Candidate gene isolation and comparative analysis of a commonly deleted segment of 7q22 implicated in myeloid malignancies.

Monosomy 7 and deletion of 7q are recurring abnormalities in malignant myeloid diseases. Here we extensively characterize an approximately 2-Mb commonly deleted segment (CDS) of 7q22 bounded by D7S1503 and D7S1841. Approximately 1.8 Mb of sequence have been generated from this interval, facilitating the construction of a transcript map that includes large numbers of genes and ESTs. The intron/exon organization of seven genes and expression patterns of three genes were determined, and leukemia samples were screened for mutations in five genes. We have used polymorphic markers from this region to examine leukemia cells for allelic loss within 7q22. Finally, we isolated mouse genomic clones orthologous to several of the characterized human genes. Fluorescence in situ hybridization studies using these clones indicate that a region of orthologous synteny lies on proximal mouse chromosome 5. These resources should greatly accelerate the pace of candidate gene discovery in this region.

Quantitative effects of Nf1 inactivation on in vivo hematopoiesis.

The NF1 tumor-suppressor gene is frequently inactivated in juvenile myelomonocytic leukemia, and Nf1 mutant mice model this myeloproliferative disorder (MPD). Competitive repopulation assays were performed to quantify the proliferative advantage of Nf1(-/-) hematopoietic cells in vivo. Nf1 mutant stem cells demonstrated a growth advantage that was greatest in myeloid lineage cells and least pronounced in T lymphocytes. Surprisingly, although low numbers of Nf1-deficient cells consistently outcompeted wild-type cells, levels of chimerism were stable over months of observation, and MPD was not observed unless threshold numbers of mutant cells were injected. These data showing that normal competitor cells can strongly modulate the growth of mutant populations in vivo have general implications for modeling cancer in the mouse. In particular, strains in which cancer-associated mutations are expressed in fields of target cells may not accurately model early events in tumorigenesis because they eliminate the requirement for a mutant clone to outcompete resident normal cells.

The hippurate ratio as an indicator of functional hepatic reserve for resection of hepatocellular carcinoma in cirrhotic patients.

Predicting the ability of the cirrhotic liver to withstand resection remains a challenge for the surgeon. This study evaluates the use of the hippurate ratio, a novel assessment of glycine conjugation of para-aminobenzoic acid by the liver, as a preoperative indicator of functional hepatic reserve. Between 1998 and 2000, sixty-one cirrhotic patients were prospectively assessed for hepatic resection using the hippurate ratio, indocyanine green retention at 15 minutes (ICG R-15), and other standard measures of liver function. Twenty-six patients were excluded as candidates for resection on the basis of inadequate functional hepatic reserve. Patients excluded from resection had significantly higher ICG R-15 values (29% +/- 9% vs. 16% +/- 12%, P = 0.001), higher Child-Pugh scores (5.9 +/- 0.9 vs. 5.3 +/- 0.4, P = 0.01), and lower hippurate ratios (30% +/- 14% vs. 45% +/- 15%, P = 0.005). There was a significant correlation between the hippurate ratio and ICG R-15. Other indicators of liver function such as factor V, factor VII, albumin, bilirubin, prothrombin time, and transaminases were no different between patients who did and those who did not undergo resection. Of the 35 patients resected, there were seven (20%) who developed varying degrees of liver failure with three perioperative deaths (8.5%). Patients who had some degree of liver failure had significantly lower hippurate ratios than patients who had no liver failure (29% +/- 10% vs. 48% +/- 14%, P = 0.002). There was no difference in ICG R-15 values between patients who had liver failure and those who did not. The hippurate ratio offers information on hepatocellular reserve that is not provided by other measures of liver function and may allow better selection of cirrhotic patients for liver resection.

Hepatic resection of noncolorectal nonneuroendocrine metastases.

Because hepatic resection is generally a safe procedure, the indications for resection of noncolorectal nonneuroendocrine (NCNNE) hepatic metastases have broadened. The prognostic features of NCNNE metastases treated surgically were reviewed to define better the value of resection. A retrospective review of patients undergoing liver resection for NCNNE metastases between 1978 and 1998 was undertaken. Thirty-seven patients were identified. Mean age was 56 years, with a median follow-up of 22 months. Primary tumor sites were grouped into gastrointestinal (GI) adenocarcinoma (small bowel, n = 4; pancreas, n = 2; esophagus, n = 1) and other (renal cell, n = 7; sarcoma, n = 7; melanoma, n = 5; adrenal, n = 3; unknown adenocarcinoma, n = 3; thyroid, n = 2; testicular, n = 1; ovarian, n = 1; breast, n = 1). All patients underwent surgery for cure. Metastases were synchronous in 14 patients. There was no surgical mortality. Overall 5-year survival rate was 45%. Five-year survival rates were better for patients with non-GI-origin metastases (60% v 0%; P =.01). Long-term survival was seen only in patients with non-GI-origin metastases. The extent of resection, presence of synchronous metastases, or disease-free interval from time of original disease to presentation with liver metastases were not predictive of outcome. We conclude that patients with NCNNE hepatic metastases can undergo liver resection with an expectation of prolonged survival. However, patients with liver metastases from GI primary tumors other than the colorectum are unlikely to show extended survival.

In vitro and in vivo effects of a farnesyltransferase inhibitor on Nf1-deficient hematopoietic cells.

Oncogenic RAS alleles encode proteins that accumulate in the guanosine triphosphate (GTP)-bound state. Because post-translational processing of Ras by farnesyltransferase is essential for biologic function, inhibitors of this enzyme have been developed as rational cancer therapeutics. We have investigated farnesyltransferase inhibitor (FTI) L-744,832 in an in vivo murine model of myeloid leukemia that is associated with inactivation of the Nf1 tumor suppressor gene. Nf1 encodes a GTPase activating protein for Ras, and Nf1-deficient (Nf1-/-) hematopoietic cells show hyperactive Ras signaling through the mitogen-activated protein (MAP) kinase pathway. L-744,832 inhibited H-Ras prenylation in cell lines and in primary hematopoietic cells and abrogated the in vitro growth of myeloid progenitor colonies in response to granulocyte-macrophage colony-stimulating factor (GM-CSF). This FTI also partially blocked GM-CSF-induced MAP kinase activation, but did not reduce constitutively elevated levels of MAP kinase activity in primary Nf1-/- cells. Injection of a single dose of 40 or 80 mg/kg of L-744, 832 increased the amount of unprocessed H-Ras in bone marrow cells, but had no detectable effect on N-Ras. Adoptive transfer of Nf1-/- hematopoietic cells into irradiated mice induces a myeloproliferative disorder that did not respond to L-744,832 treatment. We speculate that the lack of efficacy in this model is due to the resistance of N-Ras and K-Ras processing to inhibition by this FTI.

Myeloid malignancies induced by alkylating agents in Nf1 mice.

Therapy-related acute myeloid leukemia and myelodysplastic syndrome (t-AML and MDS) are severe late complications of treatment with genotoxic chemotherapeutic agents. Children with neurofibromatosis type 1 (NF1) are predisposed to malignant myeloid disorders that are associated with inactivation of the NF1 tumor suppressor gene in the leukemic clone. Recent clinical data suggest that NF1 might be also associated with an increased risk of t-AML after treatment with alkyating agents. To test this hypothesis, we administered cyclophosphamide or etoposide to cohorts of wild-type and heterozygous Nf1 knockout mice. Cyclophosphamide exposure cooperated strongly with heterozygous inactivation of Nf1 in myeloid leukemogenesis, while etoposide did not. Somatic loss of the normal Nf1 allele correlated with clinical disease and was more common in 129/Sv mice than in 129/Sv x C57BL/6 animals. Leukemic cells showing loss of heterozygosity at Nf1 retained a structural allele on each chromosome 11 homolog. These studies establish a novel in vivo model of alkylator-induced myeloid malignancy that will facilitate mechanistic and translational studies.

Surgical management of hepatocellular carcinoma: resection or transplantation?

Liver resection or transplantation offers the best opportunity for cure of hepatocellular carcinoma (HCC). To determine the relative roles for resection and transplantation and to evaluate the patient and tumor characteristics that might predict survival, the records of 125 patients treated for nonfibrolamellar HCC at The Toronto Hospital between 1981 and 1996 were reviewed. No adjuvant chemotherapy or antiviral protocols were used. Resection was the first operation in 67 patients; one underwent re-resection. Sixty patients underwent transplantation including two who had previously had a resection; 40 had known or suspected HCC and 20 had incidental tumors identified in the explanted liver. The incidence of cirrhosis was 49% for resection and 88% for transplantation. The incidence of hepatitis B virus (HBV) was 58% and 33%, respectively. The operative mortality rate for resection was 4.4% (9.4% in cirrhotic and 0 in noncirrhotic patients) and 13.3% for transplantation. The 5-year cumulative recurrence rate was 55% following resection and 20% following transplantation (P <0.001). The 5-year Kaplan-Meier survival rates were 38% for resection and 45% for transplantation-60% for transplanted HBV-negative and 17% for HBV-positive patients (P <0.001). After resection, recurrent HCC accounted for 86% of deaths, whereas recurrent HBV was responsible for 42% of deaths after transplantation. By univariate analysis, following resection, vascular invasion, advanced stage, multiple tumors, and lack of a capsule were predictive of survival; cirrhosis, HBV, age, tumor size, number, and grade were not. By multivariate analysis, only vascular invasion was predictive for resection and HBV for transplantation. Resection and transplantation are complementary methods of treating HCC. With the current organ shortage, resection should be considered first-line treatment. HBV-positive patients with HCC should only undergo transplantation in combination with effective antiviral therapy.

Aggressive surgical management of fibrolamellar hepatocellular carcinoma.

Fibrolamellar hepatocellular carcinoma (FLHC) is recognized as a distinct clinicopathologic variant of hepatocellular carcinoma. Ten consecutive patients with FLHC undergoing operative management at our institution were reviewed. At the initial presentation seven patients had stage II disease (pT2N0M0), whereas three patients were in stage III (pT2N0M0 or pT3N0M0). Initial procedures included formal right or left hepatectomy in four patients, right or left trisegmentectomy in two patients, left lateral segmentectomy or nonanatomic resection in three patients, and in one patient considered for liver transplantation, only exploration with biopsy of positive nodes was performed. Four stage II patients required a second procedure for resection of recurrent disease from 8 months to 6 years after the initial resection and one patient required a third procedure after 13 years. Reoperations included hepatic re-resection, resection of extrahepatic disease, and liver transplantation. Overall 5- and 10-year Kaplan-Meier survival was 70%. There were no deaths among stage II patients (follow-up 96 to 180 months). All stage III patients (i.e., lymph node involvement, vascular invasion, or multiple tumors) died within 5 years. Patients with stage II disease had better survival than patients with stage III disease (P = 0.011, log-rank test). Aggressive treatment of FLHC including reoperation and liver transplantation is justified, especially in patients with stage II disease.

A study of prognostic factors for hepatic resection for colorectal metastases.

BACKGROUND: Liver resection is accepted treatment for selected patients with colon cancer metastatic to the liver. There remains some controversy regarding the selection criteria, particularly which preoperative features are useful predictors of long survival postresection. METHODS: One hundred and twenty-three patients who had liver resection for colorectal metastases on the Hepato Pancreatic Biliary Service at The Toronto Hospital between August 1977 and June 1993 were studied. Seventy-seven had solitary lesions, 15 had single lesions with satellite nodules, and 31 had multiple lesions. Synchronous liver metastases were found in 40 patients and 83 patients had metachronous lesions. Fifty-one patients had formal lobectomies and 21 had extended lobectomies. RESULTS: Postoperative complications were seen in 28% of patients, but there were no operative or postoperative deaths. Overall actuarial 5-year survival was 34%. There was a significant difference in survival according to the number of metastases. Patients with single lesions had a 5-year survival of 47% compared with 16% for single lesions with satellite nodules, and 17% for multiple lesions. There were no significant differences in survival based on age, sex, synchronous versus metachronous lesions, status of lymph nodes at the time of original surgery, intraoperative blood replacement, or size of tumor. CONCLUSIONS: An aggressive approach to the surgical management of colorectal liver metastases is possible with low risk in centers specializing in liver surgery, and results in prolonged survival in one third of patients. The most reliable predictor of long-term survival is the number of metastases in the liver.

Presidential address 1996: "Dear Ministers of Health...." Canadian Association of General Surgeons.

The Canadian Association of General Surgeons, representing community and academic general surgeons throughout Canada, is concerned about the widespread effects of health care restructuring on clinical care, education and research. The general surgeon remains one of the critical members of the health care team and should have an adequate voice in ongoing discussions regarding health care reform.

Evaluation of liver transplantation for high-risk indications.

BACKGROUND: Appropriate use of orthotopic liver transplantation (OLT) requires continued assessment of the indications for transplantation as a number of diseases are associated with a poor prognosis. High-risk patients are those who have a poor survival or high incidence of recurrent disease (patients with tumours, hepatitis B- or hepatitis C-induced cirrhosis, fulminant hepatic failure or primary graft non-function). In addition, retransplantation may be associated with a poor outcome. METHODS: A retrospective review was made of the records of all adult patients undergoing OLT at this hospital between October 1985 and July 1994. RESULTS: A total of 396 liver transplants were performed in 364 patients. The 1- and 3-year actuarial survival rates were 81 and 69 per cent respectively. The overall survival rate of high-risk patients was significantly lower than that for all OLT recipients (P < 0.05). While no patients transplanted for hepatitis C have developed graft failure, recurrent hepatitis occurred in 15 of 35 patients. CONCLUSION: Strict selection criteria and appropriate perioperative investigations and interventions are required to improve the results of OLT in these high-risk patients.

ZBP-89, a Krüppel-like zinc finger protein, inhibits epidermal growth factor induction of the gastrin promoter.

We have shown previously that a GC-rich element (GGGGCGGGGTGGGGGG) conferring epidermal growth factor (EGF) responsiveness to the human gastrin promoter binds Sp1 and additional undefined complexes. A rat GH4 cell line expression library was screened by using a multimer of the gastrin EGF response element, and three overlapping cDNA clones were identified. The full-length rat cDNA encoded an 89-kDa zinc finger protein (ZBP-89) that was 89% identical to a 49-kDa human factor, ht(beta), that binds a GTGGG/CACCC element in T-cell receptor promoters. The conservation of amino acids between the zinc fingers indicates that ZBP-89 is a member of the C2H2 zinc finger family subclass typified by the Drosophila Krüppel protein. ZBP-89 is ubiquitously expressed in normal adult tissues. It binds specifically to the gastrin EGF response element and inhibits EGF induction of the gastrin promoter. Collectively, these results demonstrate that ZBP-89 functions as a repressor of basal and inducible expression of the gastrin gene.

The use of household surveys in community-oriented primary care health needs assessments.

OBJECTIVE: This study demonstrates the use of a community household survey and how it can provide information beyond that obtained with secondary epidemiologic data alone. METHODS: Adults in 215 randomly selected households in an inner-city neighborhood in Bronx, NY, were assessed by in-person interviews in homes of neighborhood residents. The survey collected data on self-reported health status, source and quality of medical care, possible barriers to obtaining medical care, and perceptions of the community. Demographic information was also obtained. Mortality rates, birth outcomes, and census data were obtained from secondary data sources. RESULTS: The overall health of the community members surveyed was poor, and rates of self-reported health status, mortality rates, and poor birth outcomes were all generally higher than city-wide rates. More than half the respondents were using hospital-based outpatient clinics or emergency room care as their primary source of medical care. Nearly half the respondents had no personal health provider, and most respondents could not obtain medical advice over the phone or be seen within a week. A number of barriers to obtaining medical care were found to be associated with either gender or ethnic group. These findings are being used as the basis for a number of community-oriented primary care (COPC) outreach efforts. CONCLUSIONS: Although household surveys are expensive to conduct, the information garnered from this survey could not be obtained from secondary data sources and was important in determining the direction of outreach and intervention programs being carried out in the community by the COPC clinic.

Nutritional status and function in patients following Whipple procedure compared with controls.

OBJECTIVE: Despite the potential for nutritional deficits in patients undergoing pancreaticoduodenectomy or Whipple procedure, long-term assessment of nutritional status has largely been ignored. This study assessed nutritional status of 24 Whipple patients compared with matched post-cholecystectomy controls. METHODS: Clinical assessment was by subjective global assessment, body composition was assessed by bioelectric impedance analysis and functional assessment was by respiratory muscle strength and skeletal muscle function performed by electrical stimulation of the ulnar nerve of the wrist and hand-grip dynamometry. RESULTS: Whipple patients studied 4.6+/-0.7 years since surgery and controls (4.8+/-0.7 years since surgery) were all judged clinically to be in a good nutritional state. Compared with controls, Whipple patients had significantly lower body weight (Whipple: 72.5+/-2.8 kg, control: 83.9+/-3.3 kg, p<0.05) however, the mean body weight of both Whipple and controls was above ideal weight (Whipple: 113.3+/-4.3%, control: 122.3+/-3.7% p = NS). No significant differences in functional performance were observed between groups. Energy intake of Whipple and controls was also comparable. In the Whipple group, neither the extent of gastric resection or the pathological diagnosis had an effect on the nutritional parameters studied. CONCLUSIONS: Long-term follow-up of patients having undergone Whipple procedure failed to reveal the presence of any nutritional or functional deficits suggesting that a full nutritional recovery is possible after this surgery.

Treatment of Budd-Chiari syndrome with portosystemic shunt or liver transplantation.

BACKGROUND: Budd-Chiari syndrome is an uncommon disorder caused by obstruction to hepatic venous outflow, causing varying degrees of hepatic injury depending on the extent, severity, and acuity of the obstruction. PATIENTS AND METHODS: We reviewed the indications for operative intervention and the results of treating 32 patients with Budd-Chiari syndrome seen at Toronto Hospital between 1968 and 1995. RESULTS: Twenty-one patients underwent portosystemic shunt (PSS) and 7 patients underwent liver transplantation (LT) as their initial operative management. Three patients who initially had PSS subsequently required LT. Patients with cirrhosis found on biopsy and preservation of hepatocellular function were treated with PSS and showed no difference in outcome when compared with patients without cirrhosis (P = 0.35). Patients who were treated by PSS with retrohepatic vena caval compression, as shown by high caval gradients had outcomes similar to those for patients with low gradients (P = 0.31). Using the Kaplan-Meier method, 5-year survival of PSS patients was 57%. Liver transplantation was used to manage patients with hepatic decompensation, as well as patients with vena caval occlusion or failed PSS. The 5-year Kaplan-Meier survival for LT was 67%. CONCLUSIONS: Both PSS and LT are effective options in the management of Budd-Chiari syndrome. Portosystemic shunt is the preferred initial approach even with cirrhosis or retrohepatic caval compression as long as there is preservation of liver function and a patent vena cava. Liver transplantation should be used as primary therapy for patients with irreversible hepatic decompensation or vena caval occlusion, and it can be an effective salvage procedure following failed PSS.

Rat phenol-preferring sulfotransferase genes (Stp and Stp2): localization to mouse chromosomes 7 and 17.

The phenol-preferring sulfotransferases aryl sulfotransferase IV and N-hydroxyarylamine sulfotransferase catalyze sulfate conjugation of N-hydroxy-2-acetylaminofluorene, a metabolite capable of causing hepatocarcinogenesis in rats. We utilized published cDNA sequences of these sulfotransferases to type the progeny of two multilocus crosses and determined that the genes, aryl sulfotransferase (Stp) and N-hydroxyarylamine sulfotransferase (Stp2), map to positions on mouse chromosomes 7 and 17.