Uninvolved Breast Tissue Research Articles

Leveraging intelligent optimization for automated, cardiac-sparing accelerated partial breast treatment planning.

Accelerated partial breast irradiation (APBI) yields similar rates of recurrence and cosmetic outcomes as compared to whole breast radiation therapy (RT) when patients and treatment techniques are appropriately selected. APBI combined with stereotactic body radiation therapy (SBRT) is a promising technique for precisely delivering high levels of radiation while avoiding uninvolved breast tissue. Here we investigate the feasibility of automatically generating high quality APBI plans in the Ethos adaptive workspace with a specific emphasis on sparing the heart. Nine patients (10 target volumes) were utilized to iteratively tune an Ethos APBI planning template for automatic plan generation. Twenty patients previously treated on a TrueBeam Edge accelerator were then automatically replanned using this template without manual intervention or reoptimization. The unbiased validation cohort Ethos plans were benchmarked via adherence to planning objectives, a comparison of DVH and quality indices against the clinical Edge plans, and qualitative reviews by two board-certified radiation oncologists. 85% (17/20) of automated validation cohort plans met all planning objectives; three plans did not achieve the contralateral lung V1.5Gy objective, but all other objectives were achieved. Compared to the Eclipse generated plans, the proposed Ethos template generated plans with greater evaluation planning target volume (PTV_Eval) V100% coverage (p = 0.01), significantly decreased heart V1.5Gy (p< 0.001), and increased contralateral breast V5Gy, skin D0.01cc, and RTOG conformity index (p = 0.03, p = 0.03, and p = 0.01, respectively). However, only the reduction in heart dose was significant after correcting for multiple testing. Physicist-selected plans were deemed clinically acceptable without modification for 75% and 90% of plans by physicians A and B, respectively. Physicians A and B scored at least one automatically generated plan as clinically acceptable for 100% and 95% of planning intents, respectively. Standard left- and right-sided planning templates automatically generated APBI plans of comparable quality to manually generated plans treated on a stereotactic linear accelerator, with a significant reduction in heart dose compared to Eclipse generated plans. The methods presented in this work elucidate an approach for generating automated, cardiac-sparing APBI treatment plans for daily adaptive RT with high efficiency.

Feasibility and Short-Term Toxicity of a Consecutively Delivered Five Fraction Stereotactic Body Radiation Therapy Regimen in Early-Stage Breast Cancer Patients Receiving Partial Breast Irradiation.

BackgroundFor appropriately selected patients with early-stage breast cancer (ESBC), accelerated partial breast irradiation (APBI) yields equivalent rates of ipsilateral breast tumor recurrence with mixed results in patient-rated cosmesis compared with whole-breast radiotherapy depending on the technique utilized. When utilizing external beam radiotherapy for APBI, techniques to reduce target margins and overall treatment volume are potentially important to decrease rates of long-term adverse cosmesis. Stereotactic body radiotherapy (SBRT) is a promising technique to deliver APBI because of its increased accuracy and sparing of uninvolved breast tissue. We report the initial results of a prospective clinical trial investigating feasibility, safety, and cosmetic outcomes of a daily five-fraction SBRT regimen for APBI.MethodsTwenty-three patients with ESBC after lumpectomy who met APBI suitability were enrolled. During lumpectomy, a bioabsorbable three-dimensional fixed array tissue marker (BioZorb™, Hologic, Marlborough, MA) was placed for enhanced visualization of the cavity boundaries. Clinical target volume (CTV) was defined as the delineable cavity plus a 1-cm isotropic expansion followed by a 3-mm isotropic planning target volume (PTV) expansion. Patients received 30 Gy delivered in five planned consecutive daily fractions in either prone or supine positioning depending on individual anatomy. Two patients completed the five-fraction treatments in 9-day interval and 11-day interval due to external circumstances. A maximum PTV of 124cc was allowed to minimize incidence of fat necrosis. Plans utilized 10-MV flattening filter–free beams delivered on a Varian Edge linear accelerator. Local control, toxicity, and nurse/patient-scored cosmesis at pre-treatment baseline, 1 month post-treatment, and at subsequent 6-month intervals were recorded.ResultsTwenty-three patients were accrued at the time of submission with median follow-up of 6 months. No patients experienced grade ≥3 acute toxicity. Of the 10 events reported probably related to SBRT, nine were grade 1 (n = 9/10, 90%). There was no evidence of difference, deterioration, or change in patient or nurse-scored cosmesis from baseline to 1 and 6 months post-treatment. One patient developed nodal failure shortly after APBI.ConclusionsAlthough longer follow-up is needed to assess long-term toxicity and local control, this study demonstrated a five-fraction SBRT regimen delivered over consecutive days is a safe, efficient, well-tolerated, and cosmetically favorable means of delivering APBI in suitable women.Clinical Trial Registrationhttps://www.clinicaltrials.gov/ct2/show/NCT03643861, NCT03643861.

Abstract P3-19-18: Feasibility and short term toxicity of a consecutively delivered 5-fraction stereotactic body radiotherapy in early stage breast cancer patients receiving accelerated partial breast irradiation

Abstract Background: For early stage breast cancer, breast conservation is an appealing option for many women. Interest in reducing the duration and volume of RT has continued in the form of accelerated partial breast irradiation (APBI). Recent work demonstrates that for appropriately selected patients, APBI yields equivalent rates of ipsilateral breast tumor recurrence (IBTR) with improvements in patient rated cosmesis compared to whole breast radiotherapy. When using external beam radiotherapy for APBI, appropriate techniques to reduce setup uncertainty and overall treatment volume are critical to reduce long term adverse cosmesis. Stereotactic body radiotherapy (SBRT) is an attractive technique to deliver APBI because of its excellent accuracy and potentially superior sparing of uninvolved breast tissue. We report here the initial results of our IRB-approved prospective clinical trial investigating feasibility, safety, and cosmetic outcomes of daily, 5-fraction SBRT for APBI in women meeting ASTRO consensus suitability criteria for partial breast irradiation. Methods: We enrolled twenty patients with early stage breast cancer after lumpectomy who met APBI suitability. During lumpectomy, a breast surgeon placed a bioabsorbable 3D fixed array tissue marker (BioZorb™, Hologic, Marlborough, MS) for enhanced visualization of the cavity boundaries. We defined the clinical target volume (CTV) as the delineable cavity plus a 1 cm isotropic expansion. We placed a 3mm isotropic planning target volume (PTV) expansion around the CTV. We treated each woman with 3000cGy delivered in 5 consecutive, daily, free-breathed fractions in either prone or supine positioning depending on individual anatomy. Patient treatment position was free-breathing in the prone position for 17 patients and supine in 3 patients using triggered imaging and fiducial tracking. A maximum PTV of 124cc was allowed to minimize incidence of fat necrosis. Plans used 10MV flattening filter free (FFF) beams delivered on a Varian Edge linear accelerator. We documented the following: adverse events (CTCAE 4.0), causality of the event to SBRT, and nurse- and patient- scored cosmesis (i.e., 1=excellent, 2=good, 3=fair, 4=poor) at pre-treatment baseline, 1 month post-treatment, and at each proceeding 6 month interval. We also clinically surveilled for disease recurrence. Results: We completed accrual and treatment of twenty patients. At time of submission, fourteen patients had completed 6-month follow-up, and eight patients had completed 12-month follow-up. No patients experienced grade ≥3 toxicity. Adverse events reported were mostly grade 1. Eighty-five percent of patient-scored cosmesis were excellent or good at baseline, with no evidence of deterioration at 1-month post-treatment (Baseline = 1.9, 1 month = 1.9; p = 0.914). Eighty-five percent of nurse-scored cosmesis reports were excellent or good at baseline, with no evidence of deterioration between baseline and 1-month post-treatment (Baseline = 1.95, 1 month = 1.90; p = 0.564). Eighty-six percent of patients had excellent or good patient- and nurse-scored cosmesis at 6 months, with either mean score of 1.71. Eighty-eight percent of patients had excellent or good patient- and nurse-scored cosmesis at 12 months, with mean cosmesis score of 1.50 and 1.63, respectively. Local control follow-up is immature, but no patients developed IBTR. Conclusions: Our study demonstrated that consecutive daily 5-fraction breast SBRT for APBI is a safe, efficient, well-tolerated, and cosmetically favorable means of delivering partial breast irradiation in suitable women. As follow-up duration matures, we will report long term cosmetic outcome and recurrence. Citation Format: Yilan Liu, Christopher Veale, Diana Hablitz, Helen Krontiras, Michael Christian Dobelbower, Rachael Lancaster, Markus Bredel, Catherine Parker, Kimberly Keene, Evan Thomas, Drexell Hunter Boggs. Feasibility and short term toxicity of a consecutively delivered 5-fraction stereotactic body radiotherapy in early stage breast cancer patients receiving accelerated partial breast irradiation [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P3-19-18.

Proton Therapy for Partial Breast Irradiation: Rationale and Considerations.

In an era of continued advancements in personalized medicine for the treatment of breast cancer, select patients with early stage breast cancer may be uniquely poised to benefit from partial breast irradiation (PBI) delivered with proton therapy. PBI presents an opportunity to improve quality of life during treatment with a significantly shorter treatment duration. By targeting less non-target breast tissue, excess radiation exposure and resulting toxicities are also reduced. Proton therapy represents a precision radiotherapy technology that builds on these advantages by further limiting the normal tissue exposure to unnecessary radiation dose not only to uninvolved breast tissue but also the underlying thoracic organs including the heart and lungs. Herein, we present a concise review of the rationale for the use of proton therapy for PBI, evidence available to date, and practical considerations in the implementation and use of proton therapy for this indication.

Abstract LB-221: Tumor microenvironment predicts aggressive breast cancer: Combination of HERV-K, immune checkpoint and activation status of CD8+ T cells

Abstract The purpose of this study is to identify useful predictive biomarkers of anti-tumor cytotoxic T cell and immune checkpoint response, to optimize targeted therapy, chemotherapy, and immunotherapy of breast cancer (BC). Immune checkpoint markers (14 circulating proteins) in BC patients were determined using a magnetic bead assay. CD27, PDL-2, LAG-3, and TIM-3 levels in the plasma, obtained from patients with invasive ductal carcinoma (IDC: n=32), ductal carcinoma in situ (DCIS: n=11), other invasive BC (n=12), prophylactic surgery (BRCA1: n=7, and BRCA2 mutations: n=4, benign phyllodes tumor: n=2) were significantly enhanced compared with healthy controls (n=12), which supports a highly immune suppressed environment. Also, these markers were compared at T1 (before surgery), T2 (6 months post-surgery), and T3 (18 months post-surgery). LAG-3, CD27 (p=0.012), 4-1BB, CD80, PD-L2, PD-L1 (p=0.045), and IDO all had a decreased plasma concentrations post-surgery (6 months or 18 months) compared with pre-surgery. Interestingly, significantly higher concentrations of BTLA, IDO, LAG-3, PD-L2, TIM-3, and PD-L1 were detected in BC patients that had higher titers of circulating human endogenous retrovirus type K (HERV-K) antibodies. We characterized the subset distribution and phenotype of the immune cells of various BC patients. CD4+ or CD8+ T cells collected from breast tissues including tumor tissues (T) or uninvolved normal breast tissues (N) were analyzed for CD4 and CD8. Higher percentages of CD8+ tumor infiltrating lymphocyte (TIL) cells were detected in N compared with autologous T. The percentage of CD38+/HLA-DR+ CD8+ T cells was low in the PBMCs compared to percentages in TILs. Activated T cells were only detected in TIL (40% to 99%; n=6) but not in their PBMCs from the same patients. Several studies have demonstrated that regulatory T cells (Tregs) within the tumor environment lead to a decrease in patient survival. Higher percentages of Treg cells were observed in aggressive BC and in TIL obtained from tumor tissues compared to TIL cells obtained from uninvolved tissues. HERV-K specific CD8 T cells (K-T cells) enhanced the percentage of CD8 T cells (62.5%; n=8) in patients with aggressive BC, and were able to kill their autologous tumor cells or mammospheres. Significantly reduced tumor weights and metastases were demonstrated in MDA-MB-231 mice treated with K-T cells. The lower percentage of CD8 T cells and higher percentage of Treg T cells we observed in breast tumor tissues, coupled with elevated concentrations of circulating immune checkpoint proteins in BC patients, suggests a highly immune suppressed environment in these aggressive tumors that could be minimized with anti-HERV-K therapy. Thus, HERV-K T cell therapies combined with blockade of immune checkpoints may be a new regimen for treating aggressive BC. Citation Format: Feng Wang-Johanning, Jia Li, Gary Johanning, Ming Li, Raghu Chivukula, Guoqing Wu. Tumor microenvironment predicts aggressive breast cancer: Combination of HERV-K, immune checkpoint and activation status of CD8+ T cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr LB-221. doi:10.1158/1538-7445.AM2017-LB-221

Intra-operative re-excision of margins in breast-conserving surgery: How well are we able to judge tumour proximity?

s / International Journal of Surgery 10 (2012) S1–S52 S15 ABSTRACTS 1127: INTRA-OPERATIVE RE-EXCISION OF MARGINS IN BREASTCONSERVING SURGERY: HOW WELL ARE WE ABLE TO JUDGE TUMOUR PROXIMITY? Lorna Cook, Debasish Debnath, Zoe Lin, Raouf Daoud, Isabella Karat, Ian Laidlaw. Frimley Park Hospital, Frimley, UK Introduction: In breast-conserving surgery (BCS), specimen margin involvement with tumour often necessitates re-operation. For this reason, extra margins are often taken at the time of primary operation. The aim of this study was to determine how often such re-excisions are appropriate. Methods: The records of 100 consecutive patients undergoing BCS for cancer during 2011 were analysed. Data was collected on demographics, tumours characteristics, surgery performed and final histology. Results: All 100 patients were female, median age 56 years (range 33-83 years). 66% had intra-operative re-excision of margins. In 26/66 (40%), re-excision was the correct decision. 25 of the 34 patients who had no extramargins taken at all were managed appropriately with the remaining 9 patients having close/positive margins. Unnecessary re-excisions were performed in 40/66 patients. The correct intraoperative decision was therefore made in 51% of patients. Conclusion: The decision regarding intra-operative margin re-excision was appropriate in just over half of all cases and a second operation was avoided in 26%. However, in order to preserve as much uninvolved breast tissue as possible, further work should be done on methods to improve accuracy and to determine whether there are any patient, radiological or pathological predictors to guide intra-operative re-excision. 1181: FACTORS PREDICTING POSITIVE MARGIN STATUS IN BREAST CANCER PATIENTS UNDERGOING BREAST CONSERVING SURGERY Euan Harris , Andrew Lee . University of Dundee, Dundee, Tayside, UK; Abertawe Bro Morgannwg University Healthboard, Swansea, West

Adenomyoepithelioma A Rare Breast Tumor: Case Studies With Review Of The Literature

Introduction : Adenomyoepithelioma is a rare, benign proliferative tumor that can involve the breast tissue. The diagnostic appearance of the lesion can give rise to a difficult differential diagnosis with breast carcinoma. There is the potential for malignant evolution of this lesion. It usually presents as a solitary, unilateral, painless mass at the periphery of the breast. Accurate diagnosis and differentiation from more aggressive tumors is important. Materials and methods :In two cases of breast lumps, fine needle aspiration cytology (FNAC) was done and stained with H & E and Papanicolaou stains. The formalin-fixed tissue sections were stained with hematoxylin and eosin and in one case, immunohistochemistry was performed with smooth muscle actin (SMA).Case 1: A 30-year-old female presented with a firm lump in the upper quadrant of the right breast along with axillary lymphadenopathy since 6 months. On ultrasound it was reported as duct ectasia. FNAC was suggestive of proliferative breast disease with atypia (possibility of duct cell carcinoma). Modified radical mastectomy was done and sent for histopathology. The histopathological diagnosis was adenomyoepithelioma – breast (right), tubular type.Case 2: A 25-year-old female presented with a firm lump in the left breast, occupying all quadrants. On FNAC the aspirates were suggestive of fibroadenoma. Peroperatively it was found to be a cystic swelling occupying the whole breast. The mass was excised and sent for histopathology. The histopathological diagnosis was adenomyeoepithelioma with apocrine metaplasia and cystic degeneration – breast (left), tubular type.Results : Two cases of breast lump were managed surgically. Both the cases were females with the mean age being 27 years. Both cases were of adenomyoepithelioma (tubular type) with one case showing apocrine metaplasia and cystic change. Conclusion : Prognosis of patients with benign adenomyoepithelioma of the breast is usually good, but it has a potential for local recurrence, especially in the tubular and lobulated variants. Total surgical excision with an adequate margin of uninvolved breast tissue is therefore recommended.

Papillary Neoplasms of the Breast A Review

Context Interpretation of papillary lesions of the breast remains a challenging task because of the wide morphologic spectrum encountered in the benign, atypical, and malignant subtypes. Data on clinical significance and outcome of papillary lesions, with superimposed atypia or areas similar to ductal carcinoma in situ partially replacing the benign elements, are sparse. Furthermore, complete excision of even a fully developed papillary carcinoma confined to a dilated or cystic duct is associated with an excellent prognosis, whereas a complex papilloma extending into multiple branches of a duct may ultimately recur as a carcinoma because of incomplete excision of microscopic foci. This makes an outcome-based classification difficult. Objective An arbitrary yet practical approach to classification is outlined, with discussion of methods to circumvent the various diagnostic difficulties. The limitations in precise diagnosis of papillary lesions in aspirates are addressed, and the implications of finding papillary lesions in core biopsies are discussed. Although the focus is on intraductal lesions, associated invasive carcinomas and invasive micropapillary carcinoma are also presented. Data sources The literature on papillary lesions and invasive micropapillary carcinoma is reviewed. Conclusions It would be prudent to completely excise any papillary lesion that has not been entirely removed by the initial core biopsy. The optimal management of localized papillary lesions is complete excision with a small rim of uninvolved breast tissue without any prior needle instrumentation if and when the papillary nature can be determined by imaging. Thus managed, most of these lesions behave indolently, and outcome is usually excellent.

Papillary Neoplasms of the Breast: A Review

Interpretation of papillary lesions of the breast remains a challenging task because of the wide morphologic spectrum encountered in the benign, atypical, and malignant subtypes. Data on clinical significance and outcome of papillary lesions, with superimposed atypia or areas similar to ductal carcinoma in situ partially replacing the benign elements, are sparse. Furthermore, complete excision of even a fully developed papillary carcinoma confined to a dilated or cystic duct is associated with an excellent prognosis, whereas a complex papilloma extending into multiple branches of a duct may ultimately recur as a carcinoma because of incomplete excision of microscopic foci. This makes an outcome-based classification difficult. An arbitrary yet practical approach to classification is outlined, with discussion of methods to circumvent the various diagnostic difficulties. The limitations in precise diagnosis of papillary lesions in aspirates are addressed, and the implications of finding papillary lesions in core biopsies are discussed. Although the focus is on intraductal lesions, associated invasive carcinomas and invasive micropapillary carcinoma are also presented. The literature on papillary lesions and invasive micropapillary carcinoma is reviewed. It would be prudent to completely excise any papillary lesion that has not been entirely removed by the initial core biopsy. The optimal management of localized papillary lesions is complete excision with a small rim of uninvolved breast tissue without any prior needle instrumentation if and when the papillary nature can be determined by imaging. Thus managed, most of these lesions behave indolently, and outcome is usually excellent.

Protein kinases C isozymes are differentially expressed in human breast carcinomas

Aims The protein kinase C (PKC) family of enzymes has been implicated in cellular proliferation, differentiation, and apoptosis. However, the distribution of specific PKC isoforms with varying functions in normal and malignant human tissues remains to be determined. The objective of this study was to investigate the expression of certain PKC isoforms (α, βI, βII, ε) in human breast cancer specimens relative to adjacent uninvolved tissue ( n = 24) and in the normal breast tissue obtained from patients undergoing reduction mammoplasty ( n = 12). Main methods Western blot analysis using PKC isoform specific antibodies was performed on tissue extracts from breast tumors, adjacent uninvolved tissues, and reduction mammoplasty tissues. Key findings Mean levels of cytosolic and membrane PKC-α, PKC-βI, and PKC-βII were significantly higher in the cancer specimens than in the adjacent uninvolved breast tissues (Wilcoxon signed-ranks test; P < 0.05 for each, after adjustment for multiple comparisons). There was a notably higher mean level of membrane PKC-βII isozyme in Her-2 positive and in poorly differentiated tumors. No significant differences were observed when normal tissue adjacent to tumor was compared to breast tissue obtained from reduction mammoplasty specimens. Significance Higher level of PKC-α, PKC-βI, and PKC-βII in cancer specimens and higher level of PKC-βII in Her-2 positive tumors require further exploration of the intracellular pathways involving PKC-α and -β isoforms in breast cancer because both could be specific targets for the development of new therapies and for the prevention and treatment of this disease.

Quantitative MRI assessment of uninvolved breast tissue in patients with locally-advanced breast cancer undergoing neoadjuvant chemotherapy.

Abstract Abstract #4016 Background: We studied the Magnetic Resonance Imaging (MRI) characteristics of breast tissue in patients with invasive cancers to better understand the relationship between tumors and their normal “host” tissue. There is a strong association between tumor morphologic patterns on MRI and both treatment outcome and survival. The objectives of this study were to assess the characteristics of normal host tissue to verify whether they were associated with treatment outcome.&amp;#x2028; Materials and Methods: 42 women with locally-advanced breast cancer underwent MRI exams prior to the start of neoadjuvant adriamycin/cytoxan (AC) chemotherapy (MRI1), following one cycle of AC (MRI2) and at the completion of chemotherapy, prior to surgery (MRI3). Patients mean age was 48.56 years (range 29.7 - 71.5). Median follow-up was 178 weeks. Pathology and radiology reports were available for all patients. Total follow-up was 3 years. We quantified tumor and surrounding normal tissue enhancement. Signal Enhancement Ratio (SER) values have been histopathologically shown to positively correlate with microvessel density within tumor. We lowered the enhancement threshold necessary to detect MRI tumor-enhancing regions, to insure that slow or non-enhancing tissue regions could be analyzed. For all patients, tumor morphologic patterns were categorized according to the degree of tumor containment from 1, denoting well-circumscribed uni-centric masses to 5, corresponding to infiltrative tumors with ill-defined borders. MRI breast tissue composition quantitative measures included 1) volumetric breast density, 2) tumor volume, 3) normal fibroglandular tissue volume and 4) SER levels in tumor and normal surrounding tissue. In order to quantify the uninvolved breast tissue volume, we used a semi-automated technique that performs a segmentation of MRI breast tissue from fat regions without user interaction, therefore avoiding subjective threshold decision issues. Regions in normal tissue surrounding tumors were automatically selected and analyzed at all time points.&amp;#x2028; Results: 10 patients recurred. We found that non-tumor mean SER did not change significantly with one cycle of chemotherapy, however the value of non-tumor mean SER after 1 cycle of chemotherapy was an independent predictor of disease free survival. We also observed a strong association between the initial (pre-treatment) tumor morphologic patterns on MRI with treatment outcome. We found that volume of normal breast tissue at baseline does relate to tumor phenotype (p&amp;lt;0.067), and change in non-tumor breast tissue volume after chemotherapy was also predictive of recurrence (p&amp;lt;0.075).&amp;#x2028; Conclusion: We found that measures of uninvolved breast tissue volume were correlated with tumor phenotypes and were shown to be predictive of recurrence. These results suggest that the host tissue may provide treatment response information. Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 4016.

Differential spatial expression and activation pattern of EGFR and HER2 in human breast cancer

Overexpression of HER family members is a well established prognostic factor and identifies potential targets for antibody-based receptor blocking strategies. While several studies have analyzed the expression of HER2 and other HER-family members in malignant tumors, considerably less is known about their expression and activation in non-involved breast tissue from breast cancer patients. We have therefore investigated the differential expression of EGFR, HER2, and their tyrosine-kinase activated forms (ptyr-1248 Her-2 and ptyr-845 EGFR) in 63 tumor specimen containing: a) malignant epithelium, b) in non-malignant tissue located at the peritumoral margin, and c) in uninvolved breast tissue obtained from tissue distant from the tumor. Using immunohistochemistry (IHC), we found significantly higher HER2 protein expression levels in malignant epithelium than in marginal and peripheral non-malignant epithelium (p=1.3 x 10(-10) Fisher's exact test). Epithelial EGFR expression did not differ between the three tissue types, but stromal EGFR protein was significantly more common in marginal and peripheral tissues when compared to tumor tissues (p=0.008, Fisher's exact test). When analyzing activated receptor forms, we found epithelial ptyr-1248 HER2 expression in one tumoral, one marginal and one peripheral sample. We did not observe ptyr-845 EGFR in any of the samples analyzed. We found a significant overall correlation between epithelial and stromal EGFR expression (r=0.442; p<0.0001; Spearman's Rho), and between stromal EGFR expression and normal tissue type (r=0.170; p<0.02; Spearman's Rho). Epithelial HER2 expression and normal tissue type (r=0.492; p<0.0001; Spearman's Rho) were inversely correlated. Taken together, we have observed a differential expression pattern of EGFR, HER2, and activated HER2 that is dependent on the spatial relation to a malignant tumor. Our findings of decreased intratumoral EGFR expression and the absence of activated EGFR suggests that, in contrast to HER2, EGFR inhibition might not be an ideal target for antibody therapy.

Increased dihydropyrimidine dehydrogenase activity in breast cancer.

Although studies have focused on modulating the bioavailability of 5-FU through inhibition of dihydropyrimidine dehydrogenase (DPD) to improve efficacy of the drug, activity of this enzyme in breast cancer has not been thoroughly examined. We measured DPD activity in primary and metastatic lesions and benign breast tumors to evaluate the clinical significance of this enzyme in the treatment of breast cancer. DPD activity was measured by catalytic assay and compared in 100 primary tumors (95 invasive carcinomas, 5 intraductal carcinomas), 26 uninvolved adjacent breast tissue specimens, 6 metastatic sites, and 7 intraductal papillomas. The enzyme level in the carcinomas was 4-fold that of adjacent uninvolved breast tissues (101 vs 23 pmol/min/mg protein, P < 0.001). Enzyme activity in intraductal papilloma (120 pmol/min/mg protein) was comparable to that in invasive carcinoma. There were no significant differences in DPD activity related to clinicopathologic features, but a tendency toward increased DPD activity was observed in progesterone receptor-negative breast cancer (P = 0.09). There was marginal correlation in enzyme activity between primary and metastatic lesions (P = 0.07). DPD activity is substantially upregulated in breast cancer tissue and is higher than that reported previously. The clinical implications of DPD inhibitors in patients being treated for breast cancer with oral fluoropyrimidine chemotherapy should be further investigated.

Identification of novel breast tumor-specific mutation(s) in the q11.2 region of chromosome 17 by RAPD/AP-PCR fingerprinting

Analysis of genetic instability in breast cancer tissues compared to uninvolved breast tissues from the same individuals by RAPD (random amplified polymorphic DNA)/AP-PCR (arbitrarily primed PCR) fingerprinting using 30 arbitrary primers revealed 190 amplified DNA fragments. Presumably, each of these represents a gene locus in a different region of the genome of breast cancer tissues. Among these amplified DNA fragments, 65 (34.2%) exhibited presence and absence or reductions and enhancements in the intensity in breast cancer tissues compared to uninvolved breast tissues from the same individuals, and 11 amplified DNA fragments (5.7%) represented polymorphisms in the uninvolved human breast tissues. Reductions and enhancements in the intensity of some of the amplified fragments were observed indicating allelic gains or losses in the breast tumor genome compared to the matched uninvolved tissue genome. The presence or absence of some of the amplified DNA fragments were observed in this study indicating homozygous deletions or insertions in the breast tumor DNA compared to the matched uninvolved tissue DNA. Notably, an insertion of a 1270 bp amplified fragment was observed in 81% (17 of 21) of the tumor samples using the primer, OPC04. This amplified fragment resolved into two, 1200 and 1300 bp, single-stranded amplified fragments on the denaturing sequencing gel. This separation into single-stranded fragments suggests that the amplified fragment contains a conformation that is semistable. The 1270 bp amplified fragment localizes to the q11.2 region of chromosome 17. Sequence analysis of this fragment showed a significant DNA base sequence similarity (93%) with one of the breast tumor-specific human EST. The similarity with EST sequences and RT-PCR analysis showed that a part of this amplified fragment is from the coding region of the genome. Any one of the events observed in this study could play an important role in the development of breast cancer or could occur during the clonal expansion of the genetically unstable breast cells.

Hormone-Regulated Genes (pS2, PIP, FAS) in Breast Cancer and Nontumoral Mammary Tissue

Expression of the hormone-regulated genes, pS2, prolactin-inducible protein (PIP) and fatty acid synthetase (FAS), was investigated by Northern blotting in primary breast carcinoma, metastatic breast cancer in axillary lymph nodes, in uninvolved breast tissue from mastectomies and in normal lymph nodes. There were considerable differences in expression of the genes between the tissues. The proportion of tissues containing PIP-mRNA decreased from uninvolved breast tissue to primary breast carcinoma to metastatic carcinoma. The reverse applied to FAS-mRNA which was found more often in metastatic cancer than in primary cancer, and least frequently in uninvolved breast tissue. Yet another pattern was observed for pS2 expression. The highest proportion of tissues demonstrating gene expression was found in primary breast cancer with both metastatic tumor and uninvolved breast tissue expressing the gene less frequently. pS2-mRNA and PIP-mRNA could only rarely be detected in trace amounts in normal lymph nodes. In contrast, FAS-mRNA was present in about one third of normal lymph nodes. Only pS2-mRNA showed an association with estrogen and progesterone receptor status.

Stromelysin‐3 expression in breast cancer biopsies: Clinico‐pathological correlations

Stromelysin-3 expression was studied by Northern blotting in 222 tissue samples including primary and metastatic breast carcinoma and normal breast tissue. Uninvolved breast tissue from mastectomy specimens, normal breast tissue from reduction mammoplasties and normal lymph nodes did not contain stromelysin-3 mRNA. About 62% of primary and metastatic breast carcinomas, but only 1 of 10 in situ ductal carcinomas, expressed stromelysin-3. Stromelysin-3 mRNA was found more often in estrogen-receptor-positive carcinomas and in histological grade-1 carcinomas. There was no significant correlation between stromelysin-3 expression and other prognostic factors, including tumor size, lymph-node involvement, age of patient, vascular invasion and cathepsin-D.

Expression of the pS2 gene in normal breast tissue.

pS2 expression in normal breast tissue removed for cosmetic reasons was significantly lower than in uninvolved breast tissues from mastectomies for breast carcinomas. It is speculated that the presence of the carcinoma, or factors related to its development, could be the reason for this difference.