Nicholas J. Petrelli

Aberrant CpG-island methylation has non-random and tumour-type-specific patterns

CpG islands frequently contain gene promoters or exons and are usually unmethylated in normal cells. Methylation of CpG islands is associated with delayed replication, condensed chromatin and inhibition of transcription initiation. The investigation of aberrant CpG-island methylation in human cancer has primarily taken a candidate gene approach, and has focused on less than 15 of the estimated 45,000 CpG islands in the genome. Here we report a global analysis of the methylation status of 1,184 unselected CpG islands in each of 98 primary human tumours using restriction landmark genomic scanning (RLGS). We estimate that an average of 600 CpG islands (range of 0 to 4,500) of the 45,000 in the genome were aberrantly methylated in the tumours, including early stage tumours. We identified patterns of CpG-island methylation that were shared within each tumour type, together with patterns and targets that displayed distinct tumour-type specificity. The expression of many of these genes was reactivated by experimental demethylation in cultured tumour cells. Thus, the methylation of particular subsets of CpG islands may have consequences for specific tumour types.

A prospective randomized trial of 5-fluorouracil versus 5-fluorouracil and high-dose leucovorin versus 5-fluorouracil and methotrexate in previously untreated patients with advanced colorectal carcinoma.

Seventy-four previously untreated patients with metastatic colorectal adenocarcinoma were prospectively randomized into one of three treatment regimens: (1) 5-fluorouracil (5-FU) 450 mg/m2 as an intravenous (IV) bolus daily for five days or toxicity, then 200 mg/m2 IV bolus every other day for six doses; (2) methotrexate (MTX) 50 mg/m2 in normal saline by IV infusion over four hours followed by an IV bolus of 5-FU 600 mg/m2. This was administered weekly for 4 weeks and then every 2 weeks. (3) Leucovorin 500 mg/m2 in a two-hour IV infusion of normal saline with 5-FU 600 mg/m2 as an IV bolus one hour after the Leucovorin began every week for 6 weeks. The combined complete and partial response rates in the three regimens were 11%, 5%, and 48%, respectively (P = .0009). The median duration of response in the 5-FU and Leucovorin regimen was 10 months. There was no statistically significant difference between the treatment regimens with respect to survival time (P = .6). Toxicity in the 5-FU and Leucovorin regimen was predominantly diarrhea (13 of 30 patients, 40%). In this regimen, eight of 13 patients (52%) who developed diarrhea not only required a dose reduction of 5-FU, but also hospitalization for IV hydration. The predominant toxicity in the 5-FU alone regimen and the 5-FU and MTX regimen was leukopenia. One drug-related death occurred in each regimen.

Age and sex are independent predictors of 5-fluorouracil toxicity: Analysis of a large scale phase III trial

Background. Cancer is most common in older age groups, but little information is available with regard to the impact of age on chemotherapy toxicity. This study was undertaken to determine if age is an independent risk factor for 5‐fluorouracil (5‐FU) toxicity.

Hamartomatous polyposis syndromes: Molecular genetics, neoplastic risk, and surveillance recommendations

Hamartomatous polyposis syndromes are characterized by an overgrowth of cells or tissues native to the area in which they normally occur. Juvenile polyposis syndrome (JPS) results from germ-line mutations in the SMAD-4 gene (18q21.1) that encodes for an enzyme involved in transforming growth factor beta(TGF-β) signal transduction. The increased neoplastic risk may result from SMAD-4 mutations in the stromal component, which stimulate epithelial dysplasia and progression to invasive malignancy. Peutz-Jeghers syndrome (PJS) is associated with germ-line mutations in the LKB1 gene (19p13.3) that encodes a multifunctional serine-threonine kinase. These mutations occur in the epithelial component, suggesting a direct tumor suppressor effect. Patients are at an increased risk of intestinal and extraintestinal malignancies, including breast, pancreatic, ovarian, testicular, and cervical cancer. Cowden’s disease is associated with germ-line mutations in the PTEN gene (10q22–23) and an increased risk of breast and thyroid malignancies. Ruvalcaba-Myhre-Smith syndrome is less common; controversy suggests that it may represent a variant of Cowden’s disease.

Sentinel node skills verification and surgeon performance: Data from a multicenter clinical trial for early-stage breast cancer

Objective:Marked variations in sentinel lymph node dissection (SLND) technique have been identified, and definitive qualifications for SLND performance remain controversial. Based on previous reports and expert opinion, we predicted that 20 to 30 cases of SLND with axillary lymph node dissection (ALND) would enable surgeons to identify sentinel lymph nodes (SLN). Summary Background Data:In 1999, the American College of Surgeons Oncology Group initiated a prospective trial, Z0010, to evaluate micrometastatic disease in the SLN and bone marrow of women with early-stage breast cancer. Eligible patients included women with biopsy-proven T1/T2 breast cancer and clinically negative lymph nodes who were candidates for lumpectomy and SLND. Methods:Participating surgeons were required to document 20 to 30 SLNDs followed by immediate ALND with failure rates less than 15%. Prior fellowship or residency training in SLND provided exemption from skill requirements. Data for 5237 subjects and 198 surgeons were available for analysis. Results:Surgeons from academic (48.4%), community (28.6%), or teaching-affiliated (19.8%) institutions qualified with 30 SLND + ALND cases (64.6%), 20 cases (22.2%), or exemption (13.1%). Participants used blue dye + radiocolloid in 79.4%, blue dye alone in 14.8%, and radiocolloid alone in 5.7% of cases, achieving a 98.7% SLN identification rate. Patient factors associated with increased SLND failure included increased body mass index and age, whereas tumor location, stage, and histology, presence of nodal metastases, and number of positive nodes were not. Surgeon accrual of fewer than 50 patients was associated with increased SLND failure; however, SLND technique, specific skill qualification, and institution type were not. Conclusions:Using a standard skill requirement, surgeons from a variety of institutions achieved an acceptably low SLND failure rate in the setting of a large multicenter trial, validating the incorporation of SLND into clinical practice.

Biofunctionalization of electrospun PCL-based scaffolds with perlecan domain IV peptide to create a 3-D pharmacokinetic cancer model

Because prostate cancer cells metastasize to bone and exhibit osteoblastic features (osteomimicry), the interrelationships between bone-specific microenvironment and prostate cancer cells at sites of bone metastasis are critical to disease progression. In this work the bone marrow microenvironment in vitro was recreated both by tailoring scaffolds physical properties and by functionalizing electrospun polymer fibers with a bioactive peptide derived from domain IV of perlecan heparan sulfate proteoglycan. Electrospun poly (epsilon-caprolactone) (PCL) fibers and PCL/gelatin composite scaffolds were modified covalently with perlecan domain IV (PlnDIV) peptide. The expression of tight junction protein (E-cadherin) and focal adhesion kinase (FAK) phosphorylation on tyrosine 397 also were investigated. The described bioactive motif significantly enhanced adherence and infiltration of the metastatic prostate cancer cells on all modified electrospun substrates by day 5 post-seeding. Cells cultured on PlnDIV-modified matrices organized stress fibers and increased proliferation at statistically significant rates. Additional findings suggest that presence of PlnDIV peptide in the matrix reduced expression of tight junction protein and binding to PlnDIV peptide was accompanied by increased focal adhesion kinase (FAK) phosphorylation on tyrosine 397. We conclude that PlnDIV peptide supports key signaling events leading to proliferation, survival, and migration of C4-2B cancer cells; hence its incorporation into electrospun matrix is a key improvement to create a successful three-dimensional (3-D) pharmacokinetic cancer model.

Photodynamic therapy in patients with colorectal cancer

A pilot study was conducted, in which photodynamic therapy (PDT), a technique in which malignant cells are destroyed by light after being previously photosensitized by a chemical compound, was tried in a group of 14 patients with recurrent or residual colorectal cancer in the pelvis. Three of the six patients with unresectable pelvic recurrences experienced a significant relief of pain after PDT. In two of the five patients who had an incomplete resection of their pelvic recurrences, there was also a substantial relief of pelvic pain after surgery and PDT. In one of these patients subsequent biopsies proved the disappearance of the residual pelvic microscopic disease after several sessions of PDT. Three patients had a recurrence from a squamous cell carcinoma primary of the anal canal. All recurrences were amenable to surgical resection. In one of the patients, PDT was used in an attempt to sterilize an area of residual tumor that was located over the left ischial tuberosity. The patient experienced good relief of pain, but died of her disease 7 months after PDT. In the other two patients, PDT was used as an „adjunct”︁ after resection of their recurrences. One of these patients was free of disease and died of an unrelated cause 12 months after PDT. The other is alive and well. This study demonstrated that PDT can be safe and tolerable in patients with pelvic malignancies. PDT is capable of tumor destruction, can be used repeatedly in areas previously exposed to ionizing radiation, and may have a role in the prevention and management of pelvic‐perineal recurrences from colorectal cancer.

Postoperative complications after splenectomy for hematologic malignancies

OBJECTIVE The authors analyzed the frequency and character of postoperative complications after splenectomy in patients with hematologic malignancies, and correlated these findings with preoperative conditions that could have predicted their outcome. SUMMARY BACKGROUND DATA Splenectomy is performed for hematologic malignancies for diagnostic and therapeutic indications. The role of splenectomy for lymphoproliferative and myeloproliferative malignancies is complex and sometimes controversial. METHODS The medical records of 135 patients undergoing splenectomies for hematologic malignancies at Roswell Park Cancer Institute from January 1, 1984 to December 31, 1993 were reviewed retrospectively. These included Hodgkins disease (HD), hairy cell leukemia (HCL), non-Hodgkins lymphoma (NHL), chronic lymphocytic leukemia (CLL), chronic myelogenous leukemia (CML), and a miscellaneous group. RESULTS The overall postoperative complication and mortality rates for all patients were 52% and 9%, respectively. The complication rate was 63% for patients whose spleens weighed greater than 2000 g, and 29% for patients whose spleens weighed less than 2000 g (p = 0.001). Seventy-three percent of the postoperative deaths were due to septic complications, only one of which was caused by an encapsulated organism. Complications occurred in less than 20% of patients with the diagnosis of HD and HCL; more than 50% of patients with NHL, CLL, and CML suffered postoperative complications. CONCLUSIONS Splenectomy performed in patients with hematologic malignancies is a potentially morbid procedure. Splenic size was the only preoperative factor found to be predictive of postoperative complications. The complication rate differed significantly between the different diagnostic subgroups.

Colonocyte differentiation is associated with increased expression and altered distribution of protein kinase C isozymes.

BACKGROUND & AIMS Colon cancer cells express reduced levels of protein kinase C (PKC). This study examines the regulation of PKC isozymes in normal colonic epithelium, as a basis for understanding the significance of alterations in this enzyme system in colon carcinogenesis. METHODS The expression and localization of PKC isozymes in mouse and rat colonocytes at different developmental stages were determined using a combined morphological and biochemical approach. PKC alpha expression was compared in colonic adenocarcinomas and adjacent normal mucosa by immunoblot analysis. RESULTS Mouse and rat colonocytes express PKC alpha, beta II, delta, epsilon, and zeta. Relatively low levels of these isozymes were detected in proliferating cells of the crypt base, predominantly in the cytosolic compartment. Coincident with colonocyte growth arrest/differentiation, PKC isozyme expression markedly increased in both the cytosolic and, more significantly, in the membrane/cytoskeletal fraction. Colonic tumors express reduced levels of PKC alpha, an isozyme that has been implicated in negative control of intestinal cell growth. CONCLUSIONS These findings are supportive of a role for certain PKC isozyme(s) in signaling pathways mediating postmitotic events in colonocytes in situ, and suggest that diminished activity of these pathway(s) may contribute to the alterations in growth control/differentiation associated with colonic neoplasia.

Genomic instability in sporadic colorectal cancer quantitated by inter-simple sequence repeat PCR analysis

Genomic instability plays a major role in cancer by facilitating tumor progression and tumor heterogeneity. Inter‐simple sequence repeat (inter‐SSR) PCR has been developed to provide a rapid and reproducible technique for quantitation of the major type of genomic instability observed in sporadic tumors, namely, that manifesting itself as amplifications, deletions, translocations, and insertions. Evaluation of 59 sporadic colorectal cancers by inter‐SSR PCR has demonstrated a wide range of instability, independent of tumor stage at diagnosis. Comparison of these data and the results of microsatellite PCR analysis reveals an association of high genomic instability with loss of heterozygosity but no association with the replication error phenomenon arising from defects in mismatch repair. Genes Chromosom. Cancer 18:19–29, 1997.