monitoring. In this study, an electrochemical disposable multiplexed immunosensing platform able to integrate difficult- and easy-to-express colorectal cancer (CRC) TAAs is reported for the sensitive determination of eight CRC-specific autoantibodies. Methods: The electrochemical immunosensing approach involves the use of magnetic microcarriers (MBs) as solid supports modified with covalently immobilized HaloTag fusion proteins for the selective capture of specific autoantibodies. After magnetic capture of the modified MBs onto screen-printed carbon working electrodes, the amperometric responses measured using the hydroquinone (HQ)/H2O2 system were related to the levels of autoantibodies in plasma. Results: The biosensing platform was applied to the analysis of autoantibodies against 8 TAAs described for the first time in this work in plasma samples from healthy asymptomatic individuals (n=3), and patients with high-risk of developing CRC (n=3), and from patients already diagnosed with colorectal (n=3), lung (n=2) or breast (n=2) cancer. The developed bioplatform demonstrated an improved discrimination between CRC patients and controls (asymptomatic healthy individuals and breast and lung cancer patients) compared to an ELISA-like luminescence test. Conclusions: The proposed methodology uses a just-in-time produced protein in a simpler protocol, with low sample volume, and involves cost-effective instrumentation, which could be used in a high-throughput manner for reliable population screening to facilitate the detection of early CRC patients at affordable cost.This work was supported by the financial support of the PI17CIII/00045 grant from the AES-ISCIII program to R.B., cofounded by FEDER funds. The financial support of the CTQ2015-64402-C2-1-R (Spanish Ministerio de Economía y Competitividad) and RTI2018-096135-B-I00 (Ministerio de Ciencia, Innovación y Universidades) Research Projects and the TRANSNANOAVANSENS-CM Program from the Comunidad de Madrid (Grant S2018/NMT-4349) to P.Y-S., S.C. and JM.P are gratefully acknowledged. G.D. acknowledges the financial support of PI15/00246 grant of the FIS and Cátedra UAM-Roche en Medicina de Innovación. M.G-A. was supported by a contract of the Programa Operativo de Empleo Juvenil y la Iniciativa de Empleo Juvenil (YEI) with the participation of the Consejería de Educación, Juventud y Deporte de la Comunidad de Madrid y del Fondo Social Europeo. A.M-C. is a recipient of a FPU fellowship from the Ministerio de Educación, Cultura y Deporte. G.S-F. is recipient of a predoctoral contract (grant number 1193818N) supported by The Flanders Research Foundation (FWO). Predoctoral contracts from the Spanish Ministerio de Economía y Competitividad (E.P.) and Universidad Complutense de Madrid (V.R.V-M.) are also gratefully acknowledged.S

Evaluation of different bowel preparations for small bowel capsule endoscopy: a prospective, randomized, controlled study

To obtain an adequate view of the whole small intestine during capsule endoscopy (CE) a clear liquid diet and overnight fasting is recommended. However, intestinal content can hamper vision in spite of these measures. Our aim was to evaluate tolerance and degree of intestinal cleanliness during CE following three types of bowel preparation. PATIENTS AND METHODS: This was a prospective, multicenter, randomized, controlled study. Two-hundred ninety-one patients underwent one of the following preparations: 4 L of clear liquids (CL) (group A; 92 patients); 90 mL of aqueous sodium phosphate (group B; 89 patients); or 4 L of a polyethylene glycol electrolyte solution (group C; 92 patients). The degree of cleanliness of the small bowel was classified by blinded examiners according to four categories (excellent, good, fair or poor). The degree of patient satisfaction, gastric and small bowel transit times, and diagnostic yield were measured. RESULTS: The degree of cleanliness did not differ significantly between the groups (P = 0.496). Interobserver concordance was fair (k = 0.38). No significant differences were detected between the diagnostic yields of the CE (P = 0.601). Gastric transit time was 35.7 +/- 3.7 min (group A), 46.1 +/- 8.6 min (group B) and 34.6 +/- 5.0 min (group C) (P = 0.417). Small-intestinal transit time was 276.9 +/- 10.7 min (group A), 249.7 +/- 13.1 min (group B) and 245.6 +/- 11.6 min (group C) (P = 0.120). CL was the best tolerated preparation. Compliance with the bowel preparation regimen was lowest in group C (P = 0.008). CONCLUSIONS: A clear liquid diet and overnight fasting is sufficient to achieve an adequate level of cleanliness and is better tolerated by patients than other forms of preparation