Bruce C. Turner

Outcome of conservatively managed early-onset breast cancer by BRCA1/2 status.

BACKGROUND Management of early-stage breast cancer in young women with mutations in BRCA1 or BRCA2 remains controversial. This study assessed the long-term risks of ipsilateral and contralateral breast cancer in a cohort of young women who underwent breast-conserving surgery followed by radiotherapy. METHODS Between 1975 and 1998, 290 women with breast cancer diagnosed at age 42 years or younger underwent lumpectomy followed by radiotherapy at our hospital. We recruited 127 of these women for complete sequencing of BRCA1 and BRCA2. Demographic, clinical, pathological, and outcome data were recorded. The primary endpoints were rates of ipsilateral and contralateral breast cancer, in relation to germline BRCA1/2 status. FINDINGS 105 women were classified as having sporadic disease (94 with wild-type or known polymorphisms and 11 with variants of unclear significance) and 22 as having genetic predisposition (deleterious mutations in BRCA1 [15] or BRCA2 [seven]). At 12 years of follow-up, the genetic group had significantly higher rates of ipsilateral (49% vs 21%, p=0.007) and contralateral events (42% vs 9%, p=0.001) than the sporadic group. The majority of events were classified as second primary tumours. No patient in the genetic group had undergone oophorectomy or was taking prophylactic agents such as tamoxifen. INTERPRETATION Patients with germline mutations in BRCA1 or BRCA2 have a high risk of developing late ipsilateral and contralateral second primary tumours. With breast-conserving therapy, chemoprophylaxis or other interventions to reduce the rate of second cancers may be valuable. Alternatively, bilateral mastectomy may be considered, to minimise the risk of second tumours in the breasts.

BRCA1/BRCA2 Germline Mutations in Locally Recurrent Breast Cancer Patients After Lumpectomy and Radiation Therapy: Implications for Breast-Conserving Management in Patients With BRCA1/BRCA2 Mutations

PURPOSE Breast cancer patients treated conservatively with lumpectomy and radiation therapy (LRT) have an estimated lifetime risk of local relapse (ipsilateral breast tumor recurrence [IBTR]) of 10% to 15%. For breast cancer patients carrying BRCA1 or BRCA2 (BRCA1/2) mutations, the outcome of treatment with LRT with respect to IBTR has not been determined. In this study, we estimate the frequency of BRCA1/2 mutations in a study of breast cancer patients with IBTR treated with LRT. PATIENTS AND METHODS Between 1973 and 1994, there were 52 breast cancer patients treated with LRT who developed an IBTR within the prior irradiated breast and who were willing to participate in the current study. From our database, we also identified 52 control breast cancer patients treated with LRT without IBTR. The control patients were individually matched to the index cases with respect to multiple clinical and pathologic parameters. Lymphocyte DNA specimens from all 52 locally recurrent patients and 15 of the matched control patients under age 40 were used as templates for polymerase chain reaction amplification and dye-primer sequencing of exons 2 to 24 of BRCA1, exons 2 to 27 of BRCA2, and flanking intron sequences. RESULTS After LRT, eight (15%) of 52 breast cancer patients had IBTR with deleterious BRCA1/2 mutations. By age, there were six (40%) of 15 patients with IBTR under age 40 with BRCA1/2 mutations, one (9.0%) of 11 between ages 40 and 49, and one (3.8%) of 26 older than age 49. In comparison to the six (40%) of 15 of patients under age 40 with IBTR found to have BRCA1/2 mutations, only one (6.6%) of 15 matched control patients without IBTR and had a BRCA1/2 mutation (P =.03). The median time to IBTR for patients with BRCA1/2 mutations was 7.8 years compared with 4.7 years for patients without BRCA1/2 mutations (P =.03). By clinical and histologic criteria, these relapses represented second primary tumors developing in the conservatively treated breast. All patients with BRCA1/2 mutations and IBTR underwent successful surgical salvage mastectomy at the time of IBTR and remain alive without evidence of local or systemic progression of disease. CONCLUSION In this study, we found an elevated frequency of deleterious BRCA1/2 mutations in breast cancer patients treated with LRT who developed late IBTR. The relatively long time to IBTR, as well as the histologic and clinical criteria, suggests that these recurrent cancers actually represent new primary breast cancers. Early onset breast cancer patients experiencing IBTR have a disproportionately high frequency of deleterious BRCA1/2 mutations. This information may be helpful in guiding management in BRCA1 or BRCA2 patients considering breast-conserving therapy.

Effective Treatment of Stage I Uterine Papillary Serous Carcinoma with High Dose-Rate Vaginal Apex Radiation (192Ir) and Chemotherapy

PURPOSE Uterine papillary serous carcinoma (UPSC) is a morphologically distinct variant of endometrial carcinoma that is associated with a poor prognosis, high recurrence rate, frequent clinical understaging, and poor response to salvage treatment. We retrospectively analyzed local control, actuarial overall survival (OS), actuarial disease-free survival (DFS), salvage rate, and complications for patients with Federation International of Gynecology and Obstetrics (FIGO) (1988) Stage I UPSC. METHODS AND MATERIALS This retrospective analysis describes 38 patients with FIGO Stage I UPSC who were treated with the combinations of radiation therapy, chemotherapy, total abdominal hysterectomy, and bilateral salpingo-oophorectomy (TAH/BSO), with or without a surgical staging procedure. Twenty of 38 patients were treated with a combination of low dose-rate (LDR) uterine/vaginal brachytherapy using 226Ra or 137Cs and conventional whole-abdomen radiation therapy (WART) or whole-pelvic radiation therapy (WPRT). Of 20 patients (10%) in this treatment group, 2 received cisplatin chemotherapy. Eighteen patients were treated with high dose-rate (HDR) vaginal apex brachytherapy using 192Ir with an afterloading device and cisplatin, doxorubicin, and cyclophosphamide (CAP) chemotherapy (5 of 18 patients). Only 6 of 20 UPSC patients treated with combination LDR uterine/vaginal brachytherapy and conventional external beam radiotherapy underwent complete surgical staging, consisting of TAH/BSO, pelvic/para-aortic lymph node sampling, omentectomy, and peritoneal fluid analysis, compared to 15 of 18 patients treated with HDR vaginal apex brachytherapy. RESULTS The 5-year actuarial OS for patients with complete surgical staging and adjuvant radiation/chemotherapy treatment was 100% vs. 61% for patients without complete staging (p = 0.002). The 5-year actuarial OS for all Stage I UPSC patients treated with postoperative HDR vaginal apex brachytherapy and systemic chemotherapy was 94% (18 patients). The 5-year actuarial OS for Stage I UPSC patients treated with HDR vaginal apex brachytherapy and chemotherapy who underwent complete surgical staging was 100% (15 patients). The 5-year actuarial OS for the 20 Stage I UPSC patients treated with combinations of pre- and postoperative LDR brachytherapy and postop WART was 65%. None of the 6 surgically staged UPSC patients treated with LDR radiation and WART/WPRT developed recurrent disease. For patients with FIGO Stage IA, IB, and IC UPSC who underwent complete surgical staging, the 5-year actuarial DFS by depth of myometrial invasion was 100, 71, and 40%, respectively (p = 0.006). The overall salvage rate for local and distant recurrence was 0%. Complications following HDR vaginal apex brachytherapy included only Radiation Therapy Oncology Group (RTOG) grade 1 and 2 toxicity in 16% of patients. However, complications from patients treated with WART/WPRT, and/or LDR brachytherapy, included RTOG grade 3 and 4 toxicity in 15% of patients. CONCLUSION Patients with UPSC should undergo complete surgical staging, and completely surgically staged FIGO Stage I UPSC patients can be effectively and safely treated with HDR vaginal apex brachytherapy and chemotherapy. Both OS and DFS of patients with UPSC are dependent on depth of myometrial invasion. The salvage rate for both local and distant UPSC recurrences is extremely poor. Complications from HDR vaginal apex brachytherapy were minimal.

Mutant p53 protein overexpression in women with ipsilateral breast tumor recurrence following lumpectomy and radiation therapy.

The p53 tumor suppressor gene encodes a nuclear phosphoprotein that is thought to be important to cell cycle regulation and DNA repair and that also may regulate induction of apoptosis by ionizing radiation. Somatic p53 gene mutations occur in 30–50% of breast carcinomas and are associated with poor prognosis. Mutations in the p53 gene result in prolonged stability of the protein that can be detected by immunohistochemical techniques. In a matched case–control study of breast carcinoma patients with ipsilateral breast tumor recurrence (IBTR) following lumpectomy and radiation therapy, the authors investigated the frequency and prognostic significance of somatic p53 mutations as well as the clinical characteristics of patients with these mutations.

Complication rates in patients with negative axillary nodes 10 years after local breast radiotherapy after either sentinel lymph node dissection or axillary clearance.

Background:We assess complication rates in node negative breast cancer patients treated with breast radiotherapy (RT) only after sentinel lymph node dissection (SLND) or axillary lymph node dissection (ALND). Materials and Methods:Between 1995 and 2001, 226 women with AJCC stage I-II breast cancer were treated with lumpectomy, either SLND or SLND+ALND, and had available toxicities in follow-up: 111/136 (82%) and 115/129 (89%) in SLND and ALND groups, respectively. RT targeted the breast to median dose of 48.2 Gy (range, 46.0 to 50.4 Gy) without axillary RT. Chi-square tests compared complication rates of 2 groups for axillary web syndrome (AWS), seroma, wound infection, decreased range of motion of the ipsilateral shoulder, paresthesia, and lymphedema. Results:Median follow-up was 9.9 years (range, 8.3-15.3 y). Median number of nodes assessed was 2 (range, 1-5) in SLND and 18 (range, 7-36) in ALND (P < 0.0001). Acute complications occurred during the first 2 years and were AWS, seroma, and wound infection. Incidences of seroma 5/111 (4.5%) in SLND and 16/115 (13.9%) in ALND (P < 0.02, respectively) and wound infection 3/111 (2.7%) in SLND and 10/115 (8.7%) in ALND (P < 0.05, respectively) differed significantly. AWS was not statistically different between the groups. At 10 years, the only chronic complications decreased were range of motion of the shoulder 46/111 (41.4%) in SLND and 92/115 (80.0%) in ALND (P < 0.0001), paresthesia 12/111 (10.8%) in SLND and 39/115 (33.9%) in ALND (P < 0.0001), and lymphedema assessed by patients 10/111 (10.0%) in SLND and 39/115 (33.9%) in ALND (P < 0.0001). Chronic lymphedema, assessed by clinicians, occurred in 6/111 (5.4%) in SLND and 21/115 (18.3%) in ALND cohorts, respectively (P < 0.0001). Conclusions:Our mature findings support that in patients with negative axillary nodal status SLND and breast RT provide excellent long-term cure rates while avoiding morbidities associated with ALND or addition of axillary RT field.

Cyclin D1 expression and early breast cancer recurrence following lumpectomy and radiation.

PURPOSE The purpose of this study was to determine the prognostic significance of cyclin D1 (cycD1) levels in ipsilateral breast tumor recurrence (IBTR) following lumpectomy and radiation therapy. METHODS AND MATERIALS A total of 98 patients (49 patients with IBTR and 49 matched cases without IBTR) selected from our conservatively treated breast cancer population served as the patient population for the current study. All patients were treated with lumpectomy followed by radiation therapy to the intact breast to a total median dose of 64 Gy. The patients were followed in our clinic with a median follow-up of 13 years. Immunohistochemical analysis of cycD1 in these 98 early-stage breast cancer patients was performed using a polyclonal antibody generated against the human cycD1 protein. All clinical, pathologic, and molecular variables were entered into a computerized data base for statistical analysis. RESULTS Low levels of immunohistochemically detected cycD1 protein correlated with IBTR (p = 0.001), but there was no association between cycD1 protein levels and metastatic disease, axillary lymph node involvement, distant disease-free survival (DDFS), and overall survival (OS). Subgroup analysis revealed that for early breast tumor relapses (within 4 years of initial breast tumor diagnosis), low levels of cycD1 were associated with IBTR (p = 0.004), but cycD1 expression was not prognostic for IBTR from breast cancer patients with late relapses (p = NS). CONCLUSION These studies provide in vivo evidence for the prognostic and biologic significance of cycD1 expression in determining response to radiation therapy in breast cancer patients.

BAG-1 Immunostaining and Survival in Early Breast Cancer

To the Editor: We read with interest the article by Turner et al1 in the February 15, 2001, issue describing the association between increased levels of cytosolic, but not nuclear, BAG-1 immunostaining and long-term survival in early breast cancer. There is, however, some difficulty in determining the precise relationship between BAG-1 expression and the biology of breast cancer. A previous report by this group using a monoclonal antibody described a significant positive correlation between high levels of nuclear BAG-1 immunostaining and overall survival, whereas a report from a different group described inferior survival in patients with high levels of nuclear BAG-1 staining using a polyclonal antibody. From the examples of immunostaining presented in the two articles from Turner and co-workers, it seems that at least some of the same patients were included in the two series with discrepant results. It would be helpful to know the degree of overlap between the two cohorts and whether any potential selection bias may have influenced the results. In particular, the low proportion of estrogen receptor–positive cases (41% as against an expected level of 70%) suggests that the sample described may not be wholly representative of an early-stage breast cancer population. Tumor grade was not included in the table of patient characteristics, and as correlations have been found between tumor differentiation and BAG-1 status by at least one group, it would be instructive to know if BAG-1 was predictive of outcome independent of tumor grade. It is also possible that the discrepant results relate to differences in methodology, such as antigen retrieval techniques, which were not described in the more recent article. It is clear that further work is required to properly describe the potential role of BAG-1 expression as a biologic variable, and it would be helpful to other investigators to have these critical details.

Homozygous mutations in the Fhit gene results in resistance to ionizing radiation and inhibition of apoptosis

Abstract Purpose: The Fhit gene was identified because it represents the most active constitutive chromosome fragile site and has functions often associated with a tumor suppressor gene. Mutations in the Fhit locus have been identified in many cancer-derived cell lines, primary human tumors including lung, head and neck, colon, breast, and esophagus, and are associated with tobacco-induced lung cancers. In this study, we examined the cellular response of mouse epithelial cells with complete loss of Fhit to therapeutic doses of ionizing radiation and the prognostic importance of Fhit protein in early stage breast cancer patients treated with breast conserving therapy. Materials and Methods: Mouse epithelial cell lines containing either homozygous mutant Fhit−/− or wild-type Fhit+/+ were derived from mice (C57BL/6J X 129/SvJ) with either wild-type or inactivated Fhit gene. Clonogenic cell survival assays were carried out on subconfluent cells in logarithmic growth using a 137Cs irradiator and survival curves were plotted as the log of the surviving cells versus dose and corrected for cloning efficiency. Apoptosis following ionizing radiation was determined by flow cytometry using the Annexin-V FITC kit and DAPI staining. Paraffin-embedded breast tumor blocks were obtained from 42 women with local breast tumor recurrence and 42 matched breast cancer patients without local cancer relapse treated with breast conserving therapy and stained with a 1:4000 dilution of polyclonal antibody to the Fhit protein and scored based on both intensity and distribution of Fhit staining within the invasive breast cancer component. Results: Treatment of Fhit−/− mouse epithelial cells with single fraction doses of ionizing radiation including 2, 4, 6, and 10 Gy result in 4-6 fold increase in cellular survival compared with isogenic parental cells from Fhit+/+ mice. Fhit−/− epithelial cells displayed 3-5 fold lower levels of apoptosis in response to both low and high doses of ionizing radiation compared to parental mammalian cells expressing wild-type Fhit protein. Finally, we demonstrated that breast tumors from breast cancer patients with local breast tumor recurrences following breast conserving therapy more often lacked immunhistochemical detection of Fhit protein compared to tumors from breast cancer patients without local breast cancer recurrence (p=0.02). Interestingly, the adjacent benign regions of these sections contained similar levels of Fhit protein expression suggesting that a somatic alteration is critical in the clinical resistance to ionizing radiation observed in these patients. Apoptotic pathways regulating the aberrant response to DNA damage-induced apoptosis in Fhit knock-out cells are currently being studied. Conclusion: Mouse epithelial cells containing homozygous Fhit mutations are resistant to single fraction low and high dose ionizing radiation with decreased levels of radiation-induced apoptotic cell death. Breast tumors from women with local breast cancer recurrence following breast conserving therapy have low levels of Fhit protein. These findings may have important biologic and treatment implications including those for cancer patients with tumors having mutations in Fhit and suggest that treatment with ionizing radiation in these patients may not result in optimal responses.

Fhit loss in familial breast cancer: is loss of DNA repair function linked to alterations at chromosome fragile sites?

The FHIT gene at 3p14.2 encompasses the common fragile site, FRA3B, and is involved in frequent chromosome rearrangements in human cancers. Fhit protein expression is reduced or lost in the majority of esophageal, lung, gastric, cervical, pancreatic, kidney and bladder cancers, and a large fraction of other cancers. Fhit expression in sporadic breast cancers has been studied by several groups, and reported to show alteration in expression of Fhit in 30-50%. Because familial breast cancers were reported to show a higher frequency of LOH at 3p14.2 than sporadic breast cancers, we were interested in whether common fragile regions might be targets for repair by the Brca1 and Brca2 proteins, and might thus be a downstream target in BRCA1- and BRCA2-induced familial breast tumors. We studied a panel of Brca2-deficient breast tumors and showed that only 18% expressed Fhit strongly, compared with 48% of sporadic tumors (P = 0.002). Very recently we completed a similar study of BRCA1 familial tumors, and observed that only 9% of these tumors showed strong expression versus more than 40% of sporadic breast tumors (P <0.001, odds ratio 0.09). We conclude that loss of BRCA1 and BRCA2 functions affect stability of the FHIT/FRA3B locus and possibly other fragile loci.