Raman spectroscopy and PCA-SVM as a non-invasive diagnostic tool to identify and classify qualitatively glycated hemoglobin levels in vivo.

In this study we identify and classify high and low levels of glycated hemoglobin (HbA1c) in healthy volunteers (HV) and diabetic patients (DP). Overall, 86 subjects were evaluated. The Raman spectrum was measured in three anatomical regions of the body: index fingertip, right ear lobe, and forehead. The measurements were performed to compare the difference between the HV and DP (22 well controlled diabetic patients (WCDP) (HbA1c <6.5%), and 49 not controlled diabetic patients (NCDP) (HbA1c ≥6.5%)). Multivariable methods such as principal components analysis (PCA) combined with support vector machine (SVM) were used to develop effective diagnostic algorithms for classification among these groups. The forehead of HV versus WCDP showed the highest sensitivity (100%) and specificity (100%). Sensitivity (100%) and specificity (60%), were highest in the forehead of WCDP, versus NCDP. In HV versus NCDP, the fingertip had the highest sensitivity (100%) and specificity (80%). The efficacy of the diagnostic algorithm by receiver operating characteristic (ROC) curve was confirmed. Overall, our study demonstrated that the combination of Raman spectroscopy and PCA-SVM are feasible non-invasive diagnostic tool in diabetes to classify qualitatively high and low levels of HbA1c in vivo.

Autologous stem cell transplantation for Hodgkin's disease: busulfan, melphalan and thiotepa compared to a radiation-based regimen.

We evaluated prognostic factors and treatment outcome of patients with relapsed/refractory Hodgkin's disease (HD) receiving autologous stem cell transplantation (ASCT). In total, 92 patients received total body irradiation, cyclophosphamide and etoposide (TBI/CY/E) (n=42) or busulfan, melphalan and thiotepa (Bu/Mel/T) (n=50) supported with ASCT. A total of 33 (66%) patients receiving the Bu/Mel/T regimen had a prior history of dose-limiting irradiation. Mucositis, hepatic and pulmonary toxicities were the main causes of morbidity and mortality, irrespective of the conditioning regimen. The transplant-related mortality was 15%. With a median follow-up of 6 years (range 2.5-11), the cumulative probabilities of survival, event-free survival (EFS) and relapse at 6 years were 55, 51 and 32%. The 6-year Kaplan-Meier (KM) probabilities of EFS for patients with less advanced disease (patients in first chemotherapy-responsive relapse or second remission (n=42)) and more advanced disease (all other patients (n=50)) were 60 and 44%. No differences in toxicities and efficacy between the conditioning regimens were found. ASCT is an effective treatment for patients with refractory/relapsed HD. Female patients and patients with less advanced disease at transplant had a better outcome. Patients with prior irradiation benefited from the Bu/Mel/T regimen.

Autologous stem cell transplantation for non-Hodgkin's lymphoma: comparison of radiation-based and chemotherapy-only preparative regimens.

The aim of this study was to compare toxicity and efficacy of total body irradiation (TBI), cyclophosphamide (CY) and etoposide (E) (TBI/CY/E) vs busulfan, melphalan and thiotepa (Bu/Mel/T) in patients receiving autologous stem cell infusion (ASCI) for malignant lymphoma (NHL). Between September 1990 and July 1998, 351 patients with NHL were treated with TBI/CY/E (n = 221) or Bu/Mel/T (n = 130) followed by ASCI. Patients in first, or second remission, first responding or untreated relapse were defined as having less advanced disease before transplantation. The median follow-up was 5 years (range 1-9) and 3.5 years (1-6) for patients receiving TBI/CY/E and Bu/Mel/T, respectively. The cumulative probabilities of survival, event-free survival (EFS) and relapse at 5 years were 44%, 32%, 49% following TBI/CY/E and 42%, 34% and 42% following Bu/Mel/T. The probability of EFS at 5 years for patients who had prior dose-limiting radiation (n = 59) was 32% after Bu/Mel/T therapy. Transplant-related mortality was 16% for TBI/CY/E and 21% for Bu/Mel/T. In univariate and multivariate analyses, more advanced disease status was associated with poor outcome (TBI/CY/E: RR 0.70, CI 0.50 to 0.97 P = 0.04; Bu/Mel/T: RR 0.61, CI 0.39 to 0.97 P = 0.03). No significant differences in toxicities and outcomes were observed between these two regimens despite the inclusion of patients who had received dose-limiting irradiation in the Bu/Mel/T regimen.

Safety of granulocyte colony-stimulating factor in normal donors.

Granulocyte colony-stimulating factor (G-CSF), which is widely used to mobilize peripheral blood stem cells (PBSC) from normal donors, has led to the use of PBSC as a major alternative to bone marrow for patients undergoing allogeneic transplants. Safety issues related to the administration of G-CSF to normal donors, however, are still under study. The short-term effects after G-CSF administration are well known and manageable. G-CSF induces a hypercoagulable state, which may predispose certain donors to thrombotic complications. A dose of 10 microg/kg/d for 5 days has been recommended for routine clinical use, but the optimal dose and schedule for PBSC collection are still being defined. Small studies to date have shown no late effects of G-CSF administration but there is insufficient information regarding any long-term adverse effects or risks. Although the administration of G-CSF to normal donors for PBSC collection appears safe, longer follow-up is required.

High-dose busulfan, melphalan and thiotepa as consolidation for non-inflammatory high-risk breast cancer.

The purpose of this study was to evaluate the toxicity and efficacy of high-dose busulfan, melphalan and thiotepa (Bu/Mel/TT) in patients with high-risk non-inflammatory breast cancer defined as stage II disease > or =10 lymph nodes (n = 52) or stage III (n = 69), and prognostic factors for treatment outcome. One hundred and twenty-one patients (median age, 46 years) were treated with high-dose Bu (12 mg/kg), Mel (100 mg/m2) and TT (500 mg/m2) (HDC) followed by autologous stem cell infusion (ASCI). One hundred patients were initially treated with surgery followed by standard adjuvant chemotherapy prior to HDC/ASCI. Twenty-one patients with stage III disease had inoperable tumors at diagnosis and were treated with neoadjuvant chemotherapy and surgery before HDC/ASCI. Transplant-related mortality was 6%. The probabilities of event-free survival (EFS) at 3 and 5 years (median follow-up of 36 months) from transplant were, for all patients: 0.62-0.60; stage II: 0.71-0.67: stage III: 0.55-0.55 (for stage III adjuvant and neoadjuvant groups: 0.60-0.60 and 0.42-0.42, respectively). Multivariate analysis did not identify variables associated with poor outcome. The efficacy of Bu/Mel/TT is similar to other HDC regimens reported for patients with high-risk non-inflammatory breast cancer. Bu/Mel/TT has high activity in stage II disease and a moderate benefit in stage III operable tumors.

H-ras and Nm23-H1 gene expression and proteolytic activity in squamous cell carcinoma of the uterine cervix.

BACKGROUND: The invasive and metastatic potential of malignant cells results from complex interactions of numerous factors not yet fully understood. Genomic alterations such as ras overexpression and nm23-H1 inhibition have been found to be frequently associated with increased invasiveness in various cancers. On the other hand, secretion of different proteinases are necessary for malignant cells to traverse a network of matrix macromolecules, but the relationship between the genomic alterations and the proteolytic phenotype is still unclear. Our aim was to investigate whether the appearance of the proteolytic phenotype had any correlation with the expression of H-ras and nm23-H1 genes in carcinoma of the uterine cervix. METHODS: Twenty-five samples from patients with carcinoma of the uterine cervix at different clinical stages were studied. Cathepsin B1, plasminogen activator, and collagenase activity were assessed in tissue cytosols using specific synthetic oligopeptides as substrates. The expression of H-ras and nm23-H1 was investigated by means of immunohistochemistry and in situ hybridization. RESULTS: Our results showed that cathepsin B1 was the most consistently elevated proteinase, demonstrating a linear correlation with clinical staging. H-ras expression was found elevated in 40% of the cases. Nm23-H1 protein immunoreactivity was positive in 40% of the cases. No correlation was found among H-ras, cathepsin B1 activity, and survival rate. Among cases with high cysteine proteinase activity, a different clinical behavior depending on the expression of Nm23-H1 was observed. The cases with Nm23-H1 protein had a markedly better survival rate than those lacking this protein. In contrast, the absence of Nm23-H1 in association with high cathepsin B1 activity was a clear indicator of a poor prognosis. CONCLUSIONS: These findings suggest a complex interaction between the proteolytic phenotype and the expression of H-ras and nm23-H1 genes in carcinoma of the cervix that influences the clinical behavior of the tumor.

Drug dose delivery and treatment outcome relationship in standard bleomycin, etoposide and cisplatin combination chemotherapy in nonseminomatous germ cell tumor patients.

This study retrospectively evaluated the influence of drug dose delivery components (DDDC) of bleomycin, etoposide and cisplatin chemotherapy for metastatic nonseminomatous germ cell tumors on treatment outcome (NSGCT). Between December 1987 and January 1995, 75 NSGCT patients were treated with a median of 4 cycles (range 3-8) of cisplatin 120 mg/m2 on day 1, etoposide 100 mg/m2 on days 1 through 5 and bleomycin 30 U on days 1, 3, and 5 every 3 weeks. DDDC, such as cumulative dose, cumulative dose in mg/m2, dose intensity (DI), relative dose intensity (RDI), dose intensity products, and relative dose intensity products by drug, were calculated and tested as possible predictors of treatment outcome in patients classified according to Indiana University (IU), and International Germ Cell Cancer Cooperative Group (IGCCCG) classifications. Overall complete response (CR) rate was 64%, and 3-year progression-free survival (PFS) was 59%. By IU classification there were statistical differences in CR and survival between moderate (89-81%) and advanced disease (42-40%) (p < 0.005), while for patients classified according to IGCCCG criteria, statistical differences in CR and PFS there were not registered. DI (mg/m2/week) and RDI values for the entire group were: cisplatin 33-0.82; etoposide 133-0.80 and bleomycin 11-0.37. We did not observe a statistically significant difference in drug dose delivery components for treatment outcome between patients who achieved a CR and incomplete response when analyzed by either extent of disease or whole group. Extent of disease was the most important predictor of treatment outcome.

The association of human papillomavirus type 16 E6 and E7 antibodies with stage of cervical cancer.

OBJECTIVE: Previous investigators have reported higher HPV type 16 antibody positivity among cervical cancer patients than among healthy women. The objective of this study was to determine the association of HPV 16 antibody levels with the stage of cervical cancer. METHODS: Pretreatment tumor biopsies and sera were obtained from 137 newly diagnosed cervical cancer patients residing in Mexico. Using peptide ELISA and radioimmunoprecipitation assay (RIPA), HPV 16 E6- and E7-specific antibodies were measured. RESULTS: By ELISA, elevated antibody titers to HPV 16 E6 and E7 were detected in 16.8 and 32.8% of the women, respectively. While sera positivity did not differ by disease stage, the mean absorbance in the E7-positive sera was 0.42, 0.62, 0.91, and 0.81 for stages I to IV, respectively. Using RIPA, anti-E6 and E7 positivity was demonstrated in 46.7 and 38.7% of the females, respectively. Although no difference across disease stage was detected for E6, increasing proportions of positivity to E7 with stage of disease was detected. The rates for increasing disease stage were 0.14, 0.37, 0.40, and 0.67. Sera from the 6-month postradiation follow-up examinations of a small group of patients demonstrated a statistically significant decrease in antibody positivity from pretreatment positivity to HPV 16 E6 (n = 14; P = 0.01) and HPV 16 E7 (n = 20; P = 0.0001) using ELISA. CONCLUSIONS: These data suggest that HPV 16 E7 antibody positivity may be associated with stage of cervical cancer. Such immune parameters may be applicable to disease staging, monitoring of recurrence and, perhaps, diagnosis. Further investigation into the relationship of HPV 16 E6 and E7 antibodies with stage of cervical cancer and response to therapy is warranted.

Proteinase activity in invasive cancer of the breast. Correlation with tumor progression.

Hydrolysis of extracellular matrix is a necessary step for malignant cells to invade, and metastasize. Three groups of proteinases, mainly serine, thiol and metalloproteinases, have been found to be secreted by cancer cells and responsible for the proteolytic cascade triggered during invasion. Previous studies from our group and others have shown that the thiol proteinase cathepsin B1 is a constant indicator of tumor invasion in carcinoma of the cervix, although others point to plasminogen activators and collagenases. So far, there are no systematic studies to correlate cathepsin B and plasminogen activator activity with advancing malignant disease and thus estimate its capability as a marker of progression. The purpose of this study was to determine the activity of cathepsin B like proteinase and plasminogen activators in invasive carcinoma of the breast at various clinical stages and with different estrogen receptor status. One hundred patients with carcinoma of the breast at different clinical stages were studied. Cathepsin B and plasminogen activators activity was assessed in tumor cytosols using different synthetic oligopeptides as substrates following the method of Smith. Estrogen receptor concentration was determined with monoclonal antibodies. A statistical analysis and correlation with different clinical stages was performed. Cathepsin B-like activity had a consistent and progressive elevation in direct correlation with clinical stage (stage I, 1.97 SE +/- 0.46; stage II, 6.67 SE +/- 1.12; stage III, 28.19 SE +/- 3.48; nmol/mg/30 min), while plasminogen activators, although constantly elevated, had no correlation with tumor progression. No relation could be found with estrogen receptor status. It is concluded that cathepsin B, but not plasminogen activator, is a good indicator of tumor progression in invasive carcinoma of the breast.

Long term results of treatment in patients with extragonadal germ cell tumours.

From 1979 to 1991 56 patients with extragonadal germ cell tumours (EGCT) received cisplatin based chemotherapy. From 16 patients with seminomatous EGCT 13 achieved complete remission (CR) with chemotherapy alone, 2 with additional radiotherapy with final CR rate of 94%. 5 (31%) patients developed relapses and at a median follow-up of 38 (5-103) months 11 (69%) are alive and 10 (62%) have no evidence of disease (NED). Only 7 patients with non-seminomatous EGCT reached CR with chemotherapy alone and 8 more with additional chemotherapy or surgery. Overall CR was 37% and 3 (20%) relapses have been observed. At a median follow-up of 26 (3-114) months 14 (35%) are alive and remain free of disease, 26 (65%) have died. By univariate analysis seminomatous EGCT patients had a significantly greater likelihood of achieving a CR, for non-seminomatous EGCT BEP induction chemotherapy was superior to VAB-6, and NSEGCT patients with serum levels > 2000 ng/ml had worse prognosis. Current staging systems are insufficient to predict the treatment outcome in EGCT.