Breast Cancer
Klijanienko J, Couturier J, Galut M, et al.
Cancer. 1999;87:312-318.
Fine-needle sampling, although a practical and noninvasive method of tissue acquisition, has rarely been used for HER-2/neu fluorescence in situ hybridization (FISH). To assess HER-2/neu gene amplification in mammary carcinoma, FISH signals on cytology and corresponding tissue biopsies were detected visually and measured by image analysis. The results were correlated with patient and tumor characteristics.
In situ HER-2/neu DNA probe hybridization was performed on 61 cytology specimens and on 47 corresponding frozen sections of breast carcinomas. Tumors were classified by visual evaluation as unamplified, moderately amplified, or highly amplified. Multiparametric image analysis was performed using the Discovery automated image analyzer (Becton Dickinson, Leiden, Netherlands). The integrated fluorescence ratio (IFR) was calculated for each sample as the integrated FISH fluorescence of the tumor cells divided by the integrated FISH fluorescence of internal control cells containing 2 spots. The percentage-positive nuclear area (PPN), calculated as the area of FISH fluorescence divided by the area of nuclear DNA fluorescence, and the PPR, ratio of the PPN of the tumor cells divided by the control cells, were also calculated for each sample.
Visual analysis yielded 46 unamplified and 15 (24.6%) amplified (7 moderately amplified and 8 highly amplified) tumors. Strong (P<.001) correlation between results on cytological and histological materials was obtained. The FISH spots on the cytological preparations were more easily visualized and scored than those on the corresponding tissue sections. Visual HER-2/neu signal scoring was strongly correlated with IFR (P=.0001) and PPR (P=.0001). Within the tumors classified as highly amplified by visual examination, quantitation of the degree of amplification fluorescence signal was possible using image analysis.
Cytologic specimens were a suitable and representative source of materials for detection and quantitation of HER-2/neu gene amplification by FISH and image analysis.
Tiny Samples Offer Potential for a Lot of Information
Editorial Comment by Britt-Marie Ljung, MD
Key words:
Fine needle samples (FNS) or fine needle aspiration biopsies (FNAB) are well established as diagnostic techniques for detecting benign and malignant lesions in breast and in many other body sites. FNS have a number of advantages over traditional open biopsies and core biopsies. They are minimally invasive since very thin 22- to 25-gauge needles are typically used. Thus FNS is very well tolerated by patients, and side effects are usually limited to an insignificant local bruise. Because cell preparations can easily be stained for preliminary examination at the bedside, material can be triaged for special studies as needed, minimizing sampling and obviating return visits for additional sampling. This also allows for a preliminary diagnosis in most cases at the time of sampling. Definitive diagnosis is typically made available within 24 hours. The accuracy in well-trained hands is similar to that of frozen section.
Traditionally, diagnosis of these samples has been based entirely upon morphology, ie, the visual appearance of the cells in the microscope interpreted by a pathologist. While morphology is still the basis for most diagnoses, molecular biology assays may now add additional prognostic information and guide treatment decisions.
One such technique is fluorescence in situ hybridization (FISH). This assay allows the evaluation of gene copy number in a nucleus by applying labeled probes directly to cells. FISH is used to determine amplification of the HER2/neu oncogene in breast cancer. Since intact individual nuclei must be visualized for FISH, fine needle samples consisting entirely of cells with intact nuclei are ideally suited for this test. Histological sections, on the other hand, have been cut into thin slices with a blade, resulting in many partial nuclei and are therefore less well suited for FISH analysis.
Other molecular biology techniques, such as comparative genomic hybridization or gene sequencing, analyze DNA extracted from cells. The number of cells obtained by FNS is sufficient for these analyses, since polymerase chain reaction (PCR) can be employed to amplify the DNA.
In the future, a combination of morphology and molecular biology assays will likely allow us to accurately diagnose and characterize a multitude of tumors to predict prognosis and response to specific therapies. Samples obtained by the minimally invasive technique of FNS are excellent material for these purposes.
Nieto Y, Cagnoni PJ, Shpall EJ, et al.
Clin Cancer Res. 1999;5:1731-1737.
The purpose of this study was to assess the efficacy of high-dose chemotherapy (HDC) with autologous stem cell transplant in stage IV breast cancer patients with minimal metastases. Eligible patients had (a) disease that could be resected en bloc and/or irradiated with curative intent using a single field and could, thus, be rendered as having no evidence of disease; and/or (b) < 5% bone marrow involvement. From September 1991 to August 1997, 40 consecutive patients were prospectively entered into the study. Pre-HDC local treatment consisted of surgery (n=31) and radiotherapy (XRT; n=3). All patients received HDC with cyclophosphamide, cisplatin, and 1,3-bis(2-chloroethyl)-1-nitrosourea and autologous stem cell transplant, with or without CD34 selection. Following HDC, 22 patients received XRT. Four patients died of treatment-related complications. Eighteen patients developed grade 3 nonhematological toxicities (15 lung, 2 cardiomyopathy, and 1 optic neuritis), which resolved with therapy. Within a median follow-up of 49 (15-91) months, 14 patients had relapsed. Twenty-five patients (62.5%) were alive, and 22 patients (55%) were alive and free of disease. Median event-free and overall survivals were 43 and 77 months, respectively. In the subset of patients with one metastatic site, 17 of 24 (68%) remained relapse-free. Grade 2 tumors, a single metastatic site, and delivery of XRT were favorable predictors of relapse-free survival in univariate but not multivariate analyses. Inclusion of HDC, as described, in the multimodal treatment of stage IV breast cancer patients with minimal metastases is promising. These results warrant prospective randomized trials with an HDC-containing arm in this patient population.
Promising Findings Overshadowed by Questionable Outcomes Data and Study Design
Editorial Comment by Debasish Tripathy, MD
Key Words:
This report describes the outcome of consecutive patients with stage IV breast cancer who have been rendered free of all gross disease through the resection or radiation of limited sites of involvement. Given the small number of patients in this category, little information exists on their outcome using standard-dose or high-dose chemotherapy. In this study, all patients received induction chemotherapy followed by high-dose cisplatin, cyclophosphamide, and BCNU with autologous bone marrow or stem cell rescue. Certain procedures, such as local radiation, purging of marrow or stem cells via positive CD34 selection, and hormonal therapy, were not administered uniformly. This therapy is not without toxicities, with 10% treatment-related mortality and significant pulmonary toxicity.
While an impressive 3-year survival of 62.5% and relapse-free survival of 55% were noted, it is also known that this group of patients already has a better than average outcome. The authors correctly point out that this group of patients differs from those that achieve a complete response to standard-dose induction chemotherapy, which is indicative of chemosensitive tumor cells. Nevertheless, this group of patients had a similar outcome in the large, multicenter, randomized trial comparing 2 years of standard-dose CMF chemotherapy with high-dose chemotherapy (STAMP V) and bone marrow transplant (Stadtmauer et al, Proc Am Soc Clin Oncol. 1999;18:1A.). In this study, of the 45 patients who achieved a complete response to induction therapy, 42% were alive after high-dose therapy and 49% were alive after CMF therapy. Reports such as this raise a larger issue: Small, single-institution, phase II studies involving a very heterogeneous patient population are generally not able to provide accurate outcomes data and certainly cannot yield comparative data. Such studies are best reserved for determining toxicity or feasibility of novel therapies or procedures, such as purging.
The regimen used in this study (STAMP I) is the most widely used high-dose chemotherapy combination, but the study does not provide any information that would alter the standard of care or integrate a novel feature into a future comparative protocol. While the jury is still out on the role of high-dose chemotherapy in either early- or advanced-stage breast cancer, it is incumbent upon the research community to design and conduct comparative trials to improve outcome and advance the standard of care. Smaller pilot studies need to contain directives that incorporate new combinations, biological therapies, and other rationally based innovations that could eventually be followed by randomized trials.
Klijanienko J, Couturier J, Galut M, et al.
Cancer. 1999;87:312-318.
Fine-needle sampling, although a practical and noninvasive method of tissue acquisition, has rarely been used for HER-2/neu fluorescence in situ hybridization (FISH). To assess HER-2/neu gene amplification in mammary carcinoma, FISH signals on cytology and corresponding tissue biopsies were detected visually and measured by image analysis. The results were correlated with patient and tumor characteristics.
In situ HER-2/neu DNA probe hybridization was performed on 61 cytology specimens and on 47 corresponding frozen sections of breast carcinomas. Tumors were classified by visual evaluation as unamplified, moderately amplified, or highly amplified. Multiparametric image analysis was performed using the Discovery automated image analyzer (Becton Dickinson, Leiden, Netherlands). The integrated fluorescence ratio (IFR) was calculated for each sample as the integrated FISH fluorescence of the tumor cells divided by the integrated FISH fluorescence of internal control cells containing 2 spots. The percentage-positive nuclear area (PPN), calculated as the area of FISH fluorescence divided by the area of nuclear DNA fluorescence, and the PPR, ratio of the PPN of the tumor cells divided by the control cells, were also calculated for each sample.
Visual analysis yielded 46 unamplified and 15 (24.6%) amplified (7 moderately amplified and 8 highly amplified) tumors. Strong (P<.001) correlation between results on cytological and histological materials was obtained. The FISH spots on the cytological preparations were more easily visualized and scored than those on the corresponding tissue sections. Visual HER-2/neu signal scoring was strongly correlated with IFR (P=.0001) and PPR (P=.0001). Within the tumors classified as highly amplified by visual examination, quantitation of the degree of amplification fluorescence signal was possible using image analysis.
Cytologic specimens were a suitable and representative source of materials for detection and quantitation of HER-2/neu gene amplification by FISH and image analysis.
Tiny Samples Offer Potential for a Lot of Information
Editorial Comment by Britt-Marie Ljung, MD
Key words:
Fine needle samples (FNS)
Fine needle aspiration biopsies (FNAB)
Fluorescence in situ hybridization (FISH)
HER-2/neu oncogene
Fine needle samples (FNS) or fine needle aspiration biopsies (FNAB) are well established as diagnostic techniques for detecting benign and malignant lesions in breast and in many other body sites. FNS have a number of advantages over traditional open biopsies and core biopsies. They are minimally invasive since very thin 22- to 25-gauge needles are typically used. Thus FNS is very well tolerated by patients, and side effects are usually limited to an insignificant local bruise. Because cell preparations can easily be stained for preliminary examination at the bedside, material can be triaged for special studies as needed, minimizing sampling and obviating return visits for additional sampling. This also allows for a preliminary diagnosis in most cases at the time of sampling. Definitive diagnosis is typically made available within 24 hours. The accuracy in well-trained hands is similar to that of frozen section.
Traditionally, diagnosis of these samples has been based entirely upon morphology, ie, the visual appearance of the cells in the microscope interpreted by a pathologist. While morphology is still the basis for most diagnoses, molecular biology assays may now add additional prognostic information and guide treatment decisions.
One such technique is fluorescence in situ hybridization (FISH). This assay allows the evaluation of gene copy number in a nucleus by applying labeled probes directly to cells. FISH is used to determine amplification of the HER2/neu oncogene in breast cancer. Since intact individual nuclei must be visualized for FISH, fine needle samples consisting entirely of cells with intact nuclei are ideally suited for this test. Histological sections, on the other hand, have been cut into thin slices with a blade, resulting in many partial nuclei and are therefore less well suited for FISH analysis.
Other molecular biology techniques, such as comparative genomic hybridization or gene sequencing, analyze DNA extracted from cells. The number of cells obtained by FNS is sufficient for these analyses, since polymerase chain reaction (PCR) can be employed to amplify the DNA.
In the future, a combination of morphology and molecular biology assays will likely allow us to accurately diagnose and characterize a multitude of tumors to predict prognosis and response to specific therapies. Samples obtained by the minimally invasive technique of FNS are excellent material for these purposes.
Koss LG, Woyke S, Olszewski W. Aspiration Biopsy: Cytologic Interpretation and Histologic Bases. 2nd ed. Igaku-Shoin Medical Publishers Inc; 1992.
Dixon JM, Anderson TJ, Lamb J, et al. Fine needle aspiration cytology, in relationship to clinical examination and mammography in the diagnosis of a solid breast mass. Br J Surg. 1984;71:593-596.
Sterrett G, Harvey J, Parsons RW, et al. Breast cancer in western Australia in 1989,III: accuracy of FNA cytology in diagnosis. Aust NZ J Surg. 1994;64:745-749.
Giard RWM, Hermans J. The value of aspiration cytologic examination of the breast: a statistical review of the medical literature. Cancer. 1992;69:2104-2110.
Nieto Y, Cagnoni PJ, Shpall EJ, et al.
Clin Cancer Res. 1999;5:1731-1737.
The purpose of this study was to assess the efficacy of high-dose chemotherapy (HDC) with autologous stem cell transplant in stage IV breast cancer patients with minimal metastases. Eligible patients had (a) disease that could be resected en bloc and/or irradiated with curative intent using a single field and could, thus, be rendered as having no evidence of disease; and/or (b) < 5% bone marrow involvement. From September 1991 to August 1997, 40 consecutive patients were prospectively entered into the study. Pre-HDC local treatment consisted of surgery (n=31) and radiotherapy (XRT; n=3). All patients received HDC with cyclophosphamide, cisplatin, and 1,3-bis(2-chloroethyl)-1-nitrosourea and autologous stem cell transplant, with or without CD34 selection. Following HDC, 22 patients received XRT. Four patients died of treatment-related complications. Eighteen patients developed grade 3 nonhematological toxicities (15 lung, 2 cardiomyopathy, and 1 optic neuritis), which resolved with therapy. Within a median follow-up of 49 (15-91) months, 14 patients had relapsed. Twenty-five patients (62.5%) were alive, and 22 patients (55%) were alive and free of disease. Median event-free and overall survivals were 43 and 77 months, respectively. In the subset of patients with one metastatic site, 17 of 24 (68%) remained relapse-free. Grade 2 tumors, a single metastatic site, and delivery of XRT were favorable predictors of relapse-free survival in univariate but not multivariate analyses. Inclusion of HDC, as described, in the multimodal treatment of stage IV breast cancer patients with minimal metastases is promising. These results warrant prospective randomized trials with an HDC-containing arm in this patient population.
Promising Findings Overshadowed by Questionable Outcomes Data and Study Design
Editorial Comment by Debasish Tripathy, MD
Key Words:
Stage IV breast cancer
Induction chemotherapy
Bone marrow transplant
Data, outcomes, comparative
This report describes the outcome of consecutive patients with stage IV breast cancer who have been rendered free of all gross disease through the resection or radiation of limited sites of involvement. Given the small number of patients in this category, little information exists on their outcome using standard-dose or high-dose chemotherapy. In this study, all patients received induction chemotherapy followed by high-dose cisplatin, cyclophosphamide, and BCNU with autologous bone marrow or stem cell rescue. Certain procedures, such as local radiation, purging of marrow or stem cells via positive CD34 selection, and hormonal therapy, were not administered uniformly. This therapy is not without toxicities, with 10% treatment-related mortality and significant pulmonary toxicity.
While an impressive 3-year survival of 62.5% and relapse-free survival of 55% were noted, it is also known that this group of patients already has a better than average outcome. The authors correctly point out that this group of patients differs from those that achieve a complete response to standard-dose induction chemotherapy, which is indicative of chemosensitive tumor cells. Nevertheless, this group of patients had a similar outcome in the large, multicenter, randomized trial comparing 2 years of standard-dose CMF chemotherapy with high-dose chemotherapy (STAMP V) and bone marrow transplant (Stadtmauer et al, Proc Am Soc Clin Oncol. 1999;18:1A.). In this study, of the 45 patients who achieved a complete response to induction therapy, 42% were alive after high-dose therapy and 49% were alive after CMF therapy. Reports such as this raise a larger issue: Small, single-institution, phase II studies involving a very heterogeneous patient population are generally not able to provide accurate outcomes data and certainly cannot yield comparative data. Such studies are best reserved for determining toxicity or feasibility of novel therapies or procedures, such as purging.
The regimen used in this study (STAMP I) is the most widely used high-dose chemotherapy combination, but the study does not provide any information that would alter the standard of care or integrate a novel feature into a future comparative protocol. While the jury is still out on the role of high-dose chemotherapy in either early- or advanced-stage breast cancer, it is incumbent upon the research community to design and conduct comparative trials to improve outcome and advance the standard of care. Smaller pilot studies need to contain directives that incorporate new combinations, biological therapies, and other rationally based innovations that could eventually be followed by randomized trials.
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