Epidermal growth factor receptor (EGFR) is a potentially useful new biological prognostic and predictive indicator in human breast cancer. Additional research on EGFR is warranted to enhance our information on: i) the method of choice for its detection and quality control issues; ii) its association with novel pathobiological markers of prognosis; iii) its prognostic value in multivariate analysis; and iv) its capability to predict response to hormone therapy and, in the future, to biological treatments using antibodies against the specific receptor or its ligands.

The type I growth factor receptor family has been found to play an important role in the control of normal growth and differentiation. Moreover, the epidermal growth factor receptor and the c-erbB-2 oncogene seem to be implicated in the pathogenesis and behaviour of several cancers, including breast cancer. c-erbB-3 is a new member of the type I receptor family for which there is currently little information available on its expression in neoplastic tissues, and on its possible prognostic significance. This study was undertaken to define the prognostic value of c-erbB-3 expression in a series of node-negative breast cancer (NNBC) patients when compared, by multivariate analysis, with expression of the c-erbB-2 protein and conventional clinicopathological features. cerbB-3 was recognised by the novel monoclonal antibody RTJ1, whereas c-erbB-2 was detected by the polyclonal antibody 21N, using immunocytochemical methods. We found that overexpression of c-erbB-3 occurs frequently in NNBC. Overall, 138 of 212 carcinomas (65%) had some degree of membrane RTJ1 staining, and 28 (13%) showed strong and generalised positivity (+++). Twenty-four per cent of carcinomas had membrane 21N staining, and 12% presented strong and generalised positivity (+++). c-erbB-3 protein expression was significantly associated only with that of c-erbB-2 (P = 0.05), whereas 21N positivity was significantly associated with small tumour size (P = 0.02) and ductal histotype (P = 0.04). No significant correlation between expression of either receptor proteins or relapse-free survival was observed after a median follow-up of 63 months. Applying multivariate analysis, only tumour size approached significance. Our results indicate that analysis of expression of c-erbB-3 and c-erbB-2 alone do not seem to be useful in identifying patients with NNBC at different risk of relapse or death.

p53 expression detected by immunocytochemistry is emerging as a novel potentially useful prognostic indicator in breast carcinoma. However, additional research is warranted because a consensus has not yet been achieved on: i) methodology and quality control issues; ii) its association with other new biological prognostic indicators; iii) its prognostic value in multivariate analysis including conventional and new pathobiological features and; iv) its clinical usefulness either as a prognostic and predictive factor. This study was undertaken in a series of 165 early-stage breast cancer patients (median follow-up of 5 years) to compare the prognostic role of p53 expression with that of several other markers that have been found to be of value, using a multivariate statistical analysis. These factors are: tumour angiogenesis, epidermal growth factor receptor (EGFR), c-erbB-2 expression, cathepsin D, growth fraction by Ki-67 antibody, DNA ploidy and S-phase fraction. The main results observed were: i) 47 of 165 (28.5%) carcinomas had pAb 1801 staining and were considered as p53-positive; ii) p53 expression was weakly associated with S-phase fraction by flow cytometry (OR=1.86; p=0.085); iii) p53 expression was significantly associated with recurrence (p53 negative [-] versus weak positive [+] tumours: p=0.07 and odds ratio of 2.21; p53 negative [-] versus high positive [++] tumours: p=0.01 and odds ratio of 2.86) and death (p53-versus +: p=0.53 and odds ratio of 1.35; p53- versus ++: p=0.05 and odds ratio of 2.53); iv) the determination of p53 is able to identify a subset of high risk patients in c-erbB-2 negative tumours, this group being generally considered at good prognosis; v) In multivariate analysis on relapse-free survival including all the above markers only tumour angiogenesis, cathepsin D, EGFR and S-phase fraction and nodal status retained significance, and for overall survival only tumour angiogenesis was significant and independent. This new information on p53 expression could be useful to the clinician for a more rationale approach in defining prognosis of breast cancer patients. The prognostic value of p53 depends on which other markers are additionally analyzed and previous studies have not always assayed tumour angiogenesis, which is the most important factor in this series. p53 still need to be assessed as a potential predictor of response to chemo or radiotherapy, because of its role in monitoring DNA damage.

ONE HUNDRED EIGHTY patients affected by cerebral arteriovenous malformations (AVMs) underwent radiosurgical treatment in our department. One hundred fifty-three patients have been treated with complete irradiation of the entire AVM nidus. In 27 patients (with large and/or three-dimensional irregular target volumes), only part of the nidus was covered with a dose adequate for obliteration. Follow-up ranged from 88 to 1 months (mean, 43.1 mo). Angiographic control was performed at 12, 24, and 36 months until complete obliteration was attained. The complete obliteration rate was 46% at 1 year and 80% at 2 years. We observed 15 hemorrhages after treatment, and five patients died from them. No bleeding took place after complete angiographic obliteration. The aim of this study is to evaluate the effect of irradiation on bleeding risk after radiosurgery and before complete obliteration. Inclusive parameters of patients considered at risk were as follows: 1) all patients in the time lapse between irradiation and demonstrated complete angiographic obliteration; 2) all patients in the time lapse between irradiation and definitive treatment either by surgery or embolization; and 3) all patients in the time lapse between irradiation and death. These groups include all irradiated patients who still had incompletely obliterated AVMs. They were stratified starting from 0 time (the date of radiosurgery), and the hemorrhages were evaluated every 6 months. In totally irradiated cases, the bleeding risk decreased from 4.8% in the first 6 months after radiosurgery to 0% starting from the 12th month of the follow-up. In partially irradiated cases, the bleeding risk increased from 4% in the first 6 months to 12 to 10% from the 6th to the 18th month and decreased to 5.5% from the 18th to the 24th month; no bleeding was observed after the 24th month.

PURPOSE:To determine the absolute and relative value of microvessel density (MVD), p53 and c-erbB-2 protein expression, peritumoral lymphatic vessel invasion (PLVI), and conventional prognosticators in predicting relapse-free (RFS) and overall survival (OS) rates in patients with node-negative breast carcinoma (NNBC).PATIENTS AND METHODS:We monitored 254 consecutive patients with NNBC for a median of 62 months. Intratumoral MVD was measured after microvessels were immunostained using anti-CD31 antibody. p53 and c-erbB-2 protein and hormone receptors were also determined immunocytochemically. Results were analyzed by both univariate and multivariate statistical analysis.RESULTS:Univariate analysis showed that MVD was significantly predictive of both RFS (odds ratio [OR], 8.30; P = .0001) and OS (OR, 4.50; P = .012) when tested as a continuous or dichotomous variable. Likewise, tumor size (OR, 3.16; P = .0012), PLVI (OR, 4.36; P = .0009), estrogen receptor (ER) status (OR, 2.35; P = .016), progesterone receptor (PR) status (OR, 2.00; P = .017), and expression of p53 protein (OR, 2.82; P = .004) were significantly associated with RFS. Tumor size (OR, 3.80; P = .0038) and expression of p53 protein (OR, 2.58; P = .024) were significantly associated with OS by univariate analysis. Multivariate analysis showed that MVD (P = .0004), p53 protein expression (P = .0063), tumor size (P = .0144), and PLVI (P = .0033) were all significant and independent prognostic factors for RFS. However, only tumor size (P = .004) and MVD (P = .047) were independent predictors for OS. c-erbB2 expression was not associated with outcome by either univariate or multivariate analysis.CONCLUSION:MVD, p53 expression, PLVI, and tumor size are independent prognostic indicators of recurrence, which are useful in selection of high-risk NNBC patients who may be eligible to receive adjuvant therapies.

228 patients affected by cerebral arteriovenous malformations (AVMs) underwent linear accelerator radiosurgery. Follow-up ranges from 1 to 100 months (mean 42 months). Complete angiographic obliteration was achieved in 47% of treated patients at one year and 80% at 2 years. 17 haemorrhages were observed after treatment and 6 patients died from them. No bleeding took place after complete angiographic obliteration. 11 patients suffered for radionecrosis. In 6 patients complete recovery was obtained with corticoid medication. The aim of this study is to present our results and to evaluate the effect of irradiation on the risk of bleeding after radiosurgery. Patients were considered at risk in the time lapse after irradiation and before angiographic obliteration or other definitive treatment or death. Patients were followed from the date of radiosurgery and the number of haemorrhages were recorded every six months. In our series the bleeding risk in patients harbouring incompletely obliterated AVMs decreases from 8% in the first year after radiosurgery to 0% starting from the 24th month of the follow-up.

This study was undertaken to define the role of P-glycoprotein (P-glyc), as a marker of multidrug resistance (MDR) and to determine if it would predict both response to chemotherapy and prognosis in 42 previously untreated patients with advanced breast carcinoma. P-glyc was assayed using the monoclonal antibody (Mab) JSB-1 using an immunocytochemical method. On the same tumours, the following biological features were also estimated: tumour angiogenesis, epidermal growth factor receptor, c-erbB-2 oncoprotein and growth fraction using immunohistochemistry with immunocytochemically factor VIII-related antigen, EGFR1 Mab, 21N antibody and Ki-67 Mab, respectively. Overall, 50% of carcinomas were JSB-1 positive and the presence of P-glyc was not significantly associated with any other factor, but it was significantly predictive of poor response with anthracycline based chemotherapy (p=0.03). Moreover, lack of initial response to chemotherapy was significantly correlated with the percentage of cells staining with the JSB-1 antibody (p=0.016). P-glyc expression and the proportion of cells staining as JSB-1 positive correlated with clinical stage and analysis showed that they significantly predicted response to chemotherapy only in stage IV patients (p=0.015 and p<0.001, respectively) whereas in stage III patients there was only a weak and non significant association with response. None of the other factors were predictive for response or prognosis. The present study indicates that intrinsic resistance seems to have an important role in the failure of chemotherapy in advanced breast cancer and the determination of P-glyc could be of value to the clinician in identifying those patients (with P-glyc-negative tumours) who are potentially responsive to conventional chemotherapy.

The aim of this presentation is to analyse the mechanical and dosimetrical parameters of the linear accelerator used in stereotactic radiosurgery. The use of the thimble and Markus chambers, TL and film in small field dosimetry are investigated. To determine the optimal irradiation technique and dose distribution, the dose volume to healthy tissue is considered.

Squamous-cell carcinoma of the head and neck includes a heterogeneous group of tumours of the upper air and food passages for which prognosis is difficult to assess. In fact, patients in comparable stages may have diverse clinical courses and responses to similar treatments. In order to better define the prognosis of each patient there is therefore a need to identify novel biological markers which reflect more accurately growth rate, progression and metastatic potential of each tumour. We assessed whether metastases correlate with microvessel counts (i.e. intratumoral vascularity) using the CD-31 monoclonal antibody (MAb) and p53 mutant protein expression, determined in the primary by immunocytochemical methods in 70 patients with locally advanced head and neck cancer. Patients were treated with concurrent chemo-radiotherapy; 50 of these presented loco-regional node metastasis at diagnosis whereas 3 cases, initially node-negative, developed distant metastasis during the period of observation. No feature was predictive for objective response to treatment. The overall mean and median blood vessel density at "hot spots" was 37.42 and 36, respectively, and 57% of the tumours expressed p53 mutant proteins. These 2 biological markers were significantly associated. Patients with metastases (loco-regional and distant) had a significantly higher mean blood-vessel density than those without tumour spread. Also, patients with p53-positive (+/++) tumours had a significantly higher incidence of metastasis than those with negative ones. Multivariate analysis showed that both vascularity and stage, but not p53 expression, are significant and independent predictors of metastasis in this series.

The study was undertaken to determine the clinico-pathobiological characteristics in a series of 49 patients who developed metachronous breast carcinoma. Possible differences between the two tumours of conventional clinico-pathological features and of some biological markers such as DNA ploidy, c-erbB-2 oncoprotein overexpression and tumour angiogenesis were evaluated. The McNemar's test for independence showed that all the characteristics analyzed between the two tumours, in the same case, were not significantly different. After a median follow-up time of 69 months the overall survival of the series was 87.5% and the only significant prognostic factor for clinical outcome was peritumoural lymphatic vessel invasion (PLVI). The median second tumour-free interval was of 32 months ( 13 to 160 months) and none of the variables analyzed on the first primary was predictive of the timing of appearance of the second tumour. To assess the association between the characteristics of the first tumour and the odds of developing a metachronous carcinoma a case-control analysis was conducted. For each woman of the present series who developed bilateral cancer (case) a woman who had unilateral breast cancer (control) was matched for the length of the follow-up. A log-logistic regression model for matched sets was also performed to assess the risk of developing the second tumour. Applying multivariate analysis we found that progesterone receptor (PgR) status was the most important prognostic factor for the odds of bilateral tumour (odds ratio 0.22, p=0.013) followed by histological grade (odds ratio 0.20, p=0.063) and presence of PLVI (odds ratio 3.13, p=0.067). These findings suggest that the knowledge on the initial primary of PgR, grading and PLVI could be important to assess the individual risk of developing metachronous breast cancer. The determination of these factors could improve our ability to identify subsets of patients operated for breast cancer with different risks for bilateral tumour, allowing for a better selection of those patients who need intensive surveillance of their contralateral breast, and eligible for chemoprevention.

Several phase II studies have shown that concurrent chemotherapy and radiotherapy (RT) improves the response rates and locoregional control in patients with advanced, unresectable squamous-cell carcinoma of the head and neck (H&N). However, it is still unclear which is the drug of choice to be given with RT. We therefore conducted a randomized comparison of cisplatin (CDDP)-RT versus carboplatin (CRP)-RT in such patients. The two platinum compounds were given at equitoxic doses. The primary objective of the study was to compare the side effects and the response rates of the two regimens. Radiotherapy was given at conventional dosages, 2 Gy for 5 days every week for a total dose of 64 Gy with CDDP 80 mg/m2 or CRP 375 mg/m2 for three cycles on days 1, 21 and 42. At present 53 patients are entered in the study: 27 in the CDDP arm and 26 in the CRP. The two arms were balanced for all the pre-treatment characteristics. Both the schedules were well tolerated. However, incidence of nausea and vomiting (p=0.0045); anemia (p=0.032) and peripheral neuropathy (p=0.032) was significantly greater with CDDP-RT as compared to CRP-RT. On the other hand, CRP-RT gives a significantly higher incidence of stomatitis (p=0.0067) and a marginally worse thrombocytopenia (p=0.09). The complete response (CR) rates were similar in the two arms (55.5% in the CDDP-RT versus 61.5% in that CRP-RT, respectively) as well as the overall response rates (92.5% versus 84.5%, respectively; p=0.36). The estimated 1 -and 2 year overall and disease-free survival rates were not significantly different in the two arms. In both the groups logistic-regression models showed that those patients with a CR (p=0.017); stage III (p=0.011); smaller primary (p=0.025) and limited node-involvement (p<0.001) had a significant better survival. We conclude that concurrent chemo-radiotherapy is an effective and safe treatment for patients with locally advanced H&N cancer. The combination CRP-RT possess a similar activity but a different, and perhaps a more favorable, spectrum of toxicity when compared to the CDDP-RT therapy. Survival results need to be assessed in a larger series and followed for a more prolonged time.

The aim of this paper is to analyse the dosimetric parameters of a linear accelerator used in radiosurgery treatments. The influence of these parameters on the resulting dose distribution are basic for delivering the predefined dose to the vascular or oncological target volume. Several dosimetric methods have been used to define the output factors for small fields. The thimble and the Markus chambers, TLD and film dosimetry are investigated; all these dosimetric systems give reliable and almost similar results if used in the correct way. In the determination of tissue maximum ratio (TMR) the response curves obtained by ionometric and film dosimetry were investigated. For TMR determination the use of the Markus chamber and the correction factors to be applied as a result of the small dimension of the field were also investigated.

The incidence of breast cancer is increasing in all Western countries. Due both to a more widespread public education and to early diagnosis by mammography screening programs, the percentage of patients with node-negative breast cancer has gone up to 70%. Thus, node-negative breast cancer is a major public health problem and, consequently, clinical research in this setting is an expanding field. A recent overview analysis confirmed the results of five prospective randomized clinical trials suggesting that systemic adjuvant therapy can benefit node-negative breast cancer patients. Because of the heterogeneity of node-negative breast cancer, it is reasonable to attempt to avoid excessive treatment morbidity and costs by using selective prognostic markers to identify patients at high risk for disease recurrence who are eligible for postsurgical systemic adjuvant therapy. It is also desirable to use predictive markers in selecting the therapy to which each patient is more likely to respond. The need for additional prognostic and predictive factors has led to identification of a plethora of potentially useful markers. As a result, the selection of patients at different risks of developing node-negative breast cancer and the choice for appropriate therapy remain difficult and confusing for the clinician. Moreover, the majority of studies have examined new markers individually rather than by multivariate analysis and retrospectively rather than prospectively. Thus, there are also important methodologic biases in such studies. This analysis consists of (a) defining the clinical "problem," (b) defining the terms of prognostic and predictive factors, (c) suggesting more appropriate laboratory and clinical approaches to properly evaluate a new indicator, (d) identifying the subsets of patients in whom the use of new prognosticators is warranted and of particular importance, and (e) providing some direction for future research on this topic. Our ultimate goals are to facilitate the understanding of node-negative breast cancer prognostic markers among clinicians, to help them select the most appropriate indicator for specific situations, and to recommend methodology for future research.

PURPOSE:A study was undertaken to define the prognostic value of the expression of the c-erbB-2 oncoprotein in a series of breast cancer patients when compared by multivariate analysis with expression of the epidermal growth factor receptor (EGFR), DNA ploidy, and conventional clinicopathologic features.PATIENTS AND METHODS:Prognostic indicators were analyzed in 165 primary breast cancers. The c-erbB-2 oncoprotein was recognized by the polyclonal antibody 21N using an immunocytochemical method. Expression of the EGFR was stated immunocytochemically using the monoclonal antibody EGFR1. DNA ploidy was assessed in paraffin-embedded sections using a standard flow-cytometric method.RESULTS:Overall, 27% of carcinomas had membrane 21N-staining and were classified as c-erbB-2-positive. Overexpression of the c-erbB-2 oncoprotein was poorly associated with EGFR expression and the conventional pathologic features, and it was weakly associated with DNA ploidy and nodal status. Univariate analysis showed that c-erbB-2 expression, nodal status, DNA ploidy, and EGFR provided significant prognostic information concerning 4-year relapse-free survival (RFS) with the odds ratios (ORs) of not relapsing of 2.94, 2.83, 2.34, and 2.20, respectively. Regarding overall survival (OS) at 4 years, only nodal status and DNA ploidy had prognostic significance, with the ORs of not dying of 2.68 and 2.80, respectively. Applying multivariate analysis to RFS, 21N when adjusted for nodal status, EGFR, and DNA ploidy (full model) failed to retain prognostic value (P = .202), whereas nodal status was the most significant indicator of relapse (P = .027) followed by DNA ploidy (P = .056) and EGFR (P = .093).CONCLUSIONS:This study suggests that overexpression of the c-erbB-2 oncoprotein appears to be an important indicator of relapse in stage I-II breast cancer when singly evaluated. Multivariate analysis shows that the determination both of nodal status and DNA ploidy improves our ability to identify subsets of patients with different prognoses, and allows for a better selection of patients for systemic adjuvant treatments.

The PC-10 monoclonal antibody to PCNA was employed to analyze proliferative grade in conventionally-formalin fixed, paraffin-embedded tumour samples of 162 patients with primary breast carcinoma. To perform the immunocytochemical method, sections were not heated, were de-waxed using alcohol, and then immersed in a phosphate-buffered saline solution and in methanol with 0.5% hydrogen peroxide to block endogenous peroxidase activity. Immunostaining was performed by a streptavidin-biotin peroxidase substrate.A semiquantitative scoring system was used to evaluate the fraction of nuclei that were PCNA-positive. The score ranged from 0% to 75% with a median value of 25%, mean of 27.8 +/- 1.5. PCNA staining was significantly associated with oestrogen receptor-negativity (p=0.011) and correlated, but not at a statistically significant level, with tumour size (p = 0.08). No significant association was observed between PCNA and node status, grading, DNA ploidy, progesterone receptor or menopausal status. Prognostic indices such as number of positive lymph nodes and DNA ploidy were significantly associated with relapse-free survival (RFS) and overall survival (OS). No significant correlation between PCNA nuclear immunostaining and RFS or OS was observed after a median follow-up of 4 years. Our results indicate that analysis of PCNA alone does not seem to be a useful marker in identifying patients at different prognosis in human breast cancer.

BACKGROUND:Axillary lymph node status has been the most important prognostic factor in operable breast carcinoma, but it does not fully account for the varied disease outcome. More accurate prognostic indicators would help in selection of patients at high risk for disease recurrence and death who are candidates for systemic adjuvant therapy. Our recent findings indicated that microvessel density (count or grade) in invasive breast carcinoma (a measure of tumor angiogenesis) is associated with metastasis and thus may be a prognostic indicator.PURPOSE:This study was designed to further define the relationship of microvessel density with overall and relapse-free survival and with other reported prognostic indicators in breast carcinoma.METHODS:In a prospective, blinded study of 165 consecutive patients, the microvessels within primary invasive breast carcinoma were highlighted by immunocytochemical staining to detect factor VIII-related antigen. Using light microscopy, we counted microvessels per 200x field in the most active areas of neovascularization and graded microvessel density. These findings were correlated, by univariate and multivariate analyses, with overall and relapse-free survival, axillary node status, and other prognostic indicators (median follow-up, 51 months).RESULTS:There was a highly significant (P < or = .001) association of microvessel density with overall survival and relapse-free survival in all patients, including node-negative and node-positive subsets. All patients with breast carcinomas having more than 100 microvessels per 200x field experienced tumor recurrence within 33 months of diagnosis, compared with less than 5% of the patients with breast carcinoma having 33 or fewer microvessels per 200x field. Moreover, microvessel density was the only statistically significant predictor of overall survival among node-negative women (P < .001). Only microvessel density (P < .001) and histologic grade (P = .04) showed statistically significant correlations with relapse-free survival in the node-negative subset.CONCLUSIONS:Microvessel density in the area of the most intense neovascularization in invasive breast carcinoma is an independent and highly significant prognostic indicator for overall and relapse-free survival in patients with early-stage breast carcinoma (I or II by International Union Against Cancer criteria).IMPLICATIONS:Such an indicator would be useful in selection of those node-negative patients with breast carcinoma who are at high risk for having occult metastasis at presentation. These patients could then be given systemic adjuvant therapy.

A retrospective analysis is reported in 232 patients with clinical Stage I endometrial adenocarcinoma diagnosed between 1980 and 1988, treated with combined surgery and adjuvant irradiation. Tumors were restaged according to the FIGO (1988) surgical staging system (SSS) in order to assess the prognostic value of this revised classification and of the conventional clinico-pathological features (age, performance status, grade and depth of myometrial invasion). The 5-year relapse-free survival (RFS) and overall survival (OS) of the series was 85% and 90.8%, respectively. Both 5-year RFS and OS were not significantly different adopting the FIGO (1971) clinical Stage (Stage IA 93% and 88% vs Stage IB 88.5 and 82%. respectively) whereas they were significant by FIGO SSS (Stage IA 96% and 93% vs Stage IB 94% and 88.5% vs Stage IC 74% and 63%, respectively) (p=0.001 and p=0.0005, respectively). Other factors that significantly affected 5-year RFS or OS at univariate analysis were age (p=0.01 and p<0.0001, respectively), performance status (p=0.035 and p=0.001, respectively), grade (p=0.015 and p<0.0001, respectively) and myometrial invasion (p=0.0017 and p=0.0003, respectively). A multivariate analysis of these prognostic variables showed that FIGO SSS was the only significant and independent (p=0.01) indicator for recurrence. However, when therapy was added to the model, FIGO SSS failed to retain significance (p=0.11). Concerning OS age (p<0.0001), performance status (p=0.04) and FIGO SSS (p=0.05) were significant and independent prognosticators, also when therapy was included in the analysis. In conclusion, this study shows that the new FIGO SSS is a useful prognosticator as well as is age and performance status, in early Stage endometrial adenocarcinoma.