Carmen Ortiz

Factors associated with breast cancer in Puerto Rican women

Background: Breast cancer (BC) is the most common cancer afflicting Puerto Rican women and accounts for more cancer-related deaths in this population than any other cancer. Methods: Demographic, anthropometric, family history, and lifestyle data, as well as DNA repair capacity (DRC), were compared in 465 BC cases and 661 controls. Crude and multiple logistic regression-derived adjusted odds ratios were used as indicators of the associations between BC and the variables under study. Results: A low DRC level, aging (>61 years), family history of BC, and low education level had statistically significant associations with increased BC risk. Endometriosis, full-term pregnancy at an earlier age, higher parity, hysterectomy before age 50, multivitamin and calcium intake, and longer duration of breastfeeding significantly decreased BC risk. Conclusions: This study discusses the major risk factors for BC in Puerto Rico (PR). Because many of these findings represent modifiable risk factors, they can translate into public health initiatives to lower BC risk. In addition, the possibility of using DRC as a simple screening tool for BC risk is explored.

Estrogen Receptor Expression Is Associated with DNA Repair Capacity in Breast Cancer.

Estrogen-receptor-positive (ER+) tumors employ complex signaling that engages in crosstalk with multiple pathways through genomic and non-genomic regulation. A greater understanding of these pathways is important for developing improved biomarkers that can better determine treatment choices, risk of recurrence and cancer progression. Deficiencies in DNA repair capacity (DRC) is a hallmark of breast cancer (BC); therefore, in this work we tested whether ER signaling influences DRC. We analyzed the association between ER positivity (% receptor activation) and DRC in 270 BC patients, then further stratified our analysis by HER2 receptor status. Our results show that among HER2 negative, the likelihood of having low DRC values among ER- women is 1.92 (95% CI: 1.03, 3.57) times the likelihood of having low DRC values among ER+ women, even adjusting for different potential confounders (p<0.05); however, a contrary pattern was observed among HER2 positives women. In conclusion, there is an association between DRC levels and ER status, and this association is modified by HER2 receptor status. Adding a DNA repair capacity test to hormone receptor testing may provide new information on defective DNA repair phenotypes, which could better stratify BC patients who have ER+ tumors. ER+/HER2- tumors are heterogeneous, incompletely defined, and clinically challenging to treat; the addition of a DRC test could better characterize and classify these patients as well as help clinicians select optimal therapies, which could improve outcomes and reduce recurrences.

Cytotoxicity and Genotoxicity Assessment of Sandalwood Essential Oil in Human Breast Cell Lines MCF-7 and MCF-10A

Sandalwood essential oil (SEO) is extracted from Santalum trees. Although α-santalol, a main constituent of SEO, has been studied as a chemopreventive agent, the genotoxic activity of the whole oil in human breast cell lines is still unknown. The main objective of this study was to assess the cytotoxic and genotoxic effects of SEO in breast adenocarcinoma (MCF-7) and nontumorigenic breast epithelial (MCF-10A) cells. Proteins associated with SEO genotoxicity were identified using a proteomics approach. Commercially available, high-purity, GC/MS characterized SEO was used to perform the experiments. The main constituents reported in the oil were (Z)-α-santalol (25.34%), (Z)-nuciferol (18.34%), (E)-β-santalol (10.97%), and (E)-nuciferol (10.46%). Upon exposure to SEO (2–8 μg/mL) for 24 hours, cell proliferation was determined by the MTT assay. Alkaline and neutral comet assays were used to assess genotoxicity. SEO exposure induced single- and double-strand breaks selectively in the DNA of MCF-7 cells. Quantitative LC/MS-based proteomics allowed identification of candidate proteins involved in this response: Ku70 (p = 1.37E − 2), Ku80 (p = 5.8E − 3), EPHX1 (p = 3.3E − 3), and 14-3-3ζ (p = 4.0E − 4). These results provide the first evidence that SEO is genotoxic and capable of inducing DNA single- and double-strand breaks in MCF-7 cells.

Variability in DNA Repair Capacity Levels among Molecular Breast Cancer Subtypes: Triple Negative Breast Cancer Shows Lowest Repair

Breast cancer (BC) is a heterogeneous disease which many studies have classified in at least four molecular subtypes: Luminal A, Luminal B, HER2-Enriched, and Basal-like (including triple-negative breast cancer, TNBC). These subtypes provide information to stratify patients for better prognostic predictions and treatment selection. Individuals vary in their sensitivities to carcinogens due to differences in their DNA repair capacity (DRC) levels. Although our previous case-control study established low DRC (in terms of NER pathway) as a BC risk factor, we aim to study this effect among the molecular subtypes. Therefore, the objectives of this study include investigating whether DRC varies among molecular subtypes and testing any association regarding DRC. This study comprised 267 recently diagnosed women with BC (cases) and 682 without BC (controls). Our results show a substantial variability in DRC among the molecular subtypes, with TNBC cases (n = 47) having the lowest DRC (p-value < 0.05). Almost 80 percent of BC cases had a DRC below the median (4.3%). Low DRC was strongly associated with the TNBC subtype (OR 7.2; 95% CI 3.3, 15.7). In conclusion, our study provides the first report on the variability among the molecular subtypes and provides a hypothesis based on DRC levels for the poor prognosis of TNBC.

Evaluation of Antioxidant Capacity of Solanum sessiliflorum (Cubiu) Extract: An In Vitro Assay

Cubiu is a vegetable of Solanaceae family, native to the Amazon, which is widely distributed through Brazil, Peru, and Colombia. It is used in food, medicine, and cosmetics by native populations. Research has shown that cubiu extracts have antioxidant activities with great biological relevance. We performed a phytochemical screening to identify the main chemical groups that could confer antioxidant activity to this extract. Several tests and qualitative precipitation specific staining for major classes of secondary metabolites were used. Antioxidant capacity in vitro tests (DPPH and ABTS) were also used to assess the extracts ability to sequester free radicals of 70% hydroethanolic and aqueous extracts of cubiu flour. Alkaloids, organic acids, phenols, flavonoid glycosides, and coumarins were found in the hydroethanolic extract while the aqueous extract presented anthocyanins, gums, tannins and mucilage, amino groups, and volatile and fixed acids. For in vitro tests, the IC50 value obtained in the DPPH assay was 606.3 ± 3.5 μg/mL while that for the ABTS assay was 290.3 ± 10.7 µg/mL. Although cubiu extracts present chemical compounds directly related to antioxidant activity, our results show that it has a low antioxidant activity. Additional studies will be needed to isolate and characterize specific compounds to further assess antioxidant activity.

Effects of Astrocaryum aculeatum Meyer (Tucumã) on Diet-Induced Dyslipidemic Rats

An in vivo study was conducted to assess the effects of the consumption of Astrocaryum aculeatum Amazon Meyer (tucumã) in the treatment of diet-induced dyslipidemia in sedentary and exercised Wistar rats. With an average weight of 350 grams, 40 male rats were divided into 4 subgroups of 10. The sedentary control group (SCG) was fed with commercial feed, while the sedentary treatment group (STG) was fed with a ration of tucumã. In addition to the sedentary groups, two exercise groups were formed. The Exercised control group (ECG) was fed with commercial food and the exercised treatment group (ETG) was fed with a ration of tucumã. Body weight gain and food intake were monitored during the experiment. Plasma was analyzed for cholesterol, triglycerides, HDL-C, LDL-C, VLDL, total protein, glucose, insulin, and leptin concentrations. Our results show that the ECG group tended to consume more food, while the groups that were fed with tucumã pulp (STG and ETG) presented a greater tendency to gain body mass. ECG group showed a tendency towards a higher concentration of cholesterol in plasma, while STG and ETG presented higher absolute values for triglycerides and VLDL. No hypolipiemic effect was observed related to tucuma ingestion.

High DRC Levels Are Associated with Let-7b Overexpression in Women with Breast Cancer

Nucleotide Excision Repair (NER) is a critical pathway involved in breast cancer (BC). We have previously published that a low DNA repair capacity (DRC) is associated with a higher risk of BC in Puerto Rican women. Let-7b belongs to a miRNA family with tumor suppressor activity that targets oncogenes. We isolated miRNAs from plasma of 153 Puerto Rican women with and without BC. DRC was measured in lymphocytes by means of a host cell reactivation assay. These women were divided into four groups according to their DRC level: High (>3.8%) and low (<3.8%). The four groups consisted of BC patients with high (n = 35) and low (n = 43) DRC and controls with high (n = 39) and low (n = 36) DRC. Epidemiologic data were collected at initial BC diagnosis and almost five years after diagnosis. A significant difference in Let-7b expression was found in BC patients with high DRC versus the remaining groups (p < 0.001). Thus, our data reveal a possible role of Let-7b on DRC during breast carcinogenesis. Our study is innovative because it provides the first evidence that Let-7b may play role in DRC regulation (through the NER repair pathway) in BC.

Abstract 3286: Correlation between vitamin D levels and DNA repair capacity in breast cancer patients stratified by molecular subtypes

Vitamin D exists as vitamin D2 and D3, which are metabolized to 25-hydroxyvitamin D [25(OH)D], the major circulating vitamin D metabolite. Besides its physiological functions, vitamin D levels have also been studied as a risk factor for several hormonal cancers including breast cancer (BC). Worldwide, BC accounts for nearly a quarter of all cancers in women. Nutritional studies report that vitamin D intake is associated with a lower BC risk. Several discrepancies exist regarding the role of serum vitamin D in BC risk. While some studies report BC risk reduction by vitamin D only in premenopausal, others propose that it only occurs in postmenopausal women. BC tumors may (+) or may not (-) have three hormonal receptors: estrogen (ER), progesterone (PR), and HER2. Based on their status, four principal molecular BC subtypes have been identified: luminal A (ER+/−, PR+/−, HER2-), luminal B (ER+/−, PR+/−, HER2+), HER2+ (ER−, PR−, HER2+), and triple negative (TN) (ER−, PR−, HER2−). If BC is analyzed in terms of molecular subtypes, low vitamin D levels have been associated with aggressive phenotypes and worse prognosis. Vitamin D also influences estrogen synthesis. Since we have previously shown that a low DNA repair capacity (DRC), measured through the nucleotide excision repair pathway, is a risk factor for BC and vitamin D has also been found to affect DNA repair, the focus of this study is to examine the role of plasma vitamin D levels and DRC in BC. The main aim is to elucidate whether there is an association between vitamin D and DRC levels among the four molecular BC subtypes. We hypothesize that a negative correlation between 25(OH)D and DRC levels will be observed among these subtypes. As an initial effort, 47 BC cases and 20 controls without BC were selected from our large BC cohort. DRC was measured in lymphocytes of untreated women using the host cell reactivation assay. Pathology reports were examined to divide BC cases according to their molecular BC subtype: luminal A (n=13), luminal B (n=11), HER2+ (n=10), and TN (n=13). Plasma 25(OH)D levels were measured using the UniCel DxC System at a CLIA-certified lab. Our results show a negative correlation between 25(OH)D and DRC levels (p=0.04). Statistically significant differences were found for vitamin D levels among the different groups (p=0.0019, ANOVA). Moreover, higher 25(OH)D levels (47.97±2.4 ng/mL) were found in ER- BC cases (p=0.03, t-test). When comparing vitamin D levels in BC subtypes, a significant difference was found in HER2+ and TN groups when compared with the control group (p Citation Format: Carmen Ortiz, Jarline Encarnacion, Ralphdy Vergne, Wanda Vargas, Jaime Matta. Correlation between vitamin D levels and DNA repair capacity in breast cancer patients stratified by molecular subtypes [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 3286. doi:10.1158/1538-7445.AM2017-3286

Abstract 1899: Let-7b overexpression is associated with higher nucleotide excision repair pathway in women with breast cancer

Nucleotide Excision Repair (NER) is a critical pathway involved in breast cancer (BC). We have previously published that a low DNA repair capacity (DRC) is associated with a higher risk of BC in Puerto Rican women. In recent years we have focused our investigations on microRNAs (miRNAs) that are differentially associated with a low and a high DRC in women with BC and controls. A discovery experiment with 29 BC cases and 27 controls produced 12 candidate miRNAs associated with DRC including let-7b. The main objective of this study was to elucidate if there is a correlation between let-7b expression and specific DRC levels. Let-7b belongs to a miRNA family with tumor suppressor activity that targets oncogenic genes such as Ras, Myc, and HmgA2. DRC was measured in lymphocytes by means of a host cell reactivation assay with a luciferase reporter gene. We isolated miRNAs from plasma of 145 Puerto Rican women with and without BC (recently diagnosed, untreated cases and controls) using the miRNeasy kit (Qiagen). These women were divided into four groups according to their DRC level: high (>3.8%) and low ( Citation Format: Jarline Encarnacion, Carmen Ortiz, Ralphdy Vergne, Wanda Vargas, Jaime L. Matta. Let-7b overexpression is associated with higher nucleotide excision repair pathway in women with breast cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1899.

Abstract 3851: Low educational level is associated with a reduced DNA repair capacity and higher risk of breast cancer

Proceedings: AACR 106th Annual Meeting 2015; April 18-22, 2015; Philadelphia, PA BACKGROUND: Breast Cancer (BC) is the leading cause of cancer deaths among females in Puerto Rico. Educational level is well established as a modifier of cancer risk. We have undertaken a large population study and have shown that a low DNA Repair Capacity (DRC) is an important risk factor for BC. It is well established that DRC is regulated by genetic, epigenetic and environmental factors. We have investigated the relationship between educational level and DRC. The purpose of this study was to evaluate the role of educational level as a modifier of the association between DRC and risk of BC in Puerto Rican women. METHODS: A total of 488 women with histopathologically confirmed BC and 607 controls were selected for the analysis. DRC was measured in lymphocytes using a host cell reactivation assay. Pathology reports were used to obtain tumor size. DRC levels were divided into low, medium and high using tertiles. Multiple logistic regression was used to assess the association among educational level and BC risk factors adjusted by DRC. For the analysis of tumor size and DRC, educational levels were divided into primary education, high school, and associate or more. Analysis of variance was performed using the Kruskal-Wallis test. RESULTS: Women with lower educational level (elementary education) had significantly higher risk of BC (OR: 5.9, 95%CI: 2.3, 14.9) followed by high school (OR: 1.4, 95%CI: 1.1, 2.1) and associate degree (OR: 1.4, 95%CI: 0.9, 2.2) using bachelors degree as a referent. Women with low education had 8.9 times the odds of having low DRC (95%CI: 5.2, 15.2), while women with high educational level had 11.7 times the odds for a low DRC (95%CI: 7.7, 17.7). The association and tumor size (cm) and educational level showed that as educational level increased, women had smaller tumor sizes (Kruskal-Wallis test p = 0.031). CONCLUSIONS: This study showed that women with the lowest levels of education, in general, had a lower DRC and significantly higher risk of BC and of having larger breast tumors when compared to controls. This study provides new insights as to how educational level can influence BC risk. This association is controversial because some studies attribute a high educational level to high risk of BC while others present the opposite. Because our study uses a biological variable (DNA repair) to measure risk, it reduces ambiguity in terms of the criterion to estimate risk. Those with the highest levels of education had smaller tumors; therefore consequently better chances for early treatment and increased chances of survival. Because education is a modifiable lifestyle factor, this study provides data that can be utilized to identify women with a higher risk of developing BC in order to design more effective BC screening, and diagnosing BC an earlier stage. Supported by grants from the NCI Diversity Training Branch through the NIH-MBRS Program grants S06 GM008239-20, 9SC1CA182846-04 to PSMHS through JM and MBRS-RISE GM082406 through CO. Citation Format: Luisa Morales, Manuel Bayona, Carmen Ortiz, Damian Adams, Carolina Alvarez-Garriga, Jaime L. Matta. Low educational level is associated with a reduced DNA repair capacity and higher risk of breast cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3851. doi:10.1158/1538-7445.AM2015-3851