Thomas A. Bonfiglio

The Expression Patterns of ER, PR, HER2, CK5/6, EGFR, Ki-67 and AR by Immunohistochemical Analysis in Breast Cancer Cell Lines

Curcumin is a compound with anti-tumor effects in a tolerable dose. A recent paper by Rowe et al described that curcumin induced DNA damage in triple negative breast cancer cells and regulated BRCA1 protein expression and modification.1 Related research and potential use of curcumin will be discussed in this article.The molecular classification for breast carcinomas has been used in clinical studies with a simple surrogate panel of immunohistochemistry (IHC) markers. The objective of this current project was to study the molecular classification of commonly used breast cancer cell lines by IHC analysis. Seventeen breast cancer cell lines were harvested, fixed in formalin and made into cell blocks. IHC analyses were performed on each cell block with antibodies to estrogen receptor (ER), progesterone receptor (PR), HER2, EGFR, CK5/6, Ki-67 and androgen receptor (AR). Among the 17 cell lines, MCF-7 and ZR-75-1 fell to Luminal A subtype; BT-474 to Luminal B subtype; SKBR-3, MDA-MD-435 and AU 565 to HER2 over-expression subtype; MDA-MB-231, MCF-12A, HBL 101, HS 598 T, MCF-10A, MCF-10F, BT-20, 468 and BT-483 to basal subtype. MDA-MB-453 belonged to Unclassified subtype. Since each subtype defined by this IHC-based molecular classification does show a distinct clinical outcome, attention should be paid when choosing a cell line for any study.I would like to welcome breast cancer research community to the first editorial of our newest journal “Breast Cancer: Basic and Clinical Research”. In pursuit of breast cancer culprits, we have come a long way since the early 90’s when the first breast cancer susceptibility gene BRCA1 was mapped and cloned. In the past few years, several new loci associated with the various degree of breast cancer risk have been identified using “Candidate Gene Association Study (CGAS) and Genome-Wide Association Study (GWAS)” approaches. This editorial is meant to quickly glance over recent findings of these population-based association studies.Among women, the most prevalent type of cancer is breast cancer, affecting 1 out of every 8 women in the United States; in Puerto Rico, 70 out of every 100,000 will develop some type of breast cancer. Therefore, a better understand of the potential risk factors for breast cancer could lead to the development of early detection tools. A gene that has been proposed as a risk factor in several populations around the world is Apolipoprotein E (apoE). ApoE functions as a mechanism of transport for lipoproteins and cholesterol throughout the body, with 3 main isoforms present in humans (apoE2, apoE3, and apoE4). Whether or not apoE4 is a risk factor for breast cancer remains controversial. Previous studies have either included test subjects of all ages (20–80) or have focused on late-onset (after age 50) breast cancer; none has concentrated specifically on early-onset (aged 50 and younger) breast cancer. The objectives of this study was to examine (in a Puerto Rican population) the differences in the relative frequency of occurrence of apoE4 in non-breast cancer versus breast cancer patients and to examine, as well, the potential differences of same in early- versus late-onset patients. We found an increased frequency of apoE4 (odds ratio 2.15) only in early-onset breast cancer survivors, which is similar to the findings of those studies that combined or adjusted for age as well as for an association between apoE4 and decreased tumor size. ApoE is also a potential risk factor for long-term cognitive effects after chemotherapy and affects response to hormone replacement. Our data supports the theory that knowing the apoE genotype of women who are at risk of developing breast cancer may be beneficial, as such knowledge would aid in the prediction of tumor size and the development of treatment regimens.The provasopressin protein (proAVP) is expressed by invasive breast cancer and non-invasive breast cancer, or ductal carcinoma in situ (DCIS). Here we demonstrate the ability of the monoclonal antibody MAG1 directed against the C-terminal end of proAVP to identify proAVP in all cases examined of human invasive cancer and DCIS (35 and 26, respectively). Tissues were chosen to represent a relevant variation in tumor type, grade, patient age, and menopausal status. By comparison, there was 65% positive staining for estrogen receptor, 61% for progesterone receptor, 67% for nuclear p53, and 39% for c-Erb-B2 with the invasive breast cancer sections. Reaction with the normal tissue types examined (67) was restricted to the vasopressinergic magnocellular neurons of the hypothalamus, where provasopressin is normally produced, and the posterior pituitary, where these neurons terminate. The breast epithelial tissue sections on the tissue microarray did not react with MAG1. Previously, we demonstrated that polyclonal antibodies to proAVP detected that protein in all breast cancer samples examined, but there was no reaction with breast tissue containing fibrocystic disease. The results presented here not only expand upon those earlier results, but they also demonstrate the specificity and effectiveness of what may be considered a more clinically-relevant agent. Thus, proAVP appears to be an attractive target for the detection of invasive breast cancer and DCIS, and these results suggest that MAG1 may be a beneficial tool for use in the development of such strategies.

Uterine sarcomas: natural history, treatment and prognosis.

Seventy‐three documented cases of uterine sarcoma were treated at the University of Rochester Strong Memorial Hospital from 1955 to 1975. Thirty‐three patients (45%) were treated with surgery only [S], 31 (43%) with surgery and radiation [S + R], and 9 (12%) with radiation alone [R]. A review of the literature with over 900 cases was also performed. Several important issues regarding these rare tumors are addressed, such as the prognosis of the several histologic variants, the role of radiation therapy in their management and what perhaps may constitute a comprehensive therapeutic approach. These tumors are characterized by local aggressiveness and early widespread dissemination. There are three main histologic varieties: mixed mesodermal sarcoma (MMS), leiomyosarcoma (LMS) and endometrial stromal sarcoma (ESS). Of the three, MMS was the most common, seen in 60% of the cases; LMS occurred in younger patients and tended to be localized to the uterine corpus (Stage I) in 80% of the instances. Tumor extent at diagnosis was the main prognosticator for survival in uterine sarcomas; patients with Stage I tumors had a significantly lower incidence of recurrences, as well as a better survival than patients with more advanced tumors. Stage‐by‐stage, there were no significant differences in survival among the pathologic variants. To ensure adequate staging, a surgical procedure is recommended first whenever possible. Adjuvant radiation therapy significantly improved disease controlability in the pelvis, although it may not have dramatically affected the final outcome. In addition to pelvic irradiation, some form of systemic therapy should be administered to decrease distant metastases.

Distribution of disease at autopsy in 100 women with ovarian cancer

Clinical and morphologic factors that affected the distribution of disease are described in 100 cases of ovarian cancer at autopsy. In addition to the expected pattern of pelvic and abdominal peritoneal spread, extensive visceral parenchymal metastases were seen: liver parenchyma (45%), lung parenchyma (39%), small and large intestinal wall (52% and 55%), lymph nodes (70%), pancreas (21%), ureter (24%), bone (11%), and brain (6%). Liver parenchymal metastases replaced more than one third of the liver in 25% of cases, whereas lung metastases always involved less than one third of the lungs. When intestinal wall invasion was seen, bowel obstruction was present more often (71%) than when only intestinal serosa was involved (30%). Lymphatic invasion was predictive of lymph node, small intestinal wall, pancreatic, and liver as well as lung parenchymal metastases. Blood vessel invasion was predictive of pancreatic and ureteral metastases. Clinical stage I at diagnosis was associated with high incidences of liver parenchymal (56%), lymph node (56%), lung parenchymal (44%), large intestinal wall (33%), and bone (33%) metastases. Thus, ovarian cancer has parenchymal metastases similar to other carcinomas in addition to its peritoneal spread. Lymphatic and blood vessel invasion is predictive of such involvement. Intestinal wall invasion predicts bowel obstruction.

Coronary arteritis, occlusion, and myocardial infarction due to lupus erythematosus

Abstract Four women with clinically documented lupus erythematosus and symptomatic coronary artery disease are presented. One, age 16, had severe coronary arteritis, thrombosis, and an acute myocardial infarct documented by postmortem examination. Three other women, ages 27, 33, and 38, had angina pectoris. The 33-year-old patient also had had a probable myocardial infarct, and has persistent, complete left bundle branch block. Coronary artery angiograms demonstrated severe focal coronary artery stenosis and multiple obstructions in all cases. Coronary arteritis may be a serious and sometimes fatal consequence of lupus erythematosus.

Uterine sarcomas. Analysis of failures with special emphasis on the use of adjuvant radiation therapy

There were 47 failures among 73 verified cases of uterine sarcoma reported at the University of Rochester Tumor Registry from 1955 to 1975; they constitute the subject of this report. Of 33 patients initially treated with surgery only [S], 19 patients (58%) failed; 20 of 31 patients (65%) treated with surgery and radiation [S + R] failed; 8 of 9 patients (89%) treated by radiation alone [R] failed. According to pathology, failures occurred in 33 of 44 patients (75%) with mixed mesodermal sarcomas (MMS), 7 of 20 patients (35%) with leiomyosarcoma (LMS), 4 of 6 patients with endometrial stromal sarcomas (ESS), and 3 of 3 patients with other types of sarcoma. Once corrected by stage, there were no significant differences in failure rates, spread patterns or survival among these main histologic variants. Twenty of 41 patients (56%) with Stage I tumors failed with an average failure time of 32 months. Twenty‐seven of 32 patients (84%) with Stages II, III, and IV tumor failed; their average failure time was only 9 months. The mean failure time for both the patients treated with [S] and [S + R] was 22 months; for patients treated by [R] it was 3 months. Isolated pelvic failures constituted only 4% of all failures, failures both in the pelvis and in distant sites, 49%, and distant metastases, 47%. There was a marked decrease in pelvic failures in patients treated with [S + R] when compared to those who received [S]. Adjuvant radiation proved to increase tumor control in the pelvis but did not influence the final outcome because over 90% of all failures developed distant spread outside the pelvis. The most common distant failures were in the upper abdomen (mainly omentum and peritoneum) and in the lungs. Lung metastases alone was the only site of failure in 16% of the instances. A comprehensive treatment approach based on the spread and failure patterns will be proposed.

THE MANAGEMENT OF CLINICAL STAGE I ENDOMETRIAL CARCINOMA

A retrospective analyses of 307 cases with clinical Stage I endometrial carcinoma was done in an attempt to determine the role of radiation therapy in the optimal treatment of this disease. A review of the modern literature with over 9000 cases served as a useful tool to corroborate inferences and conclusions. The present series has 155 patients (51%) treated with preoperative megavoltage external pelvic radiation with a variation in doses of less than 6%. Five‐year survival estimates (79%‐83%) in clinical Stage I endometrial carcinoma are similar among the several main treatment combination that are employed; they become a useless parameter for any comparison. The pelvic failure rate constitutes a more useful guideline in assessing the most adequate therapy. The pathologic grade of the tumor is the main prognosticator in endometrial carcinoma. Intimately related to the tumor grade is the depth of myometrial invasion of the carcinoma. The size of the uterus and/or its cavity carry less prognostic significance than traditionally thought. For grade I lesions, there is little error in diagnosis, few pelvic failures and excellent survival (96%); they could be approached with initial surgery and postoperative radiation reserved for selected patients. For grade 2 tumors, the error in diagnosis and the failure rate increases with an overall survival of 87%. For grade 3 tumors, the error in diagnosis and failure rates are quite high with a 5 year survival of only 70%. Preoperative radiation, especially external beam therapy, is suggested for grades 2 and 3 Stage I tumors. The use of this treatment modality yields only 3% pelvic failure and an overall 5 year survival of almost 90%.

Endometrial carcinoma: Analysis of failures with special emphasis on the use of initial preoperative external pelvic radiation

Abstract This paper analyzes in detail a total of 75 failures occurring among 364 patients with endometrial carcinoma. Approximately one half of all patients received preoperative external pelvic radiation for initial treatment. One-half of all failures occurred in the pelvis, the other half in distant sites. Failures in endometrial carcinoma are associated with a higher pathologic grade and clinical stage. An increase in grade and/or stage is associated with a higher incidence and depth of myometrial invasion and pelvic nodal involvement. The use of preoperative external pelvic radiation has eliminated vaginal recurrences in clinical Stages I and If and has reduced the overall incidence of pelvic failures to only 3%. Approximately one-third of patients with failures localized to the pelvis after initial surgery can be salvaged by the use of external pelvic radiation. Patients with distant metastases benefit significantly from therapy other than supportive measures.

Stage III ovarian tumors of low malignant potential treated with cisplatin combination therapy (A Gynecologic Oncology Group Study)

By serendipity we have had the opportunity to evaluate cis-platin-based chemotherapy in ovarian tumors of low malignant potential (LMP). Optimal (less than 1 cm residual disease) FIGO stage III ovarian carcinomas were randomly assigned to treatment with cisplatin plus cyclophosphamide with or without doxorubicin on a prospective Gynecologic Oncology Group Study. On review by the Gynecologic Oncology Group Pathology Committee, 32 of these cases were determined to represent low malignant potential tumors. Mean age of patients with these lesions was 48 years (range, 25-75 years). After initial cytoreduction, 19 patients had residual disease less than 1 cm and 13 had no residual. Twenty (62.5%) received cisplatin plus cyclophosphamide and 12 cisplatin, cyclophosphamide, and doxorubicin chemotherapy; 75% of patients received six or more courses. Second-look surgery was done in 15 cases; only six were negative. However, with a median follow-up of 31.7 months (range, 1-75), only 1 patient has died; no cancer was found at autopsy. The remaining patients are alive without clinical evidence of disease at a median of 30 months. The need for adjunctive therapy in patients with advanced LMP tumors remains speculative.

Identification of Novel Methylation Markers in Cervical Cancer Using Restriction Landmark Genomic Scanning

Aberrant methylation of CpG islands in gene promoters often represents an early clonal event in carcinogenesis. Accordingly, defining methylation profiles may be useful for developing marker panels for early detection or predicting the risk of cancer precursors. To identify specific genes frequently methylated in cervical cancer, we conducted methylation profiling of 20 primary human cervical cancers using NotI-based restriction landmark genomic scanning (RLGS). Of 2,172 RLGS fragments analyzed (average, 1,753 CpG islands per patient), 186 RLGS fragments were lost in at least one tumor and 40 were lost in three or more. Methylation was identified in 19 (95%) of 20 tumor samples compared with normal DNA. Bisulfite sequencing was conducted to confirm RLGS results. Of the confirmed markers frequently methylated, we developed Methylight assays for two corresponding genes, nucleolar protein 4 (NOL4), and lipoma HMGIC fusion partner-like protein 4 (LHFPL4), which were methylated in 85% and 55% of cancers, respectively. Using these assays, we further confirmed frequent CpG island methylation in the original cancers and in another independent series of 15 cervical cancers. We also showed methylation at a reduced frequency in a set of carefully reviewed cytology specimens demonstrating cells exfoliated from cancer precursor lesions. In summary, we identified, for the first time, NOL4 and LHFPL4 as novel methylation targets specific for cervical cancer. Inclusion of NOL4 and LHFPL4 in evaluating methylation panels for early detection, risk prediction, and etiologic research on cervical cancer is warranted.

Granulomatous pseudotumors in total joint replacement

Fourteen patients (15 joints) developed a foreign body reaction to methylmethacrylate, polyethylene, or metal adjacent to a total joint implan, a condition we would like to term granulomatous pseudotumors. There were eight male and six female patients. Their average age was 61 years. The hip was involved in 14 joints (femoral component 11 times, acetabulum 7, and great trochanter once). One patient presented with granulomatous pseudotumors of the knee. The principal findings included increasing pain and radiographic evidence of loosening occurring on average 2.7 years following the implant. This was followed by a characteristic and gradually developing radiographic pattern of discrete rounded lucencies. These developed into large ovoid lytic areas, destroying both methylmethacrylate and bone. Histologically, the appearances were characterized by histiocytic infiltration and the presence of multiple foreign body giant cells. Foreign material was identified in 9 of 11 cases. The pathogenesis is unknown but appears related to micromovement or loosening of the implant.