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The aim of this study was to provide an up-to-date review of the knowledge on the role of the main growth factors involved in the onset and progression of epithelial ovarian cancer (Transforming Growth Factor a-TGFa, Trans-forming Growth Factor-TGFb, Epidermal Growth Factor-EGF, Insulin-Like Growth Factor-IGF, Vascular Endothelial Growth Factor-VEGF). Relevant articles published between 1991 and 2001 were identified using the Medline database. Publications identified by the search were reviewed and critically evaluated for their relevance to growth factors role in ovarian cancer. This review may be useful for clinicians wishing to study the biological mechanisms involved in epithelial ovarian neoplasms, in order to evaluate the possible value of Growth Factors as prognostic or predictive markers which could lead to novel therapeutic regimens, fitting individual needs based on single biological variations.

Many members of the human kallikrein gene family were found to be differentially expressed in various malignancies and some are useful cancer diagnostic/prognostic markers. KLK9 is a newly discovered human kallikrein gene that is expressed in several tissues including thymus, testis, spinal cord, salivary gland, ovary, and skin. Like other kallikreins, the KLK9 gene was found to be regulated by steroid hormones in cancer cell lines. Our purpose is to examine whether quantitative analysis of KLK9 expression has prognostic value in ovarian cancer. We studied the expression of KLK9 by quantitative reverse transcription-PCR in 168 consecutive ovarian tumors of different stages, grades, and histological types, and correlated the expression with clinicopathological parameters, response to chemotherapy, and patients' survival. We found that KLK9 expression was significantly higher in patients with early disease stages (I or II; P = 0.044) and in patients with optimal debulking (P = 0.019). Kaplan-Meier survival curves demonstrated that patients with KLK9-positive tumors have substantially longer progression-free and overall survival (P < 0.001 and P = 0.016, respectively). When the Cox proportional hazard regression analysis was applied to subgroups of patients, KLK9 expression was found to be a significant predictor of progression-free survival in the subgroup of patients with low-grade tumors [hazard ratio (HR), 0.13; P = 0.0015], early stage (HR, 0.099; P = 0.031); and those with optimal debulking (HR, 0.26; P = 0.012). After adjusting for other known prognostic variables, KLK9 retained its independent prognostic value in all of these subgroups of patients. A negative correlation was found between the expression levels of CA125 and KLK9 (rs, 0.350; P = 0.002). Our results indicate that KLK9 is under steroid hormone regulation in ovarian and breast cancer cell lines. Immmunohistochemically, human kallikrein protein (hK9) was localized in the cytoplasm, but not in the nuclei, of the epithelial cells of ovarian cancer tissues. We conclude that KLK9 is a potential new independent favorable prognostic marker for early stage, low-grade, optimally debulked ovarian cancer patients.

Purpose: Kallikrein gene 4 (KLK4, also known as prostase/KLK-L1), located on chromosome 19q13.4, is one of the newly discovered members of the human KLK-like gene family. This gene is up-regulated by androgens in the LNCaP prostatic carcinoma cell line and by androgens and progestins in the BT-474 breast cancer cell line. On the basis of its apparent association with hormonally regulated tissues, we have undertaken to examine the prognostic value of KLK4 expression in 147 malignant ovarian tissues., Experimental Design: Tumors were pulverized, total RNA was extracted, and cDNA was prepared by reverse transcription. KLK4 was amplified by PCR using gene-specific primers, and its identity was verified by sequencing. Ovarian tissues were then classified as KLK4-positive or -negative, based on ethidium bromide visualization of the PCR product on agarose gels., Results: KLK4 was found to be expressed in 69 (55%) of 147 of ovarian cancer samples. We found a strong positive association between KLK4 expression and tumor grade (P = 0.02) and clinical stage (P < 0.001). Univariate survival analysis revealed that patients with ovarian tumors positive for KLK4 expression had an increased risk for relapse and death (P = 0.003 and 0.001, respectively). Whereas knowledge of KLK4 status did not significantly increase the prognostic power of the multivariate models, additional analyses did determine that KLK4 was an independent unfavorable prognostic factor in patients with grade 1 and 2 tumors., Conclusions: Our findings indicate that KLK4 expression is associated with more aggressive forms of ovarian cancer.

Purpose: Human kallikrein 10 (hK10; also known as the normal epithelial cell-specific 1 gene and protein) is a secreted serine protease, which belongs to the human kallikrein family. It has been reported that hK10 is down-regulated in breast and prostate cancer cell lines and that it may function as a tumor suppressor. Recently, we developed a highly sensitive and specific immunoassay for hK10 and found that this protein is abundantly expressed in ovarian tissue. In this study, we measured quantitatively hK10 levels in ovarian cancer cytosolic extracts and evaluated the prognostic value of this biomarker in ovarian cancer., Experimental Design: Specimens from eight normal ovarian tissues, eight ovarian tissues with benign disease, and 182 ovarian tumors were investigated., Results: hK10 concentration in ovarian tumor cytosols ranged from 0 to 84 ng/mg of total protein, with a median of 2.6. This median was highly elevated in comparison with normal and benign ovarian tissues (P < 0.001). A cutoff of 1.35 ng/mg was selected to categorize tumors as hK10 high and hK10 low. With chi(2) test and Fisher's exact test, high concentration hK10 was found to be associated with advanced disease stage, serous histological type, suboptimal debulking, and large residual tumor (>1 cm; all P < 0.05). hK10 status was additionally correlated with clinical outcome, including progression-free (PFS) and overall survival (OS) using the Cox model. In univariate analysis, we found that patients with hK10 high tumors were more likely to die and relapse, in comparison with patients with hK10 low tumors (hazards ratios for PFS and OS were 1.93 and 2.42, respectively; P < 0.05). Although this correlation disappeared after the entire patient population was subjected to multivariate analysis, it remained significant in the subgroup of patients with stage III/IV ovarian cancer (hazards ratios for PFS and OS were 1.98 and 2.12, respectively; P < 0.05)., Conclusions: Our results indicate that hK10 is a new, independent, unfavorable prognostic marker, especially for late-stage ovarian cancer.

Kallikreins (KLKs) belong to the serine protease family of proteolytic enzymes. Human pancreatic/renal KLK (KLK1) encodes for an enzyme that is involved in posttranslational processing of polypeptide precursors. The function of the other members of this gene family is currently unknown, but growing evidence suggests that many KLKs are implicated in carcinogenesis. By using the positional candidate approach, we were able to identify a new human KLK-like gene, KLK14 (also known as KLK-L6). This new gene maps to chromosome 19q13.3-q13.4 and is formed of seven exons (two untranslated and five coding exons) and six intervening introns. KLK14 was defined as a KLK gene based on structural and mapping criteria, in relation to other known KLK genes. KLK14 is expressed in a variety of tissues, but the highest levels of KLK14 are found in the central nervous system, including brain, cerebellum, and spinal cord. Our preliminary results show that KLK14 is down-regulated, at the mRNA level, in breast, testicular, prostatic, and ovarian cancer.

Unlabelled: KLK8 (neuropsin/ovasin) is a new member of the human kallikrein gene family, which consists of enzymes with serine protease enzymatic activity. Recent reports have implicated KLK8 in ovarian cancer. KLK8 may have potential clinical value for disease diagnosis or prognosis and it may also be a useful therapeutic target., Purpose: We undertook this study to evaluate the prognostic value of KLK8 in ovarian carcinoma by examining its expression in ovarian tumors., Experimental Design: The KLK8 gene was analyzed by reverse transcription-PCR and direct sequencing in several human normal tissues. Subsequently, its expression was studied in a set of ovarian tumors, and statistical analysis was performed., Results: We have identified two novel mRNA splice variants of the KLK8 gene, which are abundantly expressed in many tissues. These new variants were named KLK8 type 3 and type 4. Study of the expression of the KLK8 gene and its spliced variants in ovarian tumors indicated that the new variants were expressed very frequently and that full-length KLK8 expression is an independent and favorable prognostic marker for ovarian cancer. Patients with higher KLK8 expression in the tumor have lower grade disease, lower residual tumor left after surgery, live longer, and relapse less frequently. In multivariate analysis, higher KLK8 expression was significantly associated with longer disease-free survival., Conclusions: These results suggest that KLK8 is a novel, favorable prognostic marker in ovarian cancer. Because KLK8 encodes for a predicted secreted protein, its detection in serum may aid in ovarian cancer diagnosis.

Insulin-like growth factor binding protein-3 (IGFBP-3) regulates the mitogenic and anti-apoptotic actions of insulin-like growth factors (IGFs). To study the role of IGFBP-3 in ovarian cancer progression, we measured IGFBP-3 concentrations in tumour tissues from 147 patients with epithelial ovarian carcinoma and examined its associations with clinicopathological features of disease and patient survival. The average age of the patients was 54.6 years (range 25-88 years) and the median follow-up time was 37 months. IGFBP-3 levels were measured with a commercial immunoassay kit. Low IGFBP-3 levels were significantly associated with unfavourable prognostic features of the disease, including advanced stage (P=0.048), large size of residual tumour (P=0.007), and suboptimal debulking outcome (P=0.007). Low IGFBP-3 levels were also associated with a significantly increased risk for disease progression (RR=1.92; 95% confidence interval (CI) 1.05-3.45; P=0.034), but the association was not sustained when other clinical and pathological variables were adjusted for in the analysis. No significant associations were observed between the IGFBP-3 level and patients' overall survival and response to chemotherapy. Findings of the study indicate that IGFBP-3 may play a role in the progression of epithelial ovarian cancer, but that it has no independent value in predicting either disease prognosis or the response of patients to chemotherapy.

Unlabelled: The Anthracyclines/Taxanes combination is often used in adjuvant and advanced breast cancer., Purpose: To evaluate the toxicity and pathological response of sequential epidoxorubicin/paclitaxel combination as primary chemotherapy for T > 3 cm and T4 breast cancer patients., Patients and Methods: Forty-eight patients with T2 > 3 cm, T3 and T4 breast tumours were treated with Epidoxorubicin (90 mg/m2, i.v.) on day one and paclitaxel (200 mg/m2 over 3 hours) on day 2 every 21 days for four courses. After the fourth cycle the patients underwent modified radical mastectomy or quadrantectomy plus axillary lymph node dissection followed by six courses of intravenous CMF regimen (days 1 and 8, every 4 weeks). Radiotherapy was given to patients undergoing conservative surgery or with T4 cancers. Tamoxifen was administered in ER or PgR positive patients., Results: Out of the 48 patients enrolled into this trial, 43 were evaluable for toxicity and pathological response. Primary chemotherapy with epidoxorubicin and paclitaxel was well tolerated: no heart toxicity was observed during primary chemotherapy and follow-up. Primary toxicity consisted of myalgia, grade 1 neuropathy and grade 3 alopecia. Disappearance of invasive tumours in the breast with node negative was observed in 11.6% of patients: pathological partial response was shown in 56% of patients. On the whole major pathological response was achieved in 67% of our series: in the remaining 33% we found a stable disease or a size reduction less than 50%. No progressive disease was observed. Conservative surgery was performed in 64.5% of T2 and T3 tumours., Conclusion: These preliminary data showed that the epidoxorubicin/paclitaxel combination was safe and effective as primary chemotherapy for patients with T > 3 cm and T4 breast cancer patients.

The prognostic values of p53 and of its downstream mediator p21WAF1/Cip1 in patients receiving adjuvant chemotherapy for epithelial ovarian cancer have not been clearly established. Tumor extracts from a series of 120 patients treated postsurgically with cisplatin or carboplatin alone or together with other chemotherapeutics for primary ovarian carcinoma were assayed both for p53 protein by an immunofluorometric assay developed by us and for p21 protein by a commercially available immunoassay. Relative risks (RRs) for cancer relapse and death after 24 months of follow-up were determined by multivariate Cox regression analysis. Disease-free (DFS) and overall survival (OS) probabilities were also examined by the Kaplan-Meier method and log-rank tests. All other procedures were similarly nonparametric and based on two-sided tests of significance. Concentrations of p53 were elevated in patients with advanced stage disease (P = 0.02) or poorly differentiated (P = 0.03), suboptimally debulked tumors (P = 0.02), as well as in patients who failed to respond to chemotherapy (P = 0.03), as assessed by computed tomography scanning, serum CA125 determination, and second-look laparotomy. Statistically significant associations between concentrations of p53 and p21 were not found, nor were relationships demonstrated between concentrations of p21 and other clinicopathological variables or treatment response. Univariate analysis showed that p53 concentrations above the median indicated significantly higher risks for relapse (P = 0.04) and death (P < 0.01) and showed trends for increasing risks for relapse (P = 0.04) and death (P < 0.01) when p53 was considered as a four-level categorical variable. Multivariate analyses adjusted for age, stage, grade, and residual tumor size confirmed these observations (RR = 1.50; P = 0.05 for DFS and RR = 1.92; P = 0.03 for OS) for median-dichotomized p53, but the trends were of borderline significance (P = 0.09 for DFS and P = 0.07 for OS). In contrast, p21 positivity was not a significant predictor of favorable outcome in univariate survival analysis, and use of a three-level variable combining positivity or negativity status for both p53 and p21 did not yield greater separation of patients into risk groups (P = 0.07 for DFS and P = 0.06 for OS) than the use of p53 alone. Assessment of p53 expression may be an independent indicator of poor prognosis in ovarian cancer patients treated with adjuvant chemotherapy. The prognostic value of p21 expression, however, could not be demonstrated in our series of ovarian cancer patients.

Aim of this study was to provide a review of the basic mechanisms of drug resistance in ovarian cancer and novel strategies to modulate drug resistance. Relevant articles published through August 1999 were identified using the Medline data base. Publications identified by the search were reviewed and evaluated critically for their relevance to drug resistance in ovarian cancer. Ovarian cancer patients have high response rates to initial chemotherapy after cytoreductive surgery. However, most will develop resistance to chemotherapy during the course of their treatment. There are multiple mechanisms resulting in drug resistance. Strategies to modulate drug resistance include dose intensity, various pharmacologic agents, and gene therapy.

Background: Intrinsic and/or acquired chemoresistance is the major obstacle to overcome in the treatment of patients with ovarian carcinoma. The aim of the present study was to investigate the prognostic value of drug resistance associated proteins P-glycoprotein (P-gp), multidrug resistance related protein (Mrp1), canalicular multispecific organic anion trans-porter (c-MOAT or Mrp2) and lung resistance protein (Lrp) in ovarian carcinoma., Methods: Expression of P-gp, Mrp1, Mrp2 and Lrp was determined by immunohistochemistry of frozen tissue sections of 115 ovarian carcinoma patients and associated to clinico-pathological factors, response to chemotherapy and (progression free) survival., Results: Expression of P-gp was observed in 20 out of 115 (17%), Mrp1 in 51 out of 115 (44%), Mrp2 in 19 out of 115 (16%) and Lrp in 85 out 115 (74%) tumors. Expression of Mrp1 was related to Mrp2 (p < 0.0001) and P-gp (p < 0.001) expression, while Lrp expression was more frequently observed in patients with stage I/II versus stage III/IV tumors (p < 0.01), grade I/II versus III tumors (p < 0.05) and residual tumor < 2 cm versus > 2 cm after laparotomy (p < 0.05). Lower stage (p < 0.001), small residual tumor after first laparotomy (p < 0.001) and lower differentiation grade (p < 0.05) were related to longer (progression free) survival. P-gp, Mrp1, Mrp2, and Lrp expression was neither related to response to first line chemotherapy (59 evaluable patients) nor to (progression free) survival (all patients). On multivariate analysis only stage and residual tumor after first laparotomy were independent prognostic factors for (progression free) survival., Conclusions: In ovarian carcinoma Mrp1 expression is associated with Mrp2 and P-gp expression, while Lrp expression is associated with favorable clinicopathological characteristics. Assessment of P-gp, Mrp1, Mrp2 or Lrp does not allow prediction of response to chemotherapy or (progression free) survival in ovarian carcinoma.

Background: Epithelial ovarian cancer is the most lethal gynecologic neoplasia. Up to date, little is known about its biology, and this makes even more difficult the definition of new therapies and the finding of early diagnostic methods. In this study, the expression of two oncogenes, Ron and Met, whose role in cancer progression has already been shown, and the possible clinical implication of their presence in the neoplastic tissue have been evaluated., Methods: Forty-eight ovarian cancer specimens, 5 borderline lesions, 4 benign ovarian tumors and 2 normal ovaries were analyzed; from frozen tissue, Rna was extracted and cDna obtained by a RT-PCR (Retrotranscriptase-Polymerase Chain Reaction). Finally, the cDna was assayed for the presence of the Ron and the Met gene by another PCR. The results were correlated with clinicopathological parameters, and patient survival., Results: Ron expression was shown in 56% of malignant lesions, and in 60% of borderline ones, while Met expression was detected in 54 and 60%, respectively. No statistically significant correlation was found between Ron and Met expression and clinicopathological features, such as histotype, grading, staging, residual tumor after debulking surgery, and response to chemotherapy, while a strong correlation (p = 0.001) was observed between overexpression of one of the oncogenes and the concomitant expression of the other., Conclusions: Even if residual tumor after debulking surgery was the most relevant prognostic factor, this study showed new data about the concomitant expression of Ron and Met oncogenes, which may suggest their cooperative role in ovarian cancer progression.

Intrinsic and/or acquired resistance to chemotherapy is the major obstacle to overcome in the treatment of patients with ovarian carcinoma. The aim of the present study was to investigate the prognostic value of drug resistance-associated proteins P-glycoprotein (P-gp), multidrug resistance-associated protein 1 (MRP1), canalicular multispecific organic anion transporter (c-MOAT/MRP2), and lung resistance protein (LRP) in ovarian carcinoma. Expression of P-gp, MRP1, MRP2, and LRP was determined by immunohistochemistry of frozen tissue sections of 115 ovarian carcinoma patients and related to clinicopathological factors, response to chemotherapy, and progression-free survival. P-gp expression was observed in 20 of 115 (17%), MRP1 in 51 (44%), MRP2 in 19 (16%), and LRP in 85 (74%) tumors. Expression of MRP1 was related to MRP2 (P<0.0001) and P-gp (P<0.001) expression, whereas LRP expression was more frequently observed in patients with early stage (P<0.01), lower grade (P<0.05), and smaller residual tumor (P<0.05). Early stage (P<0.001), smaller residual tumor (P<0.001), and lower differentiation grade (P<0.05) were related to longer (progression-free) survival. P-gp, MRP1, MRP2, and LRP expression were neither related to response to first-line chemotherapy in 59 evaluable patients nor to progression-free survival in all patients. On multivariate analysis, only stage and residual tumor were independent prognostic factors for survival. In conclusion, in ovarian carcinoma, MRP1 expression is associated with MRP2 and P-gp expression, whereas LRP expression is associated with favorable clinicopathological characteristics. Assessment of P-gp, MRP1, MRP2, or LRP does not allow prediction of response to chemotherapy or survival in ovarian carcinoma.

p53 alteration, detected as mutation of the p53 gene or as accumulation of mutant p53 protein, is a common feature of most malignancies, including ovarian carcinoma, and may identify patients with unfavorable prognosis and resistance to chemotherapy. Tumor tissues from 55 patients with well or poorly differentiated (grades 1 or 3) primary epithelial ovarian carcinoma were assessed both for p53 protein overexpression by a sensitive time-resolved immunofluorometric assay employing DO-1 and CM-1 antibodies, and for genetic p53 abnormalities by direct sequencing of PCR-amplified exons 5 to 9. Sixteen p53 mutations (29%), including 3 deletions causing frameshifts as well as one nonsense and 12 missense point mutations were found in all exons except exon 9. Overexpression of p53 protein, defined as a concentration exceeding the 75th percentile, was found in 15 cases (27%), 10 of which had missense mutations (P < 0.01). Tumors with nonsense and frameshift mutations were p53-negative by immunoassay. Both p53 mutation (P = 0.04) and p53 protein accumulation (P < 0.01) were associated with stage III-IV disease, while p53 mutation was more closely related to grade 3 lesions (P = 0.04) and serous histotype (P = 0.01). These results indicate that p53 protein accumulation correlates well with missense point mutation in carcinoma of the ovary and, together with other evidence that p53 abnormality may be prognostic of outcome in this disease, suggest that the immunoassay of p53 protein may have clinical value.

The prognostic value of p53 protein accumulation in breast cancer, especially as detected by methods other than immunohistochemistry, has not been established unequivocally. A sensitive immunofluorometric assay of p53 protein employing DO-1 and CM-1 antibodies was used in this study to assay extracts of 171 breast carcinomas from northern Italy. p53 over-expression, demonstrated in 36 (21%) tumours, was associated with lack of oestrogen receptor (ER) expression but was not related to patient age, stage, lymph node status, tumour size, histologic type, grade or progesterone receptor (PR) expression status in contingency tables. An increased risk for cancer relapse of p53-positive patients compared to p53-negative patients was determined using multivariate Cox regression analysis, which also showed that p53 protein over-expression was an independent predictor of reduced disease-free survival in node-positive and ER+ patients but not in node-negative or ER- individuals. The equivalent analysis for assessing the impact of p53 status on overall survival was not statistically significant, possibly reflecting the short patient follow-up. Our results suggest that an immunoassay of p53 protein, applicable to cytosolic extracts prepared for steroid hormone receptor analyses, may provide information for breast cancer prognosis.

Klippel-Trenaunay-Weber syndrome (KTWS) is a rare, congenital disorder characterized by vascular nevus formation, deep venous thrombosis, varicosities, and hypertrophy of affected tissues. A patient with known thrombosis of his splanchnic circulation and visceral KTWS presented with life-threatening hemorrhage from rectosigmoid varices. Portosystemic shunting was not feasible. Endoscopic sclerosis, variceal ligation, and proctocolectomy were not possible due to the size and number of the varices. Previous treatment with epsilon-aminocaproic acid had been unsuccessful and complicated by thrombophlebitis. Conservative treatment with blood transfusions, cryoprecipitate, fresh frozen plasma, vitamin K, propanolol, and somatostatin analog failed to stop the bleeding. The patient was given the antifibrinolytic agent, tranexamic acid, with cessation of his hemorrhage. Serial thromboelastograms confirmed improved reaction time, coagulation time, clot formation rate, and maximum amplitude. We conclude that tranexamic acid may be a useful adjunct in the medical treatment of high-risk patients with KTWS and other vascular nevi complicated by coagulopathy.

Missense point mutations, leading to inactivation of the p53 tumor suppressor gene product, are currently the most frequent alterations in human cancer. Little, however, is known about small intragenic deletions or insertions occurring in this locus of chromosome 17. We have analyzed 56 primary ovarian tumors for the presence of such abnormalities. The analysis was based on multiplex PCR amplification of exons 1 through 11 of the p53 gene and fragment analysis of the generated PCR products. Mutations were detected in 14% (8 of 56) of the tumors. Deletions were much more prevalent than insertions (seven vs one). Six of the deletions and the insertion affected exon 5, and the other deletion was in exon 7. Two deletions and the insertion did not disrupt the reading frame; the protein product was expressed in the tumor at high concentrations in all three cases. The other five deletions generated a frameshift, which is predicted to result in the production of a truncated protein product. In the case of the deletions, a 2-5-bp repeat was present close to the detected deletion, whereas the insertion duplicated the sequence immediately upstream of the insertion site. Overall our findings indicate that small intragenic p53 deletions/insertions are not rare events in ovarian cancer, and that p53 exon 5 is the target in the vast majority (88%) of the cases.

The expression patterns of members of the transforming growth factor-beta (TGF-beta) family were analysed in 96 primary ovarian tumours by RNAse protection assay. mRNA for the three mammalian isoforms, TGF-beta 1, TGF-beta 2 and TGF-beta 3, was detected in 46, 66 and 66% of 74 malignant tumours, respectively, with the predominant patterns of expression being either dual or triple co-expression. TGF-beta II receptor expression, detected by reverse-transcription PCR, was present in 92% malignant tumours. Expression patterns were similar between malignant, borderline and benign tumours, although TGF-beta 1 incidence was reduced in benign tumours. In malignant tumours, the incidence of TGF-beta 1 expression was less than that of either TGF-beta 2 (P = 0.02) or TGF-beta 3 (P = 0.0014), while in both malignant and borderline tumours, TGF-beta 2 and TGF-beta 3 tended to be co-expressed. Aneuploid tumours were more likely than diploid tumours to express multiple rather than single forms of TGF-beta (P = 0.018). The incidence of TGF-beta 1 expression was reduced in PR-moderate/rich (PR > 20 fmol/mg protein) relative to PR-negative/poor tumours (P = 0.048), while TGF-beta 3 expression was increased in ER-moderate/rich (ER > 20 fmol/mg protein) tumours compared to ER-negative/poor tumours (P = 0.0012). Expression of TGF-beta 3, but not TGF-beta 1 or TGF-beta 2, was associated with advanced stage disease (P = 0.014) and, in the malignant group, reduced survival (P = 0.02) with a hazard ratio of 2.6. These data suggest a possible role for TGF-beta 3 in the progression of ovarian cancer.

Evidence of the involvement of cyclin genes in genetic alterations in human cancer is growing. In the present study, we investigated the amplification, in human breast and ovarian cancer, of 5 cyclin genes; cyclin A, cyclin D1, cyclin D2, cyclin D3 and cyclin E. For this purpose, a series of 1,171 breast and 237 ovarian tumors tested for DNA amplification by Southern blotting and a subset of 132 breast and 22 ovarian cancers were analyzed for RNA expression levels by slot-blot and Northern blotting. In breast tumors, only cyclin D1 was found to be activated in a sizeable fraction of the tumors (amplification 12.6%, overexpression 19%). Cyclin A, D2, D3, and E genes never, or only on rare occasions, showed increased DNA copy numbers and were never found overexpressed at the RNA level. Amplification of cyclin D1 correlated with ER+ breast cancer and the presence of lymph-node metastasis. Interestingly, we were also able to determine an association with invasive lobular carcinoma. Our data suggest that cyclin D1 activation determines the evolution of a particular subset of estrogen-responsive tumors. Data obtained in ovarian tumors contrasted with observations in breast cancer. Cyclin D1 DNA amplification was much less frequent in ovarian than in breast tumors (3.3% vs. 12.6%), whereas cyclin E amplification and overexpression were observed in a significant number of cases (12.5% and 18.0% respectively). Cyclin A, cyclin D2 and D3 rarely showed anomalies at the DNA level and were never overexpressed. No clear correlation could be observed between amplification of the cyclin E gene and tumor type, stage or grade in ovarian cancer. Data presented here suggest distinct pathways of cyclin activation in human breast and ovarian cancer.

Background: Pharmacologic intervention to minimize postbypass bleeding and blood product transfusions has received increasing attention for both medical and economic reasons., Methods: Two hundred ten patients were entered into a double-blinded, prospective, randomized study to receive either 10 g of fibrinolytic inhibitor tranexamic acid before incision (n = 104) or 250 mL of placebo saline solution (n = 106). All subjects requiring cardiopulmonary bypass were deemed suitable, including those having first-time coronary bypass grafting, valve replacement, and reoperation., Results: There were no statistically significant differences between the groups with respect to demographic or operative characteristics. The tranexamic acid group had a 48% reduction in 24-hour blood drainage (p < 0.001) and received 69% fewer total units of packed red blood cells, 83% fewer total units of plasma, and 75% fewer platelet transfusion units than controls. Only 13 of 104 tranexamic acid patients received blood products versus 33 of 106 controls (p < 0.001). The incidences of thrombotic complications, perioperative myocardial infarction, renal failure, and neurologic complications were not significantly different between the two groups. The tranexamic acid group had 0% mortality versus 1.9% for controls (not significant)., Conclusions: Tranexamic acid is safe and effective in reducing blood loss and blood use in a wide variety of cardiac surgical patients.

The regulatory subunits of protein kinase A, or cyclic AMP-binding proteins, were measured in a series of 107 human ovarian tumors (89 malignant, 7 borderline, and 11 benign tumors) and related to tumor clinicopathological features and patient survival. Total cyclic AMP-binding protein levels were not significantly different between malignant tumors and either borderline or benign tumors. However, serous tumors showed significantly higher levels of total cyclic AMP-binding proteins than other malignant tumors (P = 0.007). Poorly differentiated tumors also possessed significantly higher levels of binding proteins as compared with well/moderately differentiated tumors (P < 0.01). Retrospective analysis of follow-up data also revealed a significant trend for patients with high tumor cyclic AMP-binding proteins to have poorer survival (P = 0.03). Individual binding proteins were identified by photoaffinity labeling, and the RI (Mr 48,000) protein was expressed as a percentage of total cyclic AMP-binding proteins detected. The percentage of the RI protein was not significantly different among malignant, borderline, or benign pathologies and was not associated with tumor stage, differentiation, or debulk status. The percentage of RI was significantly increased in serous tumors compared to other common epithelial malignancies (P = 0.01). In malignant tumors there was a significant positive correlation between the percentage of the RI protein and total cyclic AMP-binding proteins (P = 0.01). These data indicate that high tumor levels of cyclic AMP-binding proteins are associated with serous histology, poor differentiation, and poor patient survival.

Creatine kinase BB isoenzyme (CK-BB) is overexpressed in many tumor tissues, including ovarian cancer. Using a highly sensitive and specific immunofluorometric method, CK-BB levels in 89 primary ovarian cancer cytosolic extracts have been measured and the associations between CK-BB and clinicopathological features of ovarian cancer have been studied. It was found that CK-BB levels are higher in endometrioid cell carcinomas. No clear association was established between CK-BB levels and patient age, menopausal status, clinical stage, histological grade or size of residual tumor. CK-BB was not associated significantly with either disease-free or overall survival of the patients. Based on these data, it was concluded that there is no prognostic value of CK-BB in ovarian cancer. Drugs that target CK-dependent energy metabolism of tumor cells may not be selective in ovarian cancer therapy.

Epithelial ovarian cancer probably occurs due to activation of several different combinations of genes, which produce cancers that vary biologically and clinically. We tested this hypothesis in 100 consecutive ovarian carcinomas by molecular biology techniques at the DNA and protein levels in three genes (erbB-2, myc, ras), which are frequently altered in this tumor system. Abnormally high expression of erbB-2 gene encoded p185 protein was observed in 31% of the samples, while erbB-2 gene amplification was detected by Southern analysis in 8%. ErbB-2 abnormal gene expression did not significantly affect the clinical outcome of patients, conferring a marginal worsening of survival. In 25 out of 96 (26%) tumor samples there was myc amplification. Higher levels of the ras-encoded p21 protein than in normal ovaries and benign ovarian tumors were found in 45% of the samples. Simultaneous overexpression of p185 and p21 was associated with shorter disease free (p = 0.02) and overall survival (p = 0.04) at significance levels notably higher than those observed for these oncoproteins singly. In addition, survival of patients with myc amplification and high p185/p21 coexpression was significantly worse (p < 0.05) than that of patients with normal levels. Our data suggest that concurrent abnormal gene expression may act synergistically to endow ovarian tumor cells with a highly aggressive phenotype. Evaluation of these genes may be helpful in the biological characterization of ovarian cancer and in defining individual patient prognosis.

Prostate-specific antigen (PSA) is thought to be produced exclusively by prostatic epithelial cells and is currently used as a tumor marker of prostatic adenocarcinoma. We recently found that 30% of breast cancers contain PSA immunoreactivity (IR-PSA). To examine the prognostic value of PSA in female breast cancer, we measured IR-PSA in tumor cytosols of 174 breast cancer patients and classified the breast cancers as either PSA positive or PSA negative based on an IR-PSA cutoff level of 0.03 ng/mg. IR-PSA was present in 27% of the patients. IR-PSA presence was associated with early disease stage, small tumors, and estrogen receptor-positive tumors. We used the Cox proportional hazards regression model to analyze survival of patients in association with PSA status and found that patients with IR-PSA-positive tumors had a reduced risk for relapse and death in univariate analysis (P = 0.02 and 0.06, respectively) and a reduced risk for relapse in multivariate analysis (P = 0.03). Further analysis indicated that the effect of IR-PSA on relapse-free survival was evident in node-positive or estrogen receptor-negative patients. Our study suggests that IR-PSA is an independent favorable prognostic marker for breast cancer and may be used to identify a subgroup of estrogen receptor-negative and/or node-positive patients who have good prognoses.

Background: It has been shown that the p53 gene is mutated in 30-80% of ovarian carcinomas and that the genetic alterations most often manifest as an accumulation of mutant p53 protein in tumor tissue. The prognostic significance of these findings for patients with ovarian cancer, however, must be established clearly., Methods: Mutant p53 protein in 90 consecutive epithelial ovarian carcinomas was quantitatively analyzed using a time-resolved immunofluorometric procedure. In contrast to immunohistochemical techniques, this method uses two anti-p53 antibodies. The Cox model was used to evaluate the strength of the associations between the prognostic markers and disease relapse or death at univariate and multivariate levels. Kaplan-Meier survival curves were calculated for patients who were p53-positive or negative and for subgroups with a different clinical stage, histologic grade, or residual postsurgical tumor., Results: The positivity rates for p53 included 1/21 (5%) with Stage I disease, 1/6 (17%) with Stage II, 29/51 (57%) with Stage III, and 8/12 (67%) with Stage IV (total = 39/90, 43%). Patients with p53-negative tumors had a significantly longer disease free survival than did patients with p53-positive tumors (P = 0.03); these results were similar for overall survival (P = 0.06). Multivariate analysis revealed that the presence of postsurgical residual tumor was the only predictor significantly associated with poor patient outcome. However, when patients were divided into groups based on histologic grade, patients with well (G1) and moderately (G2) differentiated tumors had a significantly higher risk of cancer relapse and death if mutant p53 protein was present in their tumors compared with patients who were negative for mutant p53 protein (< 0.01)., Conclusions: The immunofluorometric measurement of mutant p53 protein accumulation in epithelial ovarian carcinomas of a low histologic grade was associated significantly with an increased risk for cancer relapse and death. A similar trend also was suggested for early stage disease and in the absence of residual tumor after surgery. These increased risks, however, were not found for patients with high grade or advanced stage cancer or for those with residual tumor. To the authors' knowledge, this is the first report suggesting that p53 tumor protein accumulation is a marker of poor prognosis in a subset of patients with ovarian cancer.

The MET oncogene encodes the receptor for Hepatocyte Growth Factor/Scatter Factor, a unique growth factor that induces not only proliferation of epithelial cells, but also cell motility and invasiveness. DNA level and expression of the Met/HGF receptor gene were examined with Southern- and Western-blot analyses, respectively, in human ovary, benign ovarian tumors and epithelial ovarian carcinomas. The Met/HGF receptor was detectable in the surface epithelium of normal ovary. The level of expression was unchanged in benign ovarian tumors of various origins. Fourteen out of 67 malignant carcinomas (20%) showed a 3- to 10-fold increase in expression. In 5 additional cases the Met/HGF protein was overexpressed over 50-fold. This represents a total of 28% of cases. Overexpression was not associated with MET gene amplification. Overexpressing tumors belonged to different histotypic variants, but showed a well-differentiated phenotype. Clinically, overexpression was associated with disease at any pathologic stage, but was significantly correlated with premenopausal status of patients. These data suggest that expression of the Met/HGF receptor may add a selective growth advantage to a narrow subset of differentiated ovarian cancers in premenopausal patients.

Several chromosomal regions are found to be consistently amplified in human breast cancers. For two of these regions, 8p12 and 10q26, we previously reported the amplification of genes encoding FGF receptors, FGFRI/FLG and FGFR2/BEK, in about 12% of breast tumors. The PLAT gene, encoding the tissue-type plasminogen activator, is also located close to or within the 8p12 region. In the present study, we show that both FGFRI and PLAT can be amplified in breast as well as ovarian carcinomas. FGFRI amplification was detected in 14.5% of breast and 7.8% of ovarian tumors, whereas PLAT was found to be amplified in 15.6% and 19.4% of the tumors, respectively. Each gene could be amplified independently of the other. These data raised the question of which gene is selected for amplification at 8p12. In most cases, the levels of expression of FGFRI and PLAT in breast tumors were comparable to their level of expression in normal mammary tissue. However, FGFRI was expressed above the normal level in a certain number of cases. This gene could be a good candidate as "driver" of the 8p12 amplification, but it cannot account for all complex molecular events taking place in this region.

The present study was undertaken to compare the efficacy of ranitidine 150 mg/nocte with that of antacids taken "on demand" in the prevention of recurrence of duodenal ulcer (DU). Two groups (A, B) of patients with DU recently healed by ranitidine or antacids were studied. Patients of Group A (110) healed by ranitidine were put on maintenance treatment with ranitide, and patients of Group B (32) healed by antacids were given antacids "on demand" for a period of one year. Endoscopy was performed at the end of the 6th and 12th months, as well as whenever symptoms appeared. Patients were followed up clinically at 2-month intervals. Relapse rates of 11% (group A) and 34% (group B) were found at the end of the 6th month, (p less than 0.005), while the total relapse rates at the end of the 12th month were 19% and 47%, respectively (p less than 0.005). Recurrences were asymptomatic in 62% (Group A) and 27% (Group B) of patients. No significant side effects were observed. It is concluded that ranitidine 150 mg nocte is significantly superior to antacids "on demand" as far as the recurrence of DU is concerned.

In this study, 30 patients with metastatic breast cancer were treated with 5-Fluorouracil (5-FU) and high-dose Folinic acid, using a new sequential dosing schedule. Our treatment consisted of one day i.v. infusion of 500 mg/m2 of Folinic acid over two hours. One hour after the beginning of Folinic acid infusion, 5-FU (500 mg/m2) was given by i.v. bolus injection. The complete and partial response rates achieved (CR+PR) were 21% in a population of patients pretreated with chemotherapy including 5-FU. Cutaneous and bone metastasis responded best to our treatment. There were no treatment related deaths or withdrawals from the study. The drug related toxicities observed in this study were usually mild to moderate and easily controllable. Thus preliminary results of our study suggest that response rate, quality of life and time to disease progression for the responders improved by this sequential treatment.

Mammography is an effective method for finding lesions of the breast which are occult at clinical examination. For occult lesions biopsy, including as little surrounding tissue as possible, it is necessary that they be located before surgery in order to improve the pathological process and the cosmetic outcome for the patients. Among the clinically occult lesions shown only by mammography, the frequency of breast cancers ranges from 10% to 47%. There are several different techniques for locating hidden lesions of the breast. We have employed the insertion of a single rigid needle into the breast, with X-ray confirmation of correct positioning. Forty-nine patients underwent this technique in our Institution and in all cases we were able to achieve a correct insertion of the needle at the X-ray check (the tip of the needle was less than 1 cm from the lesion). The target lesion was removed (as confirmed by X-ray of the surgical specimen) in all cases at the first attempt. In our study we found 11 invasive and 7 in situ tumours (36%). No complications delaying the surgical biopsy or the recovery of the patients were observed.

The preoperative localisation of occult mammary lesions can be performed using fine needle biopsies, thus limiting esthetic damage in those patients in whom histological tests prove the lesion to be benign. Various methods have been proposed over the past 20 years. The purpose of this paper is to evaluate published data in order to determine which is the most appropriate method in terms of simplicity and efficacy.

Cyclic AMP (cAMP) analogues that selectively bind to either one of the two binding sites of cAMP-dependent protein kinase demonstrate a potent inhibition of the growth stimulated by estrogen in MCF-7 human breast-cancer cells in culture. The site-selective analogues, which are more potent activators of protein kinase than the analogues studied earlier, exhibit growth inhibition at micromolar concentrations. Among the analogues tested, 8-Cl-cAMP (Site I-selective) and N6-benzyl-cAMP (Site 2-selective) are the 2 most potent inhibitors, causing 40-70% inhibition of the estrogen-stimulated growth at 10-20 microM concentrations with no sign of toxicity. 8-Cl-cAMP (1 microM) in combination with N6-benzyl-cAMP (0.5 microM) almost completely blocks estrogen-stimulated growth, demonstrating synergism between the Site 1- and Site 2-selective analogues. The growth inhibition parallels an increase in the R11 cAMP receptor protein with a decrease in the R1 receptor as well as reduction of c-myc and c-ras oncoproteins, whereas growth inhibition by tamoxifen does not affect the levels of the cAMP receptor proteins or the c-myc and c-ras protein levels. Site-selective cAMP analogues are antagonistic to estrogen stimulation of breast-cancer cell growth through a mechanism different from that of tamoxifen.

Site-selective cyclic AMP analogs bind to site 1 or site 2 of the known cAMP-binding sites depending on the position of substituents on the purine ring, either at C-2 and C-8 (site 1) or at C-6 (site 2). The growth inhibitory effect of such site-selective cAMP analogs used in this investigation with 15 human cancer cell lines surpassed that of analogs previously tested. The most potent analogs were 8-chloro, N6-benzyl and N6-phenyl-8-p-chlorophenylthio-cAMP. The combination of a C-8 with an N6 analog had synergistic effects. The 24 site-selective analogs tested produced growth inhibition ranging from 30 to 80% at micromolar concentrations with no sign of toxic effects. Growth inhibition was not due to a block in a specific phase of the cell cycle but paralleled a change in cell morphology, an increase of the RII cAMP receptor protein and a decrease of p21 ras protein. Since the adenosine counterpart of the 8-chloro analog produced G1 synchronization without affecting the RII and p21 ras protein levels, it is unlikely that an adenosine metabolite is involved in the analog effect. Site-selective cAMP analogs thus provide a new biological tool for control of cancer growth.

A case of alpha-chain disease (alpha-CD) is described presenting as a malabsorption syndrome complicated by severe kalipenic nephropathy. Antibiotics and cyclophosphamide were effective in controlling the malabsorption and thr renal dysfunction but did not affect the basic pathological lesion. Eventually the disease became generalized, affected lymph nodes, liver and bone marrow and caused multiple intestinal perforations. Detailed histological studies failed to demonstrate the development of reticulum-cell sarcoma as a terminal complication of the disease and are suggestive of a single malignant plasmacytic cell process.

The physiologic role of cyclic adenosine monophosphate (cAMP) in the growth control of a spectrum of human cancer lines, including leukemic lines, and v-rasH oncogene-transformed NIH/3T3 cells is demonstrated by the use of site-selective cAMP analogs. These cAMP analogs, which can select either of the two known cAMP binding sites of the cAMP receptor protein, induce potent growth inhibition, phenotypic change, and differentiation (leukemic cells) of cancer cells at micromolar concentrations with no sign of cytotoxicity. The growth inhibition parallels selective modulation of cAMP-dependent protein kinase isozymes, type I versus type II, and suppression of cellular proto-oncogene expression. Site-selective cAMP analogs thus provide new biological tools for investigating cell proliferation and differentiation and also for the improved management of human cancers.

Site-selective cyclic AMP (cAMP) analogues inhibit growth and induce changes in morphology in a spectrum of human cancer cell lines (D. Katsaros et al., FEBS Lett., 223:97, 1987). The cellular events underlying such effects of cAMP analogues include differential regulation of type I versus type II cAMP-dependent protein kinase isozymes (S. Ally et al., Proc. Natl. Acad. Sci. USA, 85: 6319, 1988). Infusion (i.p.) of 8-Cl-cAMP, the most potent site-selective cAMP analogue, for 7 days produced regression of LX-1 lung carcinoma in athymic mice in a dose-dependent manner. The tumor regression correlated with the changing levels of cAMP receptor proteins, RI alpha and RII beta, the regulatory subunits of cAMP-dependent protein kinase type I and type II, respectively. By photoaffinity labeling with 8-N3-[32P]cAMP and immunoblotting with a monospecific anti-RII antibody, RI alpha (Mr 49,000) and RII beta (Mr 51,000) were identified in the untreated control tumors. 8-Cl-cAMP treatment induced a rapid increase of both RI alpha and RII beta in tumor cytosols and translocation (within 1 h) of only RII beta from the cytosol to the nucleus. RII beta in both cytosols and nuclei remained elevated during 8-Cl-cAMP treatment, whereas RI alpha in the cytosols gradually decreased with time of treatment after its initial transient increase. Northern blot analyses demonstrated that the RII beta mRNA level increased within 6 h of 8-Cl-cAMP treatment and remained elevated during treatment, whereas the RI alpha mRNA level decreased to below that of the untreated control tumor level after its transient increase during 1-6 h of treatment. 8-Cl-cAMP treatment also caused a sharp decrease in both N-ras and c-myc mRNA levels. These results suggest that the fundamental basis for the antineoplastic activity of 8-Cl-cAMP may reside in the restoration of normal gene regulation in neoplasms in which cAMP receptor proteins play a role.