Gianluca Paraluppi

The genomic landscape of response to EGFR blockade in colorectal cancer

Colorectal cancer is the third most common cancer worldwide, with 1.2 million patients diagnosed annually. In late-stage colorectal cancer, the most commonly used targeted therapies are the monoclonal antibodies cetuximab and panitumumab, which prevent epidermal growth factor receptor (EGFR) activation. Recent studies have identified alterations in KRAS and other genes as likely mechanisms of primary and secondary resistance to anti-EGFR antibody therapy. Despite these efforts, additional mechanisms of resistance to EGFR blockade are thought to be present in colorectal cancer and little is known about determinants of sensitivity to this therapy. To examine the effect of somatic genetic changes in colorectal cancer on response to anti-EGFR antibody therapy, here we perform complete exome sequence and copy number analyses of 129 patient-derived tumour grafts and targeted genomic analyses of 55 patient tumours, all of which were KRAS wild-type. We analysed the response of tumours to anti-EGFR antibody blockade in tumour graft models and in clinical settings and functionally linked therapeutic responses to mutational data. In addition to previously identified genes, we detected mutations in ERBB2, EGFR, FGFR1, PDGFRA, and MAP2K1 as potential mechanisms of primary resistance to this therapy. Novel alterations in the ectodomain of EGFR were identified in patients with acquired resistance to EGFR blockade. Amplifications and sequence changes in the tyrosine kinase receptor adaptor gene IRS2 were identified in tumours with increased sensitivity to anti-EGFR therapy. Therapeutic resistance to EGFR blockade could be overcome in tumour graft models through combinatorial therapies targeting actionable genes. These analyses provide a systematic approach to evaluating response to targeted therapies in human cancer, highlight new mechanisms of responsiveness to anti-EGFR therapies, and delineate new avenues for intervention in managing colorectal cancer.

Genetic and Expression Analysis of MET, MACC1, and HGF in Metastatic Colorectal Cancer: Response to Met Inhibition in Patient Xenografts and Pathologic Correlations

Purpose: We determined the gene copy numbers for MET, for its transcriptional activator MACC1 and for its ligand hepatocyte growth factor (HGF) in liver metastases from colorectal carcinoma (mCRC). We correlated copy numbers with mRNA levels and explored whether gain and/or overexpression of MET and MACC1 predict response to anti-Met therapies. Finally, we assessed whether their genomic or transcriptional deregulation correlates with pathologic and molecular parameters of aggressive disease. Experimental Design: One hundred three mCRCs were analyzed. Copy numbers and mRNA were determined by quantitative PCR (qPCR). Thirty nine samples were implanted and expanded in NOD (nonobese diabetic)/SCID (severe combined immunodeficient) mice to generate cohorts that were treated with the Met inhibitor JNJ-38877605. In silico analysis of MACC1 targets relied on genome-wide mapping of promoter regions and on expression data from two CRC datasets. Results: No focal, high-grade amplifications of MET, MACC1, or HGF were detected. Chromosome 7 polysomy and gain of the p-arm were observed in 21% and 8% of cases, respectively, and significantly correlated with higher expression of both Met and MACC1. Met inhibition in patient-derived xenografts did not modify tumor growth. Copy number gain and overexpression of MACC1 correlated with unfavorable pathologic features better than overexpression of Met. Bioinformatic analysis of putative MACC1 targets identified elements besides Met, whose overexpression cosegregated with aggressive forms of colorectal cancer. Conclusions: Experiments in patient-derived xenografts suggest that mCRCs do not rely on Met genomic gain and/or overexpression for growth. On the basis of pathologic correlations and bioinformatic analysis, MACC1 could contribute to CRC progression through mechanisms other than or additional to Met transcriptional upregulation. Clin Cancer Res; 17(10); 3146–56. ©2011 AACR.

Severe Course of Primary Hyperoxaluria and Renal Failure After Domino Hepatic Transplantation

We report herein a domino orthotopic liver transplantation (LT), from a 38‐year‐old woman undergoing liver‐kidney transplantation (LKT) for primary hyperoxaluria type I (PH1) to a recipient with cirrhosis and hepatocellular carcinoma. Delayed onset of PH1 and renal failure and 10% residual alanine‐glyoxylate aminotransferase (AGT) activity in domino liver justified its use for domino procedure. The clinical course after LKT was similar to that described in other series, including ours. Renal function started promptly and maintained despite sustained hyperoxaluria from dissolution of oxalotic deposits. Conversely, the domino recipient manifested severe hyperoxaluria and developed nephrolithiasis and renal insufficiency with rapid progression over 2 months. A new LT resulted in slow decrease of oxaluria and improvement of renal function. Therefore, PH1 behaved quite differently in these two patients, leading us to conclude that domino LT using livers from PH1 patients should be considered very carefully, only as a bridge to definitive LT in recipients with critical clinical conditions.

Intraoperative placement of transparietohepatic biliary drainage in remedial hepaticojejunostomy: technique and clinical experience

Remedial biliary surgery most often entails a Roux-en-Y hepaticojejunostomy. Sometimes the duct wall at the porta hepatis has been so damaged by inflammatory changes that the postoperative external drainage of bile away from a biliodigestive suture at risk of dehiscence is advisable. A technique of intraoperative placement of transparietohepatic biliary drainage was devised. The maneuver implies retrograde cannulation of a major intrahepatic duct with a vascular irrigation needle that is pushed to create the transhepatic path. Of 220 remedial hepaticojejunostomies performed in 211 patients (including 151 liver transplant recipients), the technique was applied in 49 (22%) of the most difficult cases in which the preoperative radiologic approach to the biliary tree had failed, was unsafe, or was unfeasible. The only major complication was a parenchymal tear needing perihepatic packing when the maneuver was performed too early after liver transplantation. Postoperative biliary fistula occurred in 2 patients (4%) and access to the biliary tract for percutaneous bilioplasty was provided in the short-term follow-up evaluation of 14 patients (29%).

Transthoracic open window hepatostomy: A salvage approach to right lobe abscesses after liver transplantation

Pyogenic abscesses in grafts after liver transplantation (LT) are observed in 0.5% to 1% of patients, with mortality rates up to 45%. Predisposing factors are hepatic artery thrombosis, presence of hepaticojejunostomy, biliary obstruction, ascendant cholangitis, bacteremia, and percutaneous procedures (liver biopsy and biliary drainage). Therapeutic options include antibiotics, immunosuppression withdrawal, percutaneous or surgical drainage, liver resection, and retransplantation. However, in extreme situations, treatments can fail and retransplantation can be contraindicated because of sepsis, a complex surgical history, or both. In this setting, a salvage approach to abscesses located in the right hepatic area was devised, inspired by the open window thoracostomy described by Clagett in 1963 for postpneumonectomy empyema. Similar to the Clagett procedure, the so-called open window hepatostomy (OWH) is based on transthoracic surgical drainage of the septic cavity and secondary wound healing. With over 1820 consecutive LT procedures performed from 1990 to 2008, the technique was applied in only 3 cases (0.2%), always resulting in control of infection and preservation of life. Repeated dressings and further interventions were needed without exception because an external biliary fistula invariably appeared. Complete skin closure was obtained in 2 cases in 12 years and 7 months, respectively. The most recent patient, whose case is illustrated here, is now at an advanced stage of the healing process. CASE REPORT

Disease-free survival after radical resection followed by adjuvant chemotherapy for primary hepatic synovial sarcoma

A 24-year-old gentleman with no significant previous medical history except smoking habit (15 cigarettes/day) was referred to our unit for a 7.5 cm S5–S8 liver lesion, discovered at ultrasound scan following an episode of fever associated with upper quadrant abdominal pain. Computed tomography (CT) confirmed an inhomogeneous S4–S5–S8 lesion presenting contrast enhancement during arterial phase and rapid wash-out at delayed phase, suggestive for hepatocellular carcinoma (Fig. 1a). At magnetic resonance imaging, the lesion was hyperintense in T2-weighed sequences, hypointense in T1-weighed sequences and presented a sharp signal increase at diffusion-weighed sequences (Fig. 1b). Magnetic resonance cholangiopancreatography showed that the lesion infiltrated right hepatic duct close to the confluence and was partially constituted by ectatic bile ducts (Fig. 1c). Alfa-fetoprotein and CA19.9 levels were normal. Upper gastro-intestinal endoscopy and colonoscopy were negative. He underwent right hepatectomy extended to S4 and had a normal postoperative course. Histological examination of resected specimen showed a poorly differentiated, highly cellular malignant neoplasm constituted prevalently by spindle-shaped cells, characterized by perineural and vascular invasion, large areas of necrosis, hemangiopericytomalike vascular pattern and high mitotic index. The lesion was surrounded by a pseudocapsule (Fig. 1d, e). Surgical resection margins were clear. Immunohistochemistry staining was diffusely positive for vimentin and CD99, focally positive for cytokeratin AE1/AE3 and epithelial membrane antigen (EMA), and negative for CD117, CD31, CLA, Chromogranin A, actin, S100, CD34, desmin and synaptophysin. These findings were in keeping with the diagnosis of poorly differentiated synovial sarcoma, grade 3 according to FNCLCC scoring system, which was confirmed by fluorescence in situ hybridization reaction using a Dual Color Breakapart probe (Abbott, IL, USA) showing typical SYT gene translocation (18q11.2) (Fig. 1f). Whole-body positron emission tomography/CT (PET/CT) obtained postoperatively ruled out distant disease leading to the final diagnosis of primary hepatic synovial sarcoma. The decision to administer adjuvant chemotherapy was carefully discussed with the patient and based on the high risk of recurrence according to the nomogram proposed by Canter et al. [1]. Patient received a total of five cycles of adjuvant chemotherapy with epirubicin and ifosfamide and was alive and recurrence free at 18-month follow-up. Synovial sarcoma is a type of soft tissue sarcoma, representing approximately 6–15% of all sarcomas of this group. Despite the adjective “synovial”, this tumor has no relation with joint tissue. It arises more frequently in the extremities but can arise at any site. There are three histological types of SS: monophasic, constituted by mesenchymal spindleshaped cells; biphasic, containing areas of epithelial and mesenchymal cells, and poorly differentiated. The vast majority of SS (95%) present typical t(X;18)(p11.2;q11.2) translocation, which is considered a hallmark of the disease [2]. Clinical behavior of SS is highly variable, ranging from relatively indolent disease to aggressive metastatic disease at onset. Long-term survival is also influenced by the tendency to recur late after surgical resection and 5-year * Renato Romagnoli renato.romagnoli@unito.it