In the Balkan and Taiwan, the relationship between exposure to aristolochic acid and risk of urothelial neoplasms was inferred from the A>T genetic hallmark in TP53 gene from malignant cells. This study aimed to characterize the TP53 mutational spectrum in urothelial cancers consecutive to Aristolochic Acid Nephropathy in Belgium. Serial frozen tumor sections from female patients (n = 5) exposed to aristolochic acid during weight-loss regimen were alternatively used either for p53 immunostaining or laser microdissection. Tissue areas with at least 60% p53-positive nuclei were selected for microdissecting sections according to p53-positive matching areas. All areas appeared to be carcinoma in situ. After DNA extraction, mutations in the TP53 hot spot region (exons 5–8) were identified using nested-PCR and sequencing. False-negative controls consisted in microdissecting fresh-frozen tumor tissues both from a patient with a Li-Fraumeni syndrome who carried a p53 constitutional mutation, and from KRas mutated adenocarcinomas. To rule out false-positive results potentially generated by microdissection and nested-PCR, a phenacetin-associated urothelial carcinoma and normal fresh ureteral tissues (n = 4) were processed with high laser power. No unexpected results being identified, molecular analysis was pursued on malignant tissues, showing at least one mutation in all (six different mutations in two) patients, with 13/16 exonic (nonsense, 2; missense, 11) and 3/16 intronic (one splice site) mutations. They were distributed as transitions (n = 7) or transversions (n = 9), with an equal prevalence of A>T and G>T (3/16 each). While current results are in line with A>T prevalence previously reported in Balkan and Taiwan studies, they also demonstrate that multiple mutations in the TP53 hot spot region and a high frequency of G>T transversion appear as a complementary signature reflecting the toxicity of a cumulative dose of aristolochic acid ingested over a short period of time.

BACKGROUND Regorafenib is the first small-molecule multikinase inhibitor which showed survival benefits in pretreated metastatic colorectal cancer (mCRC) patients. Beside classical adverse events of this drug class, hepatotoxicity has been described as a frequent side effect. MATERIAL AND METHODS Patients with refractory mCRC treated with regorafenib in our institution were reviewed. Severe treatment-related liver toxicity was investigated. Clinical history, liver histology and genetic assessment (sequence analysis) of cytochrome P 3A4 (CYP3A4) and uridine diphosphate-glucuronosyltransferase 1A9 (UGT1A9) involved in regorafenib metabolization were here reported for patients with severe hepatotoxicity. RESULTS Among the 93 reviewed patients, 3 presented severe and icteric toxic hepatitis which was fatal for 1 patient. Histopathological liver lesions were different depending on the onset of hepatotoxicity (acute or subacute): acinar zone 3 necrosis in case of acute symptoms and portal tract inflammation with porto-central bridging and fibrosis in the delayed presentation. None patients had CYP3A4 gene mutations. Similar polymorphisms in UGT1A9 gene promoter region (UGT1A9 variant -118T9>10 (rs3832043)) were found in both patients who presented acute hepatitis. Moreover, it appears retrospectively that both of them already experienced significant toxicity under irinotecan-based chemotherapy. CONCLUSION This is the first report of severe hepatotoxicity with available liver histology and genetic assessment of enzymes involved in regorafenib metabolization. This report also reminds the importance of a close liver tests monitoring during regorafenib treatment. This article is protected by copyright. All rights reserved. Accepted ArticleThis article has been accepted for publication and undergone full peer review but has not been through the copyediting, typesetting, pagination and proofreading process, which may lead to differences between this version and the Version of Record. Please cite this article as doi: 10.1111/liv.13217 This article is protected by copyright. All rights reserved. Accepted ArticleThis article is protected by copyright. All rights reserved.Abstract: 221 words.

Purpose:To investigate the value of p53 functional analysis of separated alleles in yeast (FASAY) as a witness of p53/p21pathway alteration and as a predictor of recurrence in superficial transitional cell carcinomas. Experimental Design: p53 transcriptional activity was prospectively analyzed in 52 newly diagnosed transitional cell carcinoma using FASAY competent for the transactivation of p21 and bax promoters.TP53 and p21gene expression was quantified by real-time PCR, and expression of corresponding proteins was assessed by immunohistochemistry. In addition to tumor stage and grade, the predictive value of FASAY, real-time PCR, and immunohistochemistry for tumor recurrence was assessed by Cox survival analysis. Results: A total (p21 and bax) or partial (bax only) loss of transcriptional activity was observed in 15 of 52 (29%) and 4 of 52 (7.7%) cases, respectively, a partial loss being consistently associated with R283H mutation. p53 nuclear overexpression grossly overestimated (f40%) or underestimated (f10%) the true incidence of p53 transcriptional abnormalities, especially inT a -T 1 grade 1 to 2 tumors. Loss of p21 transactivation significantly correlated with decreased p21 gene expression and lack of expression of p21 (P = 0.001). FASAY had a better predictive value for recurrence than p53 immunohistochemistry (Cox hazard ratio, 6.57 versus 3.95; P = 0.0002 versus 0.019, respectively), whereas neither p21 immunohistochemistry (hazard ratio, 1.9; P = 0.29) norTP53 or p21 gene expression were significant predictors of recurrence. The prognostic difference between FASAY and p53 immunohistochemistry was maintained in the subgroup of T a -T 1 grade 3 tumors. Conclusions: FASAY is a valuable surrogate marker for assessing p53/p21 pathway alteration and predicts transitional cell carcinoma recurrence better than p53 immunohistochemistry.Transitional cell carcinoma (TCC) of the bladder is the fourth most common malignancy in men and the eight most common cancer in women. Eighty percent of TCCs are initially superficial (TMN97: T a -T 1 ) and easily treated by transurethral resection of bladder tumor. Despite initial complete resection of the tumor, recurrence occurs in 30% to 90% of the cases (1, 2). Progression to muscle invasive and/or metastatic stages requires radical and/or systemic therapies in 15% to 20% of the cases. To decrease recurrence and progression rates, intravesical administration of chemotherapy and/or Bacillus CalmetteGuerin is considered as standard therapy (2, 3). Because these therapies are not devoid of local and systemic toxicity, their use should better be guided by individual risks for recurrence and progression (4).Over the years, many attempts have been made to identify molecular markers that would supersede clinical features, such as tumor stage, grade, and multifocality (5, 6). Expression of TP53 gene has been one of the most frequently assessed genetic alterations in bladder TCC. In an overwhelming majority of studies, the prognostic significance of TP53 alterations in...

The predictive performance of the clinical model was only slightly improved by adding MGRS questioning the real clinical added value with regards to the cost of genetic testing and performance of current inexpensive clinical risk-calculators.