Jean Saric

Transcriptome classification of HCC is related to gene alterations and to new therapeutic targets

Hepatocellular carcinomas (HCCs) are a heterogeneous group of tumors that differ in risk factors and genetic alterations. We further investigated transcriptome‐genotype‐phenotype correlations in HCC. Global transcriptome analyses were performed on 57 HCCs and 3 hepatocellular adenomas and validated by quantitative RT‐PCR using 63 additional HCCs. We determined loss of heterozygosity, gene mutations, promoter methylation of CDH1 and CDKN2A, and HBV DNA copy number for each tumor. Unsupervised transcriptome analysis identified 6 robust subgroups of HCC (G1‐G6) associated with clinical and genetic characteristics. G1 tumors were associated with low copy number of HBV and overexpression of genes expressed in fetal liver and controlled by parental imprinting. G2 included HCCs infected with a high copy number of HBV and mutations in PIK3CA and TP53. In these first groups, we detected specific activation of the AKT pathway. G3 tumors were typified by mutation of TP53 and overexpression of genes controlling the cell cycle. G4 was a heterogeneous subgroup of tumors including TCF1‐mutated hepatocellular adenomas and carcinomas. G5 and G6 were strongly related to β‐catenin mutations that lead to Wnt pathway activation; in particular, G6 tumors were characterized by satellite nodules, higher activation of the Wnt pathway, and E‐cadherin underexpression. Conclusion: These results have furthered our understanding of the genetic diversity of human HCC and have provided specific identifiers for classifying tumors. In addition, our classification has potential therapeutic implications because 50% of the tumors were related to WNT or AKT pathway activation, which potentially could be targeted by specific inhibiting therapies. (HEPATOLOGY 2007;45:42–52.rpar;

Hepatocellular Adenoma Subtype Classification Using Molecular Markers and Immunohistochemistry

Hepatocellular adenomas (HCA) with activated β‐catenin present a high risk of malignant transformation. To permit robust routine diagnosis to allow for HCA subtype classification, we searched new useful markers. We analyzed the expression of candidate genes by quantitative reverse transcription polymerase chain reaction QRT‐PCR followed by immunohistochemistry to validate their specificity and sensitivity according to hepatocyte nuclear factor 1 alpha (HNF1α) and β‐catenin mutations as well as inflammatory phenotype. Quantitative RT‐PCR showed that FABP1 (liver fatty acid binding protein) and UGT2B7 were downregulated in HNF1α‐inactivated HCA (P ≤ 0.0002); GLUL (glutamine synthetase) and GPR49 overexpression were associated with β‐catenin–activating mutations (P ≤ 0.0005), and SAA2 (serum amyloid A2) and CRP (C‐reactive protein) were upregulated in inflammatory HCA (P = 0.0001). Immunohistochemistry validation confirmed that the absence of liver‐fatty acid binding protein (L‐FABP) expression rightly indicated HNF1α mutation (100% sensitivity and specificity), the combination of glutamine synthetase overexpression and nuclear β‐catenin staining were excellent predictors of β‐catenin–activating mutation (85% sensitivity, 100% specificity), and SAA hepatocytic staining was ideal to classify inflammatory HCA (91% sensitivity and specificity). Finally, a series of 93 HCA was unambiguously classified using our 4 validated immunohistochemical markers. Importantly, new associations were revealed for inflammatory HCA defined by SAA staining with frequent hemorrhages (P = 0.003), telangiectatic phenotype (P < 0.001), high body mass index, and alcohol intake (P ≤ 0.04). Previously described associations were confirmed and in particular the significant association between β‐catenin–activated HCA and hepatocellular carcinomas (HCC) at diagnosis or during follow‐up (P < 10−5). Conclusion: We refined HCA classification and its phenotypic correlations, providing a routine test to classify hepatocellular adenomas using simple and robust immunohistochemistry. (HEPATOLOGY 2007.)

Hepatocellular adenoma management and phenotypic classification: The Bordeaux experience

We took advantage of the reported genotype/phenotype classification to analyze our surgical series of hepatocellular adenoma (HCA). The series without specific known etiologies included 128 cases (116 women). The number of nodules varies from single, <5, and ≥5 in 78, 38, and 12 cases, respectively. The resection was complete in 95 cases. We identified 46 HNF1α‐inactivated HCAs (44 women), 63 inflammatory HCAs (IHCA, 53 women) of which nine were also β‐catenin–activated, and seven β‐catenin–activated HCAs (all women); six additional cases had no known phenotypic marker and six others could not be phenotypically analyzed. Twenty‐three of 128 HCAs showed bleeding. No differences were observed in solitary or multiple tumors in terms of hemorrhagic manifestations between groups. In contrast, differences were observed between the two main groups. Steatosis (tumor), microadenomas (resected specimen), and additional benign nodules were more frequently observed in HNF1α‐inactivated HCAs (P < 0.01) than in IHCAs. Body mass index > 25, peliosis (tumor), and steatosis in background liver were more frequent in IHCA (P < 0.01). After complete resection, new HCAs in the centimetric range were more frequently found during follow‐up (>1 year) in HNF1α‐inactivated HCA. After incomplete resection (HCA left in nonresected liver), the majority of HCA remained stable in the two main groups and even sometimes regressed. Six patients of 128 developed hepatocellular carcinoma (HCC) (all were β‐catenin–activated, whether inflammatory or not). Conclusion: There were noticeable clinical differences between HNF1α–inactivated HCA and IHCA; there was no increased risk of bleeding or HCC related to the number of HCAs; β‐catenin–activated HCAs are at higher risk of HCC. As a consequence, we believe that management of HCA needs to be adapted to the phenotype of these tumors. (HEPATOLOGY 2009.)

Hepatic Resection for Noncolorectal Nonendocrine Liver Metastases: Analysis of 1452 Patients and Development of a Prognostic Model

Objective:To determine the utility of hepatic resection (HR) in the treatment of patients with noncolorectal nonendocrine liver metastases (NCNELM). Summary Background Data:The place of HR in the treatment of NCNELM remains controversial, primarily due to the limitations of previously published reports and the heterogeneity of primary tumor sites and histologies. Methods:A multivariate risk model was developed by analyzing prognostic factors and long-term outcomes in 1452 patients with NCNELM treated with HR at 41 centers from 1983 to 2004. Results:Hepatic metastases were solitary in 56% and unilateral in 71% (mean diameter, 50.5 mm). Extrahepatic metastases were present in 22%. The most common primary sites were breast (32%), gastrointestinal (16%), and urologic (14%). The most common histologies were adenocarcinoma (60%), GIST/sarcoma (13.5%), and melanoma (13%). R0 resection was achieved in 83% of patients with a 60-day mortality rate of 2.3% and a major complication rate of 21.5%. Tumor recurred in 67% of patients (liver, 24%; extrahepatic, 18%; both, 25%). Overall and disease-free survivals at 5 years were 36% and 21% and at 10 years were 23% and 15%, respectively. In multivariate analysis, factors associated with poor prognosis were patient age >60 years, nonbreast origin, melanoma or squamous histology, disease-free interval <12 months, extrahepatic metastases, R2 resection, and major hepatectomy (all P ≤ 0.02). A prognostic model based on these factors effectively stratified patients into low-risk (0–3 points, 46% 5-year survival), mid-risk (4–6 points, 33% 5-year survival), and high-risk (>6 points, <10% 5-year survival) groups (P = 0.0001). Discussion:HR for NCNELM is safe and effective, with outcomes mainly dependent on primary tumor site and histology. For individual patients, a statistical model based on key prognostic factors could validate the indication for hepatic resection by predicting long-term survivals.

Possible involvement of pregnane X receptor-enhanced CYP24 expression in drug-induced osteomalacia.

Vitamin D controls calcium homeostasis and the development and maintenance of bones through vitamin D receptor activation. Prolonged therapy with rifampicin or phenobarbital has been shown to cause vitamin D deficiency or osteomalacia, particularly in patients with marginal vitamin D stores. However, the molecular mechanism of this process is unknown. Here we show that these drugs lead to the upregulation of 25-hydroxyvitamin D(3)-24-hydroxylase (CYP24) gene expression through the activation of the nuclear receptor pregnane X receptor (PXR; NR1I2). CYP24 is a mitochondrial enzyme responsible for inactivating vitamin D metabolites. CYP24 mRNA is upregulated in vivo in mice by pregnenolone 16alpha-carbonitrile and dexamethasone, 2 murine PXR agonists, and in vitro in human hepatocytes by rifampicin and hyperforin, 2 human PXR agonists. Moreover, rifampicin increased 24-hydroxylase activity in these cells, while, in vivo in mice, pregnenolone 16alpha-carbonitrile increased the plasma concentration of 24,25-dihydroxyvitamin D(3). Transfection of PXR in human embryonic kidney cells resulted in rifampicin-mediated induction of CYP24 mRNA. Analysis of the human CYP24 promoter showed that PXR transactivates the sequence between -326 and -142. We demonstrated that PXR binds to and transactivates the 2 proximal vitamin D-responsive elements of the human CYP24 promoter. These data suggest that xenobiotics and drugs can modulate CYP24 gene expression and alter vitamin D(3) hormonal activity and calcium homeostasis through the activation of PXR.

Preoperative Radiochemotherapy and Sphincter-Saving Resection for T3 Carcinomas of the Lower Third of the Rectum

ObjectiveTo evaluate the complications and oncologic and functional results of preoperative radiochemotherapy and sphincter-saving resection for T3 cancers of the lower third of the rectum. Summary Background DataCarcinomas of the lower third of the rectum (i.e., located at or below 6 cm from the anal verge) are usually treated by abdominoperineal resection, especially for T3 lesions. Few data are available evaluating concomitant chemotherapy with preoperative radiotherapy for increasing sphincter-saving resection in low rectal cancer. MethodsBetween 1995 and 1999, 43 patients underwent preoperative radiochemotherapy with conservative surgery for a low rectal tumor located a mean of 4.5 cm from the anal verge (range 2–6); 70% of the lesions were less than 2 cm from the anal sphincter. There were 40 T3 and 3 T4 tumors. Patients received preoperative radiotherapy with a mean dose of 50 Gy (range 40–54) and concomitant chemotherapy with 5-FU in continuous infusion (n = 36) or bolus (n = 7). Sphincter- saving resection was performed 6 weeks after treatment, in 25 patients by using intersphincteric resection. Coloanal anastomoses were associated with a colonic pouch in 86% of the patients, and all patients had a protecting stoma. ResultsThere were no deaths related to preoperative radiochemotherapy and surgery. Acute toxicity was mainly due to diarrhea, with 54% of grade 1 to 2. Four anastomotic fistulas and two pelvic hematomas occurred; all patients but one had closure of the stoma. Distal and radial surgical margins were respectively 23 ± 8 mm (range 10–40) and 8 ± 4 mm (range 1–20) and were negative in 98% of the patients. Downstaging (pT0–2N0) was observed in 42% of the patients (18/43) and was associated with a greater radial margin (10 vs. 6 mm;P = .02). After a median follow-up of 30 months, the rate of local recurrence was 2% (1/43), and four patients had distal metastases. Overall and disease-free survival rates were both 85% at 3 years. Functional results were good (Kirwan continence I, II) in 79% of the available patients (n = 37). They were slightly altered by intersphincteric resection (57 vs. 75% of perfect continence; NS) but were significantly improved by a colonic pouch (74 vs. 16%;P = .01). ConclusionsThese results suggest that preoperative radiochemotherapy allowed sphincter-saving resection to be performed with good local control and good functional results in patients with T3 low rectal cancers that would have required abdominoperineal resection in most instances.

Influence of postoperative morbidity on long-term survival following liver resection for colorectal metastases

Survival after resection of colorectal liver metastases may be influenced by the patient, the primary tumour and the liver metastases. Postoperative morbidity is associated with poor survival in several cancers. The aim of this retrospective study was to evaluate prognostic factors of survival after resection of colorectal liver metastases, including postoperative morbidity.

Laparoscopic intersphincteric resection with coloplasty and coloanal anastomosis for mid and low rectal cancer

The feasibility of laparoscopic rectal resection in patients with mid or low rectal cancer was studied prospectively with regard to quality of mesorectal excision, autonomic pelvic nerve preservation and anal sphincter preservation.

Comparison of Functional Results and Quality of Life Between Intersphincteric Resection and Conventional Coloanal Anastomosis for Low Rectal Cancer

PURPOSE:The technique of intersphincteric resection permits sphincter preservation with good oncologic results in very low rectal cancer. This study aimed to investigate functional results and quality of life after intersphincteric resection compared with conventional coloanal anastomoses.METHODS:From 1990 to 2000, 170 patients underwent total mesorectal excision with coloanal anastomosis for low rectal tumors. Questionnaires were obtained from 77 patients alive without colostomy: 37 had a conventional coloanal anastomosis and 40 had intersphincteric resection. Both groups were similar according to age, gender, anastomotic stenosis, colonic pouch, anastomotic leakage, preoperative radiotherapy, and follow-up (median, 56 months). Assessment included one functional and two quality-of-life questionnaires: the SF-36 Health Status and the Fecal Incontinence Quality of Life score.RESULTS:There was no difference in stool frequency, fragmentation, urgency, dyschesia, and alimentary restriction between patients with and without intersphincteric resection. Patients with intersphincteric resection had significantly worse continence (Wexner score, 10.8 vs. 6.9; P < 0.001) and needed more antidiarrheal drugs (60 vs. 35 percent; P = 0.04) than those without. Compared with conventional coloanal anastomoses, quality of life was altered by intersphincteric resection for the subscale embarrassment (P < 0.01) in the Fecal Incontinence Quality of Life score, whereas no difference of quality of life was observed with SF-36.CONCLUSIONS:Compared with conventional coloanal anastomoses, patients with intersphincteric resection have a higher risk of fecal incontinence and a slightly altered quality of life.

Intersphincteric resection with excision of internal anal sphincter for conservative treatment of very low rectal cancer

PURPOSE: Standard surgical treatment for low rectal cancer situated below 5 cm from the anal verge or at less than 1 cm from the anal ring is abdominoperineal resection. This is because of the necessity both to achieve a sufficient distal margin and to preserve the whole of the anal sphincter. The aim of this study was to evaluate morbidity, oncologic, and functional results of intersphincteric resection with excision of the internal anal sphincter and low coloanal anastomosis for carcinomas of the anorectal junction. METHODS: From January 1990 to December 1996, 16 patients were studied prospectively. All patients had an infiltrating adenocarcinoma (5 T2 and 11 T3), located between 2.5 and 4.5 (mean, 3.6) cm from the anal verge. Rectal resection with a minimum distal margin of 2 (mean, 2.4) cm was performed in all cases; six patients underwent partial resection of the internal sphincter, and ten patients had a subtotal resection. A colonic J-pouch was associated with coloanal anastomoses in eight cases. Twelve patients had preoperative radiotherapy, 3 with concomitant chemotherapy; 5 patients had postoperative chemotherapy. RESULTS: There was no post-operative mortality. Morbidity occurred in four patients, of whom two underwent permanent colostomy after pelvic hemorrhage or anovaginal fistula. After a median follow-up of 44 (range, 11–92) months, no local recurrence was observed, and two patients died of distal metastases. The five-year actuarial survival rate was 75 percent. Continence was normal in one-half of patients and was altered in the other patients who suffered from occasional minor leaks. The median resting pressure was lower after subtotal than after partial resection of the internal sphincter (40vs. 70 cm H2O;P=0.02), but functional results were similar in the two groups. CONCLUSION: These preliminary results suggest that intersphincteric resection can be an alternative to abdominoperineal resection for selected rectal tumors situated at the anorectal junction, without compromising chance of cure. Functional results and continence were not altered by subtotal resection of the internal anal sphincter.