Early Blood Flow Abnormalities in Axillary and Brachial Arteries Precede the Onset of Persistent Lymphedema in Women Treated Surgically for Breast Cancer.

Background: surgery to treat breast cancer (BC) is associated with upper limb (UL) lymphedema, which in some cases may become permanent. It is uncertain whether lymphedema results from injury to either lymphatic or blood vessels, or to both. Methods and Results: a cohort of 200 BC patients was examined 1, 3, 6, 12, and 24 months after surgery. Axillary and brachial blood vessels were evaluated using Doppler Ultrasound, and patients had their UL examined for lymphedema at each visit. Patients who developed lymphedema 24 months after surgery presented with higher mean flow velocity (MFV) and end diastolic velocity (EDV) in both axillary (MFV = 13.57 vs. 10.7 cm/s, p = 0.02; EDV = 5.62 vs. 3.47 cm/s; p = 0.004) and brachial (MFV = 11.44 vs. 8.74 cm/s; p = 0.03; EDV = 5.08 vs. 3.04; p = 0.04) arteries as early as 1 month after surgery. Similar associations were found 3, 6, and 12 months after surgery. Early abnormalities of the resistive and pulsatility indexes were also significantly associated with persistent lymphedema. EDV measured 1 month after surgery had the best performance to detect patients who will later develop long-term lymphedema, (sensitivity = 73.7%; specificity = 71.2%; negative predictive value = 57.6%). Conclusion: vascular abnormalities precede and are possible causal factors for UL lymphedema in BC patients.

Metabolomics by NMR Combined with Machine Learning to Predict Neoadjuvant Chemotherapy Response for Breast Cancer.

Neoadjuvant chemotherapy (NACT) is offered to patients with operable or inoperable breast cancer (BC) to downstage the disease. Clinical responses to NACT may vary depending on a few known clinical and biological features, but the diversity of responses to NACT is not fully understood. In this study, 80 women had their metabolite profiles of pre-treatment sera analyzed for potential NACT response biomarker candidates in combination with immunohistochemical parameters using Nuclear Magnetic Resonance (NMR). Sixty-four percent of the patients were resistant to chemotherapy. NMR, hormonal receptors (HR), human epidermal growth factor receptor 2 (HER2), and the nuclear protein Ki67 were combined through machine learning (ML) to predict the response to NACT. Metabolites such as leucine, formate, valine, and proline, along with hormone receptor status, were discriminants of response to NACT. The glyoxylate and dicarboxylate metabolism was found to be involved in the resistance to NACT. We obtained an accuracy in excess of 80% for the prediction of response to NACT combining metabolomic and tumor profile data. Our results suggest that NMR data can substantially enhance the prediction of response to NACT when used in combination with already known response prediction factors.

Longitudinal, Long-Term Comparison of Single- versus Multipoint Upper Limb Circumference Periodical Measurements as a Tool to Predict Persistent Lymphedema in Women Treated Surgically for Breast Cancer: An Optimized Strategy to Early Diagnose Lymphedema and Avoid Permanent Sequelae in Breast Cancer Survivors.

PURPOSE: We aim to evaluate whether upper limb (UL) circumference (ULC) and UL swelling sensation (ULSS) performed shortly after surgery or later on during follow-up can predict long-term/persistent forms of lymphedema in women who underwent surgery for breast cancer. PATIENTS AND METHODS: Eighty-five women completed at least 24 months of follow-up. At each follow-up visit (1, 3, 6, 12, and 24 months after surgery), patients were tested for lymphedema using ULC and ULSS. Two different approaches to ULC were compared: (1) a "positive" lymphedema diagnosis if a difference ≥ 2 cm between the affected and contralateral UL was detected in at least two contiguous measurement points (MPs) and (2) a "positive" result if just one MP ≥ 2 cm. Patients were also questioned about their perception of weight, swelling, and/or tension (ULSS). The gold standard for long-term lymphedema was a water displacement difference between the UL ≥ 200 mL 24 months after surgery (ULWD). RESULTS: Twenty-four months after surgery, 19 (22.4%) women were diagnosed with long-term lymphedema. Using 24-month data, comparison of log-likelihoods denoted a clear superiority of the ULC approach 1 compared with 2 for the diagnosis of long-term lymphedema (p < 0.001). Using approach 1, the best prediction of a woman developing long-term lymphedema if she had a positive ULC in the follow-up was obtained at 6 months after surgery (posterior probability of 60%). CONCLUSIONS: Our study reveals that performing ULC 6 months after surgery, regarding as "positive" only women with a difference ≥ 2 cm at two contiguous MPs, is the best strategy to identify women at increased risk of later developing permanent forms of lymphedema.

Multiplatform Investigation of Plasma and Tissue Lipid Signatures of Breast Cancer Using Mass Spectrometry Tools.

Plasma and tissue from breast cancer patients are valuable for diagnostic/prognostic purposes and are accessible by multiple mass spectrometry (MS) tools. Liquid chromatography-mass spectrometry (LC-MS) and ambient mass spectrometry imaging (MSI) were shown to be robust and reproducible technologies for breast cancer diagnosis. Here, we investigated whether there is a correspondence between lipid cancer features observed by desorption electrospray ionization (DESI)-MSI in tissue and those detected by LC-MS in plasma samples. The study included 28 tissues and 20 plasma samples from 24 women with ductal breast carcinomas of both special and no special type (NST) along with 22 plasma samples from healthy women. The comparison of plasma and tissue lipid signatures revealed that each one of the studied matrices (i.e., blood or tumor) has its own specific molecular signature and the full interposition of their discriminant ions is not possible. This comparison also revealed that the molecular indicators of tissue injury, characteristic of the breast cancer tissue profile obtained by DESI-MSI, do not persist as cancer discriminators in peripheral blood even though some of them could be found in plasma samples.

Copy number alterations associated with clinical features in an underrepresented population with breast cancer.

BACKGROUND: As the most incident tumor among women worldwide, breast cancer is a heterogeneous disease. Tremendous efforts have been made to understand how tumor characteristics as histological type, molecular subtype, and tumor microenvironment collectively influence disease diagnosis to treatment, which impact outcomes. Differences between populations and environmental and cultural factors have impacts on the origin and evolution of the disease, as well as the therapeutic challenges that arise due to these factors. We, then, compared copy number variations (CNVs) in mucinous and nonmucinous luminal breast tumors from a Brazilian cohort to investigate major CNV imbalances in mucinous tumors versus non-mucinous luminal tumors, taking into account their clinical and pathological features. METHODS: 48 breast tumor samples and 48 matched control blood samples from Brazilian women were assessed for CNVs by chromosome microarray. Logistic regression and random forest models were used in order to assess CNVs in chromosomal regions from tumors. RESULTS: CNVs that were identified in chromosomes 1, 5, 8, 17, 19, and 21 classify tumors according to their histological type, ethnicity, disease stage, and familial history. CONCLUSION: Copy number alterations described in this study provide a better understanding of the landscape of genomic aberrations in mucinous breast cancers that are associated with clinical features.

Molecular Signatures of High-Grade Cervical Lesions.

Cervical cancer is the fourth most common neoplasia in women and the infection with human papilloma virus (HPV) is its necessary cause. Screening methods, currently based on cytology and HPV DNA tests, display low specificity/sensitivity, reducing the efficacy of cervical cancer screening programs. Herein, molecular signatures of cervical cytologic specimens revealed by liquid chromatography-mass spectrometry (LC-MS), were tested in their ability to provide a metabolomic screening for cervical cancer. These molecules were tested whether they could clinically differentiate insignificant HPV infections from precancerous lesions. For that, high-grade squamous intraepithelial lesions (HSIL)-related metabolites were compared to those of no cervical lesions in women with and without HPV infection. Samples were collected from women diagnosed with normal cervix (N = 40) and from those detected with HSIL from cytology and colposcopy (N = 40). Liquid-based cytology diagnosis, DNA HPV-detection test, and LC-MS analysis were carried out for all the samples. The same sample, in a customized collection medium, could be used for all the diagnostic techniques employed here. The metabolomic profile of cervical cancer provided by LC-MS was found to indicate unique molecular signatures for HSIL, being two ceramides and a sphingosine metabolite. These molecules occurred independently of women's HPV status and could be related to the pre-neoplastic phenotype. Statistical models based on such findings could correctly discriminate and classify HSIL and no cervical lesion women. The results showcase the potential of LC-MS as an emerging technology for clinical use in cervical cancer screening, although further validation with a larger sample set is still necessary.

Prognostic significance of PD-L1 and PD-L2 in breast cancer.

PD-L1 and PD-L2 constitute an important antitumor immune response. In breast cancer, their prognostic value is still to be defined. In this study, we investigate the correlation between PD-L1 and PD-L2 protein expressions with clinical and pathologic features and disease-free survival and overall survival. To assess PD-L1 and PD-L2 expressions, we conducted immunohistochemistry studies using a breast cancer tissue microarray encompassing a total of 192 breast cancer cases, stages I, II, and III, with detailed clinical and outcome data. PD-L1 expression was present in 56.6% (107/189), and PD-L2 expression was identified in 50.8% (97/191) of breast cancer cases. Younger age at diagnosis, lymph node positivity, negative estrogen receptor, and recurrence at distant sites were all associated with both PD-L1 and PD-L2 expressions. The presence of larger tumors was associated only with PD-L1 expression. In our study, PD-L1 expression was significantly associated with better overall survival (P = .04) in breast cancer patients. Despite its association with poor clinical and pathologic features, PD-L1 expression emerges as a positive prognostic biomarker in breast cancer. This survival result might be due to the presence of a strong antitumor immune response leading to PD-L1 expression.

Analysis of the contribution of immunologically-detectable HER2, steroid receptors and of the "triple-negative" tumor status to disease-free and overall survival of women with epithelial ovarian cancer.

We assessed associations between steroid receptors including: estrogen-alpha, estrogen-beta, androgen receptor, progesterone receptor, the HER2 status and triple-negative epithelial ovarian cancer (ERα-/PR-/HER2-; TNEOC) status and survival in women with epithelial ovarian cancer. The study included 152 women with primary epithelial ovarian cancer. The status of steroid receptor and HER2 was determined by immunohistochemistry. Disease-free and overall survival were calculated and compared with steroid receptor and HER2 status as well as clinicopathological features using the Cox Proportional Hazards model. A mean follow-up period of 43.6 months (interquartile range=41.4 months) was achieved where 44% of patients had serous tumor, followed by mucinous (23%), endometrioid (9%), mixed (9%), undifferentiated (8.5%) and clear cell tumors (5.3%). ER-alpha staining was associated with grade II-III tumors. Progesterone receptor staining was positively associated with a Body Mass Index≥25. Androgen receptor positivity was higher in serous tumors. In stand-alone analysis of receptor contribution to survival, estrogen-alpha positivity was associated with greater disease-free survival. However, there was no significant association between steroid receptor expression, HER2 status, or TNEOC status, and overall survival. Although estrogen-alpha, androgen receptor, progesterone receptor and the HER2 status were associated with key clinical features of the women and pathological characteristics of the tumors, these associations were not implicated in survival. Interestingly, women with TNEOC seem to fare the same way as their counterparts with non-TNEOC.

Tissue aluminum concentration does not affect the genomic stability of ERBB2, C-MYC, and CCND1 genes in breast cancer.

It has long been hypothesized that body tissue uptake of aluminum may have biological implications in breast cancer. In vitro and in vivo studies have shown that aluminum may trigger genomic instability by interfering with DNA strands. The objective of this study was to examine the relationship between aluminum concentrations in the peripheral and central areas of breast tumors with the instability of three key genes in breast cancer, ERBB2, C-MYC, and CCND1 and aneuploidy of the chromosomes harboring these genes. Tissue samples of 118 women treated for breast cancer were obtained. Evaluation of aluminum content was carried out using graphite furnace atomic absorption spectrometry. A tissue microarray slide containing the tumor samples was used in FISH assays to assess ERBB2, C-MYC, and CCND1 expressions as well as the statuses of their respective chromosomes 17, 8, and 11. Clinicopathological data were obtained from patient's records. Aluminum levels of >2.0 mg/kg were found in 20.3 and 22.1% of the central and peripheral breast tumor areas, respectively. Amplification and/or aneuploid-positive statuses for ERBB2/CEP17, C-MYC/CEP8, and CCND1/CEP11 were detected in 24, 36.7, and 29.3% of the tumors, respectively. We found that aluminum concentration was not related to these altered gene statuses. Our findings suggest that aluminum concentration does not affect genomic stability in breast tissues. Tissue microenvironment modifications, due to the presence of aluminum compounds, seem more appealing as a possible target for future studies to determine the implications of aluminum in breast carcinogenesis.

Immediate breast reconstruction with a Latissimus dorsi flap has no detrimental effects on shoulder motion or postsurgical complications up to 1 year after surgery.

BACKGROUND: Mastectomy negatively affects scapulothoracic and glenohumeral kinematics. Breast reconstructive methods such as the latissimus dorsi flap can result in anatomical modifications that may in theory further affect the shoulder apparatus. The purpose of this study was to examine the effects of latissimus dorsi flap reconstruction on the recovery of shoulder motion and other postsurgical problems during the first year after mastectomy. METHODS: This was a prospective cohort study of 104 consecutive mastectomies (47 with immediate latissimus dorsi flaps). Shoulder range of motion was assessed before and at 1, 3, 6, and 12 months after surgery. Pain, tissue adhesion, scar enlargement, and web syndrome were assessed during follow-up. RESULTS: There was a 30 percent decrease of shoulder range of motion 1 month after surgery, with gradual recovery over time. However, mean abduction and flexion capacities did not reach baseline levels and were on average 5 to 10 percent lower than baseline, even after 1 year. Over time, the latissimus dorsi flap was not associated with restriction of flexion or abduction. Scar enlargement (at the first month, p = 0.009) and tissue adhesion (at month 12, p = 0.032) were significantly less common in the latissimus dorsi flap group. CONCLUSIONS: The authors' study clearly suggests that the additional anatomical manipulation required for the latissimus dorsi flap procedure does not further affect shoulder kinematics and is associated with a lower incidence of tissue adhesion. CLINICAL QUESTION/LEVEL OF EVIDENCE: Therapeutic, II.

Aluminum concentrations in central and peripheral areas of malignant breast lesions do not differ from those in normal breast tissues.

BACKGROUND: Aluminum is used in a wide range of applications and is a potential environmental hazard. The known genotoxic effects of aluminum might play a role in the development of breast cancer. However, the data currently available on the subject are not sufficient to establish a causal relationship between aluminum exposure and the augmented risk of developing breast cancer. To achieve maximum sensitivity and specificity in the determination of aluminum levels, we have developed a detection protocol using graphite furnace atomic absorption spectrometry (GFAAS). The objective of the present study was to compare the aluminum levels in the central and peripheral areas of breast carcinomas with those in the adjacent normal breast tissues, and to identify patient and/or tumor characteristics associated with these aluminum levels. METHODS: A total of 176 patients with breast cancer were included in the study. Samples from the central and peripheral areas of their tumors were obtained, as well as from the surrounding normal breast tissue. Aluminum quantification was performed using GFAAS. RESULTS: The average (mean ± SD) aluminum concentrations were as follows: central area, 1.88 ± 3.60 mg/kg; peripheral area, 2.10 ± 5.67 mg/kg; and normal area, 1.68 ± 11.1 mg/kg. Overall and two-by-two comparisons of the aluminum concentrations in these areas indicated no significant differences. We detected a positive relationship between aluminum levels in the peripheral areas of the tumors, age and menopausal status of the patients (P = .02). CONCLUSIONS: Using a sensitive quantification technique we detected similar aluminum concentrations in the central and peripheral regions of breast tumors, and in normal tissues. In addition, we did not detect significant differences in aluminum concentrations as related to the location of the breast tumor within the breast, or to other relevant tumor features such as stage, size and steroid receptor status. The next logical step is the assessment of whether the aluminum concentration is related to the key genomic abnormalities associated with breast carcinogenesis.

HER2 expression in Brazilian patients with estrogen and progesterone receptor-negative breast carcinoma.

The aim of the study was to evaluate the relationship between clinical and pathological factors and survival in patients with double negative HER2-overexpressing carcinoma and triple negative carcinoma. One hundred and sixty-one (161) patients diagnosed with breast cancer negative for estrogen receptor (ER) and progesterone receptor (PR) were included. Of the total, 58 patients had double negative HER2-overexpressing (ER/PR-negative and HER2-positive) and 103 had triple negative (ER-negative, PR-negative and HER2-negative). ER and PR expression was assessed through immunohistochemistry (IHC) and HER2 expression was measured by immunohistochemistry and Fluorescent in situ Hybridization (FISH) analysis in tissue microarray. More than 80% had stages II and III disease and histologic grade III and nuclear grade 3. Patients with triple negative breast carcinoma had undifferentiated histologic types in 11% of cases and vascular invasion in 14.5%. Both groups had more than 50% visceral metastases. HER2 expression (p=0.42) and vascular invasion (p=0.05) did not interfere with survival. Survival of patients with Stages I-II disease was significantly longer than in those with Stage III disease both for double negative HER2-overexpressing carcinomas (p<0.0001) and triple negative carcinomas (p=0.03). The study shows that hormone receptor-negative breast carcinomas were undifferentiated and diagnosed at advanced stages and that HER2 expression was not associated with overall survival.

Metastatic mucinous carcinomas in the ovary: a practical approach to diagnosis related to gross aspects and to immunohistochemical evaluation.

Ovarian mucinous carcinomas are uncommon, and the differential diagnosis is metastatic carcinoma mainly from the gastrointestinal tract. The aim was to verify the importance of immunohistochemical reactions and the algorithm described in literature on the basis of laterality and tumor size. Twenty-five cases identified as metastatic mucinous adenocarcinomas were reviewed, along with clinical records; a tissue microarray was created, and immunohistochemical reactions for CK7, CK20, Ca125, hormonal receptors, WT1, DPC4, ß-catenin, and Cdx2 were determined. The median age was 51, and only 9 patients had a history of cancer. Sixteen patients (64%) had bilateral tumors, with sizes ranging from 5 to 36 cm (average, 20.5 cm); 9 (36%) had unilateral tumors varying from 5.5 to 38 cm (average, 21.8 cm). Algorithm agreement was 76%; most unilateral tumors were >13 cm. Common positive markers were Dpc4 (88%), Cdx2 (68%), CK20 (60%), and CK7 (44%). The useful markers were CK7, CK20, and Cdx2, although there were cases with overlapping results. The most common primary tumor was of colorectal origin (14 cases). The mean survival age was 32.6 mo. Although the proposed algorithm and immunohistochemical reactions are useful tools for diagnosis, some mucinous tumors cannot be definitively classified as primary or metastatic without further clinical evaluation, emphasizing the limits of this challenging diagnosis.

Expression of cyclooxygenase-2 (COX-2) and p53 in neighboring invasive and in situ components of breast tumors.

The aim of the study was to assess the relationship between the expression of COX-2 and p53, hormone receptors and HER-2 in the in situ (DCIS) and invasive components of ductal carcinomas (IDC) of the same breast. The expression of COX-2, p53, and hormone receptors was assessed in 87 cases of IDC with contiguous areas of DCIS. Results showed that there was no difference in COX-2 expression comparing the in situ and invasive components of the tumors. In the in situ component, there was a statistically borderline increase in p53 expression in tumors that also expressed COX-2. ER-positive specimens were more common in the group of tumors that expressed COX-2 in the invasive component. From this study we conclude that the expression of COX-2 was similar in the in situ and invasive components of the breast carcinomas. COX-2 positivity was marginally related with the expression of p53 in the in situ components, and with the ER expression in the invasive components.

Image compression impact on quantitative angiogenesis analysis of ovarian epithelial neoplasms.

OBJECTIVE: This study aims to investigate the impact of digital image compression on manual and semiautomatic quantification of angiogenesis in ovarian epithelial neoplasms (including benign, borderline, and malignant specimens). DESIGN: We examined 405 digital images (obtained from a previously validated computer-assisted analysis system), which were equally divided into 5 groups: images captured in Tagged Image File Format (TIFF), low and high compression Joint Photographic Experts Group (JPEG) formats, and low and high compression JPEG images converted from the TIFF files. MEASUREMENTS: Microvessel density counts and CD34 endothelial areas manually and semiautomatically determined from TIFF images were compared with those from the other 4 groups. RESULTS: Mostly, the correlations between TIFF and JPEG images were very high (intraclass correlation coefficients >0.8), especially for low compression JPEG images obtained by capture, regardless of the variable considered. The only exception consisted in the use of high compression JPEG files for semiautomatic microvessel density counts, which resulted in intraclass correlation coefficients of <0.7. Nonetheless, even then, interconversion between TIFF and JPEG values could be successfully achieved using prediction models established by linear regression. CONCLUSION: Image compression does not seem to significantly compromise the accuracy of angiogenesis quantitation in the ovarian epithelial tumors, although low compression JPEG images should always be preferred over high compression ones.

Reduced prevalence of the C825T polymorphism of the G-protein beta subunit gene in women with breast cancer.

OBJECTIVE: To evaluate the prevalence of the C825T polymorphism in the GNB3 gene in women with and without breast cancer and its possible association with clinical or pathological features of breast disease. SUBJECTS AND METHODS: We included 134 women with breast cancer and a control group of 129 healthy women. The case group responded to a questionnaire on lifestyle, reproductive factors and family history. Clinical data were also evaluated. The risk for cancer was calculated and PCR was carried out for the detection of the polymorphism. Statistical analysis was performed using the package R Environment, with confidence intervals of 95% and a significance level of 5% (p and lt;0.05). RESULTS: The frequency of the TT genotype was significantly greater in women of the control group (30.2%) than women with breast cancer (14.9%) (p=0.02). The polymorphism was not associated with clinical features, age at diagnosis (p=0.07), age at menarche (p=0.17), and age at menopause (p=0.60). The TT genotype did not have a higher frequency in patients with high BMI (p=0.98). The risk for cancer showed no correlation with the presence of the polymorphism. CONCLUSIONS: Our data indicate that the C825T polymorphism in the GNB3 gene has no relationship to the risk for breast cancer or the characteristics of the disease.

Hormonal contraceptives and the length of their use are not independent risk factors for high-risk HPV infections or high-grade CIN.

AIMS: To evaluate the role of hormonal contraceptives as a risk factor of high-risk human papillomavirus (HR-HPV), cervical intraepithelial lesions (CIN) and cervical cancer in our multi-center population-based LAMS (Latin American Screening) study. METHODS: A cohort study with >12,000 women from Brazil and Argentina using logistic regression to analyze the covariates of hormonal contraception (HOC - oral, injections, patches, implants, vaginal ring and progesterone intrauterine system) use followed by multivariate modeling for predictors of HR-HPV and CIN2+. RESULTS: HR-HPV infection was a consistent risk factor of high-grade CIN in all three groups of women. The length of HOC use was not significantly related to high-grade squamous intraepithelial lesions (HSIL)+ Pap (p = 0.069), LSIL+ Pap (p = 0.781) or ASCUS+ (p = 0.231). The same was true with the length of HOC use and histology CIN3+ (p = 0.115) and CIN2+ (p = 0.515). Frequently, HOC users have previously shown more HPV-related lesions, as well as lower HPV prevalence if they were current smokers. But HOC use and time of usage were not independent risk factors of either HR-HPV infection or high-grade CIN using multiple logistic regressions. CONCLUSIONS: No evidence was found for an association between the use of HOC with an increased risk for HR-HPV infection or high-grade CIN in this cohort.