Hepcidin and ferritin blood level as noninvasive tools for predicting breast cancer.

BACKGROUND: Currently used CA15-3 and CEA have found their clinical application particularly in the follow-up of patients with advanced disease. Novel biomarkers are urgent, especially for improving early diagnosis as well as for discriminating between benign and malignant disease. PATIENTS AND METHODS: In the present study, we used a proteomic approach based on surface-enhanced laser desorption/ionization-time of flight-mass spectrometry screening with the aim of identifying differentially expressed 2-30 kDa proteins in plasma of patients with malignant (65 cases) and benign (88 cases) breast lesions with respect to 121 healthy controls. RESULTS: We found that the most promising SELDI peaks were those corresponding to hepcidin-25 and ferritin light chain. We evaluated the capability of these peaks in predicting malignant and benign breast lesions using the area under the receiver operating characteristic curve (AUC). The results showed a good capacity to predict malignant breast lesions for hepcidin-25 [AUC: 0.82; 95% confidence interval (CI) 0.75-0.90] and ferritin light chain (AUC: 0.86; 95% CI 0.79-0.92). Conversely, a weak and satisfactory capability to predict benign breast lesion was observed for hepcidin-25 (AUC: 0.63; 95% CI 0.41-0.85) and ferritin light chain (AUC: 0.73; 95% CI 0.49-0.97). A significant association between HER2 status and hepcidin-25 was observed and the distribution of transferrin and ferritin were found significantly different in patients with breast cancer when compared with that of controls. CONCLUSIONS: This study provides evidence that hepcidin and ferritin light chain level in plasma may be of clinical usefulness to predict malignant and benign disease with respect to healthy controls.

Blood-based genomics of triple-negative breast cancer progression in patients treated with neoadjuvant chemotherapy.

BACKGROUND: As neoadjuvant chemotherapy (NAC) is increasingly used in triple-negative breast cancer (TNBC), we investigated the value of circulating tumor DNA (ctDNA) for patient monitoring prior, during, and after NAC, and circulating tumor cells (CTCs) for disease characterization at clinical progression. MATERIALS AND METHODS: Forty-two TNBC patients undergoing NAC were prospectively enrolled. Primary tumor mutations identified by targeted-gene sequencing were validated and tracked in 168 plasma samples longitudinally collected at multiple time-points by droplet digital polymerase chain reaction. At progression, plasma DNA underwent direct targeted-gene assay, and CTCs were collected and analyzed for copy number alterations (CNAs) by low-pass whole genome sequencing. RESULTS: ctDNA detection after NAC was associated with increased risk of relapse, with 2-year event-free survival estimates being 44.4% [95% confidence interval (CI) 21.4%-92.3%] versus 77.4% (95% CI 57.8%-100%). ctDNA prognostic value remained worthy even after adjusting for age, residual disease, systemic inflammatory indices, and Ki-67 [hazard ratio (HR) 1.91; 95% CI 0.51-7.08]. During follow-up, ctDNA was undetectable in non-recurrent cases with the unique exception of one showing a temporary peak over eight samples. Conversely, ctDNA was detected in 8/11 recurrent cases, and predated the clinical diagnosis up to 13 months. Notably, recurrent cases without ctDNA developed locoregional, contralateral, and bone-only disease. At clinical progression, CTCs presented chromosome 10 and 21q CNAs whose network analysis showed connected modules including HER/PI3K/Ras/JAK signaling and immune response. CONCLUSION: ctDNA is not only associated with but is also predictive of prognosis in TNBC patients receiving NAC, and represents an exploitable tool, either alone or with CTCs, for personalized TNBC management.

p53 as an independent prognostic marker in lymph node-negative breast cancer patients.

BACKGROUND: At present, most decisions concerning the use of adjuvant therapy in lymph node-negative breast cancer patients are made on the basis of traditional factors such as tumor size, nodal status, and histopathologic features. However, prognostic factors are being investigated that could identify high-risk groups and that could better address treatment efforts for those patients. Identification of more accurate prognostic markers, such as the expression of the mutant p53 protein encoded by the p53 (also known as TP53) tumor suppressor gene, that are reproducible, easily assessable, and independent in predicting clinical outcome would have a beneficial impact on cancer treatment decisions. PURPOSE: Our purpose was to analyze the predictive relevance of mutant p53 protein expression on 6-year relapse-free and overall survival in node-negative breast cancer patients in relation to menopausal status, tumor size, cell kinetics, and estrogen receptor status. METHODS: Expression of mutant p53 protein was detected by an immunohistochemical technique using a 1:50 dilution of PAb1801 monoclonal antibody on paraffin-embedded tumor specimens obtained from 256 axillary lymph node-negative breast cancer patients, with long-term follow-up (median, 72 months). The [3H]thymidine labeling index, a measure of cell kinetics, was evaluated on histologic sections after fresh tumor tissue was labeled with [3H]thymidine. Estrogen receptor status was determined by the dextran-coated charcoal absorption technique. Statistical comparisons were made for levels of p53 protein expression, [3H]thymidine labeling index, estrogen receptor status, tumor size, and menopausal status with respect to 6-year relapse-free survival and overall survival. RESULTS: Overexpression of the p53 protein, defined as the presence of more than 5% positive cells, was detected in 113 (44%) of 256 tumors. Odds ratios (ORs) for multiple regression analysis of 6-year relapse-free survival were significantly higher for p53 (OR = 3.24; 95% confidence limits [CL] = 2.01-5.23) and [3H]thymidine labeling index (OR = 1.92; 95% CL = 1.19-3.12), both of which appeared to be the most relevant indicators of relapse, than for tumor size (OR = 1.49; 95% CL = 0.94-2.38) and estrogen receptor status (OR = 0.91; 95% CL = 0.55-1.51). Overexpression was found to be unrelated to menopausal status. CONCLUSIONS: Immunohistochemically detected p53 overexpression is an independent marker for shortened 6-year relapse-free and overall survival in node-negative patients with resectable breast cancers. Based on these findings, p53 overexpression should be used with other established prognostic factors, such as [3H]thymidine labeling index and estrogen receptor status, to further refine the prognostic assessment of node-negative breast cancer.

The Bcl-2 protein: a prognostic indicator strongly related to p53 protein in lymph node-negative breast cancer patients.

BACKGROUND: The bcl-2 gene (also known as BCL2) encodes for a mitochondrial protein thought to prevent apoptosis of normal cells. The protein has been detected by immunohistochemical procedures in hormonally regulated epithelia. PURPOSE: We analyzed the predictive relevance of Bcl-2 expression on 6-year relapse-free and overall survival in lymph node-negative breast cancers in relation to pathologic (tumor size) and biologic ([3H]thymidine-labeling index, p53 protein expression, and estrogen receptor [ER] status) features. METHODS: The expression of Bcl-2 and p53 was detected by immunohistochemistry on paraffin-embedded sections from 283 node-negative resectable breast cancers treated with local-regional therapy alone until relapse. The [3H]thymidine-labeling index was evaluated on histologic sections after incubation of fresh tumor tissue with [3H]thymidine, and ER content was determined by the dextran-coated charcoal absorption technique. RESULTS: A significantly higher fraction of Bcl-2-positive cells was observed in small, ER-positive, slowly proliferating, and p53-negative tumors than in large, ER-negative, rapidly proliferating, and p53-positive tumors. A stronger association was observed between Bcl-2 and p53 expression than between these variables and [3H]thymidine-labeling index. In univariate analysis, Bcl-2 and p53 expression, [3H]thymidine-labeling index, tumor size, and ER status were indicators for relapse-free and, with the exception of tumor size, overall survival within 6 years of surgery. In multivariate analysis, Bcl-2 failed to maintain its prognostic role for relapse-free and overall survival in the presence of p53 expression, whereas the [3H]thymidine-labeling index was still statistically significant as a predictor for both events. CONCLUSION: The predictive role of Bcl-2 expression on 6-year relapse-free and overall survival was mainly dependent on p53 expression.

Expression of p53, glutathione S-transferase-pi, and Bcl-2 proteins and benefit from adjuvant radiotherapy in breast cancer.

BACKGROUND: In clinical breast cancer research, the utility of certain biomarkers as predictors of response to surgery, chemotherapy, or hormonal therapy has been studied intensively. Much less research has been done on the relevance of biologic predictors of response to radiotherapy, which represents an effective local-regional treatment for breast cancer. PURPOSE: The utility of biomarkers involved in DNA damage repair (p53 protein), control of programmed cell death (p53 and Bcl-2 proteins), and cellular detoxification (glutathione S-transferase-pi [GST-pi] enzyme) in predicting local breast cancer recurrence was analyzed retrospectively in two cohorts of breast cancer patients. These patients had had no detectable metastases in the axillary lymph nodes (i.e., node-negative) or in distant sites and had had similar distributions of clinicopathologic and biologic prognostic features. They had been treated by conservative surgery alone (139 case patients) or by conservative surgery followed by adjuvant radiotherapy (496 case patients) during the period from 1984 through 1990. METHODS: The expression of the p53, GST-pi, and Bcl-2 proteins in the specimens of primary breast tumor obtained from these patients was determined by use of immunohistochemistry; cell proliferation activity and levels of steroid receptors were determined by use of a [3H]thymidine-labeling index assay and the dextran-coated charcoal technique, respectively. The median time of follow-up of patients was 6 years. In the analyses of patient outcomes, only local failures that presented as first events were considered. RESULTS: After surgery alone, the risk of local recurrence at 6 years was higher for patients with tumors exhibiting elevated levels of p53 and GST-pi protein expression than for patients with low levels (hazard ratio [HR] = 3.1, 95% confidence interval [CI] = 1.3-7.7, two-sided P = .012; HR = 2.7, 95% CI = 1.1-6.4, two-sided P = .026, respectively). Weak or no observable expression of Bcl-2 protein was only suggestive of a higher frequency of local failures. Adjustment for patient age, tumor size, cell proliferation, and estrogen receptor status did not change these findings. Conversely, in the series of patients given conservative surgery followed by radiotherapy, there was no difference in local tumor recurrence between patients with tumors expressing or not expressing each of the three markers. CONCLUSIONS: Our study provides indirect evidence of a benefit from radiation therapy in preventing local breast cancer relapse, particularly among node-negative patients with tumors that express elevated levels of the p53 or GST-pi proteins or that express little or no Bcl-2 protein.

p53 and bcl-2 expression correlates with clinical outcome in a series of node-positive breast cancer patients.

BACKGROUND AND PURPOSE: The tumor-suppressor gene TP53 and the proto-oncogene bcl-2 encode, respectively, for a nuclear phosphoprotein and for a mitochondrial protein involved in multiple cellular functions. The proteins provide prognostic information in node-negative breast cancer and are supposed to influence treatment responsiveness. We analyzed the predictive role of p53 and bcl-2 expression, alone and in association with other variables, in postmenopausal women with node-positive, estrogen receptor-positive (ER+) breast cancers treated with radical or conservative surgery plus radiotherapy and adjuvant tamoxifen for at least 1 year. PATIENTS AND METHODS: On 240 resectable cancers, we determined the expression of p53 and bcl-2, using immunohistochemistry, cell proliferation (3H-thymidine labeling index [3H-dT LI]), and ER and progesterone receptors (PgR). RESULTS: p53 expression and 3H-dT LI were weakly related to one another and both were unrelated to bcl-2. Relapse-free and distant metastasis-free survival at 5 years were significantly lower for patients with tumors that highly expressed p53 (P = .0001) and for those that weakly expressed or did not express bcl-2 (P = .02). However, p53, but not bcl-2, provided prognostic information independent of tumor size, axillary node involvement, steroid receptors, and 3H-dT LI. Moreover, the simultaneous p53 overexpression and lack of PgR identified patients at maximum risk of relapse, whereas bcl-2 overexpression, associated with a low 3H-dT LI or the presence of PgR, improved the prognostic resolution for low-risk patients. CONCLUSION: p53 expression appears to be indicative of clinical outcome in postmenopausal patients treated with tamoxifen. Whether p53 overexpression and weak bcl-2 expression are indicators of biologic aggressiveness, regardless of treatment, or of hormone resistance remains to be defined.

Clinical studies of Bcl-2 and treatment benefit in breast cancer patients.

Interest in translational studies aimed at investigating the role of biologic markers in predicting clinical outcome of breast cancer patients and, in particular, response to specific treatments, has progressively increased. Among biologic variables presently under investigation, apoptosis markers, in particular Bcl-2 and Bax expression, are receiving much attention for their relationship with the cellular response to genotoxic damage in experimental tumors. Retrospective, independent studies were carried out by several research groups on about 5000 patients with breast cancer at different stages and with an adequate follow-up. The outcome of separate analyses as a function of treatment generally demonstrated that Bcl-2 overexpression, which correlates with biologic features of a differentiated phenotype (slow proliferation, high steroid receptor levels, absence of p53 and c-erB-2 expression), is associated with a favorable outcome. Such a finding is mainly evident following surgery as well as endocrine treatment. Conversely, no or weak Bcl-2 expression, alone or in association with bax overexpression, appears indicative of a radiation response, and preliminary emerging evidence supports the involvement of such an association of apoptosis-related markers even as predictors of long-term response to neoadjuvant cytotoxic treatment. Although the findings of an involvement of Bcl-2 and Bax as determinants of treatment response should be confirmed within the context of randomized clinical trials, they indicate a combined consideration of proteins that negatively and positively regulate apoptosis in translational studies on the effect of chemical and physical agents at a cellular level.

Expression of P-glycoprotein and in vitro or in vivo resistance to doxorubicin and cisplatin in breast and ovarian cancers.

The expression of P-glycoprotein (P-gp) was studied by immunocytohistochemistry, using the C219 monoclonal antibody, in 39 locally advanced breast cancers and 20 ovarian cancers from previously untreated patients. P-gp was expressed in 46 and 35% of breast and ovarian tumours, respectively. A significant association was observed in both tumour types between P-gp expression and in vitro resistance to doxorubicin. We also observed a higher clinical response rate to doxorubicin +/- vincristine in patients with breast cancers not expressing P-gp. Conversely, no correlation was found between P-gp expression and in vitro resistance to cisplatin or in vivo response to cisplatin +/- cyclophosphamide treatment in ovarian cancers. Our results support the relevance of P-gp expression as a specific indicator of resistance to certain drugs, such as doxorubicin and vincristine, involved in the phenomenon of multidrug resistance in breast and ovarian cancer cells.

P53 and cathepsin-d are independent of established prognostic factors in breast-cancer.

p53 and cathepsin D expression was investigated in 300 primary breast cancers by the avidin-biotin immunoperoxidase method using two murine monoclonal antibodies: PAb1801 and anti-procathepsin D, respectively. The frequency of p53- and cathepsin D-positive cells varied widely among different tumors and most tumors (82% and 86%) at least occasionally showed positive cells. The two biological markers were unrelated to one another, to cell proliferative rate and ploidy and were differently related to other biological and pathological features. In particular, p53 was directly related to tumor size and nodal involvement and inversely related to the presence of steroid receptors. Conversely, cathepsin D was directly related only to nodal involvement.

Biological predictors of response to radiotherapy in cervical carcinomas.

On 21 patients with T1b-T3b tumours subjected to external radiotherapy and brachytherapy, the expression of P53 and glutathione S-transferase pi [GST-pi], immunohistochemically detected, the S-phase cells fraction (H-3-thymidine labeling index, TLI) and DNA content evaluated by image analysis were determined on biopsies before and after the first 10 Gy. P53 accumulation was reduced in 60% of P53-overexpressing tumours and not induced in P53-negative tumours, GST-pi was induced in about 40% of pretreatment GST-pi-negative cases, TLI was reduced in 70% of the cases regardless of pretreatment values, and DNA profiles remained unchanged in two-thirds of the cases. P53 accumulation was a predictor of 3-year relapse-free survival after radiotherapy, followed by GST-pi expression, whereas TLI did not influence prognosis.

Biological markers in transurethral biopsies from bladder-cancer - preliminary-results.

A biologic profile including proliferative.activity, evaluated as H-3-thymidine labeling index (H-3-dT LI), DNA ploidy, p53 tumor-suppressor gene and P-glycoprotein (P-170), as an expression of the multidrug resistance gene, was defined for 50 primary transitional cell carcinomas of the bladder. H-3-dT LI was evaluated by autoradiography on histologic sections after incubation of fresh tumor biopsies with H-3-thymidine. Ploidy was defined by flow cytometric analysis of DNA content on nuclei suspensions obtained from frozen material. Expression of p53 protein and P-170 glycoprotein was detected by immunohistochemistry using the PAb1801 and C219 monoclonal antibody respectively, on sections from paraffin-embedded tumor biopsies. Invasive tumors showed a higher median H-3-dT LI (12.7% vs 4.2%) and a higher frequency of aneuploidy (73% vs 43%) and more frequently expressed p53 (82% vs 36%) than superficial tumors. Further analysis showed that proliferative activity was higher in invasive than in superficial cancers only in p53-positive or aneuploid tumors and not in p53-negative or diploid tumors. Moreover, proliferative activity and p53 overexpression, but not ploidy, were directly related to histologic grading and tumor stage. Generally, P-170 was not significantly related to any biologic or clinico-pathologic factor. Kinetic and phenotypic biologic markers are differently related to clinico-pathologic factors. A panel of biologic features can be easily evaluated on small transurethral biopsies at diagnosis, during endocavitary treatment or follow-up in bladder cancer patients.

Bcl-2 protein and prognosis in patients with potentially curable non-small-cell lung cancer.

The bcl-2 proto-oncogene functions as a cell death suppressor, and its expression prolongs cell survival by blocking apoptosis. Data available on the clinical relevance of bcl-2 protein expression in patients with non-small-cell lung cancer (NSCLC) are controversial. We analysed the role of bcl-2 protein expression on 6-year relapse-free survival in 229 patients with stage I-IIIa NSCLC (101 squamous cell carcinomas and 128 adenocarcinomas) subjected to surgery, with curative intent. Immunohistochemical analysis was performed on archival material by using a monoclonal antibody anti-bcl-2 (clone 124). Bcl-2 protein expression, which was detected in 22% of the cases, was significantly related to stage, histology and grading, and was an indicator of clinical outcome. The probability of relapse-free survival at 6 years was longer for patients with bcl-2-positive tumours (74%) than for those with bcl-2-negative tumours (57%) (P=0.02). This finding was mainly evident for the subgroups of patients with stage IIIa tumours (P=0.05), squamous cell carcinoma (P=0.03) or moderately/poorly differentiated tumours (P=0.02). However, multivariate analysis by Weibull's regression model indicated that bcl-2 protein expression was not an independent prognostic risk factor in patients with curable NSCLC when the information provided by stage was available.

Biological indicators of survival in patients treated by surgery for squamous cell carcinoma of the oral cavity and oropharynx.

Squamous cell carcinomas of the head and neck are a heterogeneous group of tumours with regard to anatomical site, natural history and response to various treatments. Assessment of the role of biomarkers as indicators of prognosis or response to treatment is thus complex. In the last decade, different biomarkers have been investigated in the search for objective and reproducible indicators of prognosis. In 69 squamous cell carcinomas of the oral cavity or oropharynx from patients treated with radical surgery alone, we determined cell kinetics, evaluated as in vitro 3H-thymidine labelling index (TLI), p53, bcl-2 and glutathione S-transferase pi (GST pi) expression, by using immunohistochemical methods. The biological variables were unrelated to one another or to established clinical and pathological prognostic factors. Univariate analysis showed that a low proliferative activity was associated to a significantly higher risk of death than that observed in patients with a high TLI, whereas p53, bcl-2 and GST pi expression did not provide prognostic information. Multivariate analysis showed that cell proliferation, gender and nodal status retained their clinical relevance. In the subset of node-negative patients, TLI and p53 expression were indicators of survival. Moreover, the combined analysis of TLI and p53 expression identified a subgroup of node-negative patients with slowly proliferating and highly p53-expressing tumours who died within 1 year of radical surgery. These results indicate that in patients with operable oral cavity and oropharyngeal cancer, biomarkers can provide important information on clinical outcome.

Potentiation of cisplatin cytotoxicity by lonidamine in primary cultures of human ovarian cancer.

The ability of lonidamine, an energolytic derivative of indazole-carboxylic acid, to modulate the cytotoxic activity of cisplatin was investigated on primary cultures obtained from 11 human ovarian carcinomas. A 72-h postincubation with lonidamine potentiated the activity of a 1-h cisplatin treatment. Statistical analysis of the dose-effect plots indicated that the interaction between the two drugs was synergistic in 4 tumors and additive in the remaining 7 tumors. The occurrence of the synergistic effect was independent of some biological characteristics of the tumor cell population, such as cell kinetics (as assessed by 3H-thymidine labeling index), DNA content and the expression of the putative factor of cisplatin resistance, glutathione-S-transferase pi.

Comparative in vitro sensitivity of human cholangiocarcinoma and colon adenocarcinoma cells to anticancer agents.

We comparatively investigated the sensitivity of a human cholangiocarcinoma cell line (SG231) and an adenocarcinoma cell line (WiDr) to mitoxantrone (MX), taxol (TX), mitomycin C (MMC), doxorubicin (DX), cisplatin (CDDP) and 5-fluorouracil (5FU) by the sulforhodamine B assay. The lower susceptibility of SG231 to SFU, to CDDP, to DX and to MMC than WiDr was observed, whereas the sensitivity of the two cell lines to MX and TX was similar. We also investigated the ability of a chemical modulator, lonidomine (LND), and hyperthermia to enhance the cytotoxic activity of the different drugs in the SG231 cell line. No potentiation of MX or CDDP activity was observed after a 2 hours treatment in hyperthermic conditions (42 degrees C). Conversely, a slight potentiation of a 2 hours pretreatment with MX and DX was obtained by a 24 hours post treatment with LND.

Comparative analysis of different approaches to investigate cell kinetics.

The potential of different methods to investigate proliferative activity of cell populations was analysed for non-Hodgkin's lymphomas. Cells in S phase and all cycling cells were determined on cell suspensions obtained from fresh lymph node material by [3H]-thymidine autoradiography [( 3H]TdR LI), a monoclonal antibody to bromodeoxyuridine (BrdU LI), and the monoclonal antibody Ki67. A good correlation was observed between the values of [3H]TdR LI and BrdU LI (rs = 0.90; P less than 0.01), [3H]TdR LI and S phase (rs = 0.62; P less than 0.01) and [3H]TdR LI and Ki67 (rs = 0.64; P less than 0.01) in individual lymphomas. Using the median values obtained from the different approaches as cut-off points to define slowly and rapidly proliferating tumours, the best agreement was observed between [3H]TdR LI and BrdU LI (91%) and poorer agreements, even though statistically significant, were observed between [3H]TdR LI and S phase (73%) or Ki67 (76%). In conclusion, the kinetic information derived from different approaches was more or less concordant and newly proposed approaches should be directly and carefully verified for their prognostic relevance before using them as alternatives to conventional methods.

Comparative analysis of [3H]-thymidine labelling index and monoclonal antibody Ki-67 in non-Hodgkin's lymphomas.

The relation between the [3H]-thymidine labelling index (3H-Thy LI), which evaluates the S-phase cells, and the monoclonal antibody Ki-67 (MoAb Ki-67), which recognizes a nuclear antigen expressed during the cell cycle, was defined in 35 non-Hodgkin's lymphomas (NHL). Significantly higher median values of [3H]-Thy LI and Ki-67-positive cells were observed for high-grade than for low-grade malignancy tumours according to the Kiel classification, but with wide and overlapping values for the two morphologic subgroups. A significant correlation was observed between [3H]-Thy LI and Ki-67-positive cells (P less than 0.0001; r = 0.62). However, the ratio between the two cell kinetic variables on individual tumours was not constant. It sharply increased with decreasing [3H]-Thy LI values and was much higher in low-grade NHL than in high-grade NHL. Follow-up studies are needed to establish the role of the tumour growth fraction as evaluated by MoAb Ki-67 as a prognostic indicator in NHL.

Morphological and clinical significance of cell kinetics in non-Hodgkin's lymphomas.

The relation between the proliferative activity and morphologic features according to Rappaport and Kiel classifications and the Working Formulation (WF) was analyzed in a series of 123 adult patients with non-Hodgkin lymphomas. Cell kinetics was determined as in vitro 3H-thymidine labelling index (LI). A significant association was observed between low LI and follicular histology or low grade of the WF, as well as between high LI and high-grade of malignancy of the Kiel and WF. The analysis of the clinical relevance of cell kinetics on objective response to treatment showed a higher frequency (91%) of complete remission in the tumors with low proliferative activity versus tumors with high proliferative activity (48%) (p = 0.00003). Moreover, cell kinetics proved to be and important indicator of 6-year survival on the whole population of patients (66% for slowly vs 18% for fast-proliferating tumors; p less than 10(-9)) and a further discriminant within diffuse histology (68% vs 9%; p less than 0.0001), low-grade malignancy according to the Kiel (64% vs 15%; p less than 10(-8)) and low (64% vs 28%; p less than 0.005) and intermediate grades of the WF (68% vs 9%; p less than 0.0001).

Contribution of 3H-thymidine labelling index and flow cytometric S-phase in predicting survival of patients with non-Hodgkin's lymphoma.

The 3H-thymidine labelling index (3H-dT LI) of cell suspensions from fresh material and the flow cytometric S-phase (FCM-S) of nuclei recovered from paraffin blocks were determined on the same pathologic lymph node specimen for 190 non-Hodgkin's lymphomas (NHLs). FCM-S was defined by a planimetric method and by an optimization procedure. Poor correlation coefficients were observed among the three cell kinetic variables. All three cell kinetic variables were significant indicators of 8-year survival and median survival time. The life-regression procedure evidenced a significant relative contribution of 3H-dT LI and FCM-S, thus suggesting a different biologic meaning of the two cell kinetic variables. This finding was further supported by evidence that simultaneous use of 3H-dT LI and FCM-S can identify groups of patients with different survival better than when either modality is used alone. Multivariate analysis indicated that the risk groups as defined by cell kinetic variables are predictors of survival even in the presence of established factors such as histology and stage.