Padma P. Tadi Uppala

Selective Inhibition of Cell Proliferation by Lycopene in MCF-7 Breast Cancer Cells In vitro: A Proteomic Analysis

Lycopene, a red pigmented carotenoid present in many fruits and vegetables such as tomatoes, has been associated with the reduced risk of breast cancer. This study sought to identify proteins modulated by lycopene during cell proliferation of the breast cancer cell line MCF‐7 to gain an understanding into its mechanism of action. MCF‐7 breast cancer cells and MCF‐10 normal breast cells were treated with 0, 2, 4, 6, 8, and 10 μM of lycopene for 72 h. 3‐(4,5‐Dimethylthiazol‐2‐yl)‐2,5‐diphenyltetrazolium bromide (MTT) tetrazolium reduction assay was used to measure cell proliferation and two‐dimensional fluorescence difference gel electrophoresis to assess the changes in protein expression, which were identified using MALDI‐ToF/ToF (matrix‐assisted laser desorption ionization tandem time‐of‐flight) and Mascot database search. MTT and cell proliferation assays showed that lycopene selectively inhibited the growth of MCF‐7 but not MCF‐10 cells. Difference gel electrophoresis analysis revealed that proteins in the MCF‐7 cells respond differently to lycopene compared with the MCF‐10 cells. Lycopene altered the expression levels of proteins such as Cytokeratin 8/18 (CK8/18), CK19 and their post translational status. We have shown that lycopene inhibits cell proliferation in MCF‐7 human breast cancer cells but not in the MCF‐10 mammary epithelial cells. Lycopene was shown to modulate cell cycle proteins such as beta tubulin, CK8/18, CK19 and heat shock proteins. Copyright

Abstract C85: Community-based participatory research leads to sustainable lifestyle intervention program for reducing breast cancer risk among African American and Latina women

Minority women are underrepresented in clinical research, which impedes progress in understanding and eliminating health disparities. Reducing health disparities is an important goal of public health community. Recent data suggests that community-based participatory research (CBPR) which systematically involves the affected communities in each stage of the research process-inception, planning, recruitment, conduct and education is the key to reducing health disparities. Between 2001 and 2005 approximately 10,000 invasive and 2,000 in situ breast cancer cases were reported in Region 5 of the San Bernardino County women in California. Although data reveals that the age adjusted incidence rate of 23.7 for in situ breast cancer cases in the Region 5 population is lower than the state-wide rate of 27.3, the number of in situ cases in the area rose from 9.6 in 1988 to 24.8 in 2007. Obesity rates for African American (AA) and Latina adults in the San Bernardino County are 34%. The emergence of the obesity epidemic worldwide has been associated with increases in metabolic syndrome, breast cancer, and type 2 diabetes in the industrialized countries. The proposed pilot study used a CBPR method to study women9s perceptions, beliefs and attitudes towards participating in a clinical breast cancer study that involved blood. Methods: A community-based participatory mixed-methods approach, including a 6-month lifestyle intervention education program was employed. Participants were recruited using purposive convenience sampling that evaluated educational program designed to increase women9s knowledge of positive attitudes and participation in breast cancer clinical studies and simple changes in lifestyle that result in reduced risk for breast cancer. To evaluate the effectiveness of the educational intervention, we used knowledge, attitudes, behavior, and perception-based questionnaire, and in-depth interviews to assess changes before and after the educational experience. Results: Fifty eight women that included 36 African American and 22 Latina women participated in the educational program, out of which twenty five women were recruited into the lifestyle intervention program that required providing blood to test for metabolic syndrome and risk for breast cancer. All the women in the intervention study were willing to provide blood for the study. Seventy six percent of the women in the lifestyle intervention program were either obese or experienced metabolic syndrome. At the end of six months, 78% of the women who participated in the study reduced their body fat by 2% change, by reducing dietary fat consumption. Approximately 93% of the women increased servings of fruits and vegetables in their diet and engaged in some form of exercise after their enrollment into the study. This program is currently sustained by community support groups, after the end of the grant. The project director meets with the women once a month and monitors their life style changes. Conclusion: Community-based participatory research projects that engage community partners through the entire process of research planning and conduct may result in effective sustainable projects that will result in resilient communities. Funding supported by Kaiser Permanente Fontana Community Benefit Grant. Citation Format: Padma Pauline Tadi Uppala, Padma Pauline Tadi Uppala, Loistine Herndon, Hildemar Dos Santos, Amanda Dupre, Persila Mohammadnia, Maheswari Senthil. Community-based participatory research leads to sustainable lifestyle intervention program for reducing breast cancer risk among African American and Latina women. [abstract]. In: Proceedings of the Sixth AACR Conference: The Science of Cancer Health Disparities; Dec 6–9, 2013; Atlanta, GA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2014;23(11 Suppl):Abstract nr C85. doi:10.1158/1538-7755.DISP13-C85

Abstract 4999: Stress, spiritual wellbeing and cancer risk among diverse racial faith-based communities: Elevated levels of stress proteomic biomarkers in breast cancer patients

Spiritual wellbeing is associated with decreased risk for cancer. Stress and depression are common among cancer patients and may be inversely associated with spiritual wellbeing. The purpose of the study is to examine if stress and depression as indicators of cancer risk are lower in a racially diverse faith-based community. Methods: 752 individuals from a faith-based community completed a behavioral assessment survey that mapped the current patterns of behavior in key categories that include: stress, depression, exercise and nutrition using the E-wellness platform FitThumb. To identify stress biomarkers in cancer patients we examined levels of stress serum proteomic biomarkers that were previously identified in our study by proteomic profiling using 2D-DIGEMS analysis and a subset of samples by shotgun LCMS technology. The serum proteomic study included serum samples from 15 African American breast cancer patients and 12 healthy controls who were from a faith-based community. Results: The behavioral assessment surveys that included 752 individuals who were racially diverse showed a low stress risk of 47.74% vs 8.24 % of high stress risk for chronic disease; and low depression risk of 85.11% vs 1.99% high risk for chronic disease. Elevated levels of stress and inflammatory serum proteomic biomarkers such as ceruloplasmin known to increase in stressed animals and humans; heptaglobin, apolipoproteins, and heat shock proteins were significantly elevated in breast cancer patients compared to healthy controls. Conclusions: Our results indicate that spiritual well-being is associated with significantly low stress and depression in a faith-based community regardless of race or ethnicity, posing a low risk for cancer as shown in previous Adventist Health Cohort studies. Future efforts will focus on validating and identifying panel of biomarkers from this cohort to gain insight into their role(s) in the mechanisms of stress hormones and cancer risk. Funded by Susan G Komen for the cure. Citation Format: Padma P. Tadi Uppala, Gretchen Krivak, Sherine Brown-Fraser, Dixon Anjejo, Alfredo Mejia, Dominique wakefield, Kumar Kolli. Stress, spiritual wellbeing and cancer risk among diverse racial faith-based communities: Elevated levels of stress proteomic biomarkers in breast cancer patients [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 4999. doi:10.1158/1538-7445.AM2017-4999

Abstract 3936: Identification and validation of the potential biomarker insulin-like growth factor binding protein acid-labile subunit for breast cancer in African American women

Breast cancer is the most frequently diagnosed cancer in women, with an estimated 40, 730 breast cancer deaths in the US in 2015. African American (AA) women have a lower breast cancer incidence rate, but a higher breast cancer death rate, than non-Hispanic White women. Research indicates that breast tumor biology in AA women is different from that in Caucasian women. AA women are more likely to be diagnosed with breast cancer at an earlier age and with more aggressive form of the disease, characterized by higher grade and negative estrogen and progesterone receptor status. Because of the aggressive nature of these tumors and current lack of targeted therapies, identification of novel relevant protein markers is of great importance. The purpose of this study was to validate serum proteins that were previously identified by serum proteomic profiling in 22 serum samples by 2D-DIGE/MS analysis and a subset of samples by shotgun LC/MS technology. Methods and new data: The current study included serum samples from 15 African American breast cancer patients and 12 healthy controls. Patients were grouped into triple negative (TN), HER2 and Luminal A and B subtypes. Proteins of biological significance were validated using western blot analysis. For ceruloplasmin, and insulin-like growth factor binding protein acid-labile subunit (IGFBP-ALS), one-way ANOVA was used to compare mean density among the three groups. For Vitamin D Binding protein (VDB), a two-sample t-test was used to compare the density between the groups. Due to the small sample size, we have also conducted nonparametric tests. IGFBP-ALS was significantly lower in triple negative breast cancer patients (p = 0.016) and in HER2 (p = 0.025) subtypes. There was no significant difference in VDB protein in the luminal A and B subtypes (p = 0.98). Future efforts will focus on validating the identified panel of biomarkers to gain insight into their role(s) in the etiology of aggressive breast tumors. Funded by Susan G Komen for the Cure and SEED grant SPH. Citation Format: Padma P. Tadi Uppala, Carlos Garberoglio, Sharon Lum, Willie Davis, Hon-Chiu Eastwood Leung, Michael Liebman, Keiji Oda, Utkarsh P. Patel. Identification and validation of the potential biomarker insulin-like growth factor binding protein acid-labile subunit for breast cancer in African American women. [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 3936.

Abstract 1783: Heterogeneity of triple negative breast cancer across race and ethnicities: Indian women at higher risk for early onset breast cancer

Breast cancer is a heterogeneous disease with several subtypes presenting various morphological and molecular features, and response to therapy. While targeted therapies are available for estrogen receptor positive and HER2-positive breast tumors, triple-negative breast cancer (TNBC) which are negative for ER, progesterone receptor PR and HER2 receptors lack suitable targeted therapies. TNBC is one of the most aggressive breast cancer subtypes and poses a clinical challenge as they lack suitable targeted therapies. While a majority of breast cancer patients in the US are postmenopausal, more than 80% of Indian patients are younger than 60 years of age, with median age being 40-55, presenting with larger tumor size, poor tumor grade, and low rates of hormone-receptor positive status. Literature review suggests that none of the standard risk factors for breast cancer had any significant associations for the early onset breast cancers among Indian women. Recent studies on breast cancer subtypes across culture and geographical regions in India have indicated that incidence of early onset breast cancer, TNBC in particular is rising at alarming rates among Indian women. The interaction of race and ethnicity with age, molecular profiles and lifestyles has contributed significantly to the heterogeneity of TNBC breast cancer. The purpose of this study is to conduct a systematic literature review to identify factors that may increase risk for breast cancer among young Indian women. Methods: All the major databases including Web of Science and PubMed were used to search the literature for early onset breast cancer among Indian women. In a study that analyzed molecular subtypes in early onset breast cancer among various races that included Indian, Chinese, non-Hispanic White (NHW), African American (AA), and Hispanic women, incidence of TNBC was significantly higher (p = 0.0369) with early onset (40 years and younger) in Indian women. Incidence of HER2 over-expression subtype was also highest among Indian women. Conclusion: Early onset breast cancer is increasing rapidly among Indian women, along with obesity and diabetes. It is suggested that epigenetic factors that include sedentary lifestyles, lack of exercise, fatty diets and increasing obesity with underlying molecular mechanisms may contribute to early onset breast cancer among Indian women. Future studies that focus on racial and ethnic differences in genetic, reproductive, lifestyle and environmental exposures of TNBC pathways will offer unique biomarkers, targeted therapies and clinical trial design leading to personalized medicine. Citation Format: Padma P. Tadi Uppala, Maheswari Senthil, Utkarsh P. Patel, Sharon Lum, Carlos Garberoglio, Larry Beeson, John Morgan. Heterogeneity of triple negative breast cancer across race and ethnicities: Indian women at higher risk for early onset 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 1783.

Abstract 2442: Community-based participatory research leads to sustainable lifestyle intervention program for reducing breast cancer risk among African American and Latina women

Proceedings: AACR Annual Meeting 2014; April 5-9, 2014; San Diego, CA Breast cancer mortality rates are higher among African American (AA) women in San Bernardino County (SBC) at 36.31 per 100,000 women compared with white women in the same region, as well as with California breast cancer rates and the national averages. In addition, approximately 30% of individuals in SBC are obese. Mortality rates for diabetes in SBC between 2009 -2011 were 33 per 100,000 adults. Recent literature suggests an association between obesity, diabetes, metabolic syndrome and triple-negative breast cancer (TNBC). The insulin-leptin adiponectin axis has been implicated in these breast tumors that are insensitive to estrogen. TNBC is common among premenopausal AA and Latina women and is unclear whether overweight women or those with metabolic syndrome are at increased risk for breast cancer. Engaging minority AA and Latina women in clinical research has been difficult. Mistrust of doctors and scientists is reported as a barrier to participation. In this pilot study, community-based participatory (CBPR) methods were used to study womens perceptions, beliefs and attitudes towards participating in a breast cancer clinical study. Methods: A CBPR mixed-methods approach, including a 6-month lifestyle intervention education program was employed. Participants were recruited using purposive convenience sampling. To evaluate the effectiveness of the lifestyle intervention educational program, we used knowledge, attitudes, behavior, and perception-based questionnaire to assess changes before and after the educational experience. Results: Fifty eight women that included 36 African American and 22 Latina women participated in the educational program, out of which twenty five women were recruited into the lifestyle intervention program that required providing blood to test for metabolic syndrome and risk for breast cancer. All the women in the intervention study were willing to provide blood for the study. Seventy six percent of the women in the lifestyle intervention program were either obese or experienced metabolic syndrome. At the end of six months, 78% of the women who participated in the study reduced their body fat by 2% change, by reducing dietary fat consumption. Approximately 93% of the women increased servings of fruits and vegetables in their diet and engaged in some form of exercise after their enrollment into the study. This program is currently sustained by community support groups, after the end of the grant. Conclusion: Community-based participatory research projects that engage community partners through the entire process of research planning and conduct may result in effective sustainable projects that will result in resilient communities. Funding supported by Kaiser Permanente Fontana Community Benefit Grant. Citation Format: Padma P. Tadi Uppala, Hildemaar Dos Santos, Amanda Dupre, Persila Mohammadnia, Maheswari Senthil. Community-based participatory research leads to sustainable lifestyle intervention program for reducing breast cancer risk among African American and Latina women. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2442. doi:10.1158/1538-7445.AM2014-2442

Health effects of environmental DDT exposure: An epidemiological and serum proteomic study

P offer a variety of benefits to the society such as increased crop production and decreased insect infestations. However, when used improperly, pesticides have the potential for causing harm. Approximately, 10,000-20,000 physiciandiagnosed pesticide poisonings occur each year among US agricultural workers. Although banned in the US in 1973, DDT, an organochlorine pesticide finds its way into the food chain as it is used in many parts of the world. The purpose of the study was to examine the health effects of DDT in a predominantly African American population who were exposed to highest levels of DDT in the US by consumption of DDTcontaminated fish. A cross-sectional study was conducted on 234 African Americans. Logistic Regression was used to calculate risk of morbidity from consumption of DDTcontaminated fish. Serum proteomic study included 21 breast cancer cases and 29 controls. Protein expression data was obtained by 2D DIGE analysis. Differentially expressed DIGE spots were excised for protein identification. Spots were digested with trypsin and the peptides were identified by QTOF LC/MS/MS or MALDI TOF analysis. Results: A non-significant increase in risk for hypertension in those who consumed DDT contaminated fish was observed (OR 1.672; 95% CI 0.324-8.628). Serum proteomic study yielded significantly increased levels of fibrinogen gamma in exposed individuals. Gamma fibrinogen is associated with increased risk of coronary artery disease. Future efforts will focus on validation of gamma fibrinogen in DDT exposed individuals. Funded by Susan G. Komen for the cure.

Abstract P5-14-01: Accrual of underrepresented minority women to breast cancer clinical trials in Inland Empire, California.

Withdrawn by Author Citation Information: Cancer Res 2012;72(24 Suppl):Abstract nr P5-14-01.

Abstract A58: Role of insulin-like growth factor binding protein acid labile complex in ER-PR- Her2+ breast cancer in African American women.

The emergence of the obesity epidemic worldwide has been associated with increases in breast cancer and type 2 diabetes. Symptoms associated with obesity collectively known as metabolic syndrome have been associated with triple negative breast tumors that are common among premenopausal women, especially those of African American (AA) descent. The insulin-leptin adiponectin axis has been implicated in these breast tumors that are insensitive to estrogen. It is suggested that insulin, insulin like growth factors and epidermal growth factors mediate the interactions between these hormones. Previous studies have shown that insulin like growth factor 2 (IGF-2) synergistically cross-talks with estrogen receptor (ER) alpha and ER-beta to promote estrogen independent breast cancer progression. This suggests that IGF-2 can activate insulin like growth factor I (IGF-1) receptor and insulin receptor to activate both ER alpha and ER beta in breast cancer cells leading to proliferation and tumorigenesis. The purpose of this study was to find the the role of insulin-like growth factor binding protein acid labile complex (IGFBP-ALS) identified in our previous serum proteomic pilot study that included six AA breast cancer women and six healthy AA controls. Methods: The serum samples were processed on IgY12 (Beckman) antibody column to remove the top 12 proteins. The flow through fraction was digested using trypsin for LC/MS analysis. The protein digest serum samples were analyzed in triplicate on a high performance LTQFT (Thermo Electron) mass spectrometer coupled to an online Surveyor LC system equipped with an autosampler. Samples were loaded onto a trap column using a sample pump. An MS pump was used for eluting the peptides onto an analytical column for further separation and online nano-ESI LC/MS/MS analysis. The raw MS data files were imported into DeCyderMS (GE Healthcare) module and data were processed for peptide/protein expression analysis. Results: DeCyderMS analysis revealed significant differences in the expression of 215 peptides between the two groups (t-test: 2). Most of these peptides were mapped to 28 proteins in the Swiss-Prot database. DecyderMS analyses on all the replicates from each sample are underway to identify the peptide pattern differences between the healthy and cancer samples. Among other differentially expressed proteins IGFBP-ALS was significantly low in sera from African American women with breast cancer with tumors possessing the ER-PR-Her2+receptor status. In a previous study, 60% of the women from the same cohort experienced metabolic syndrome. Conclusions: It is suggested that IGFBP-ALS is necessary for the binding of IGF-1 or IGF-2 to the plasma IGF protein BP-53. Low levels of IGFBP-ALS subunit in the sera of ER-PR-HER2+ AA cancer patients could serve as biomarker for early-stage, breast cancer with aggressive features. Additionally this protein could reveal an association between obesity and breast cancer. Efforts are currently underway to validate this biomarker to gain insight into the etiology of aggressive breast tumors in AA women. Funded by The Susan G. Komen For the Cure Breast Cancer Foundation. Citation Format: Padma Uppala. Role of insulin-like growth factor binding protein acid labile complex in ER-PR- Her2+ breast cancer in African American women. [abstract]. In: Proceedings of the Fifth AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2012 Oct 27-30; San Diego, CA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2012;21(10 Suppl):Abstract nr A58.

Accrual of under-represented minority women to breast cancer clinical trials in Inland Empire, California.

64 Background: Ethnic minority women are currently underrepresented in breast cancer clinical trials. The age-adjusted incidence and mortality rates for breast cancer are higher in Riverside and San Bernardino Counties (Inland Empire), California, than the statewide average. During the 10 years of enrollment at Loma Linda University Cancer Center (LLUCC) between 1999 and 2009, a total of 151 patients were enrolled to breast cancer clinical trials. Among the enrolled patients, only 26.7% were ethnic minority women in contrast with the 59% of ethnic minority women living in Inland Empire. METHODS A countywide breast cancer awareness campaign using billboard, print ads, and newsletters, combined with direct mailing sent to regional physician offices and breast imaging centers was launched. We specifically addressed previously identified barriers to participation in clinical trials, such as lack of medical insurance, time, invitation, awareness of available studies, access to the institution or doctors conducting the trial, and coordination among clinics. In addition, we utilized a culturally sensitive multidisciplinary team to address the breast cancer care for minority women. RESULTS During the two year period since implementation of the regional culturally sensitive breast cancer awareness campaign, we successfully enrolled 32 patients into breast cancer clinical trials. Minority breast cancer patient accrual increased from a 10-year average of 26.7% to 46.9% in comparison with non-Hispanic white patients (p=0.02). The accrual rate of Hispanic population increased from 10% to 21.9%. The accrual rate of black patients increased from 5.3% to 12.5%. CONCLUSIONS The use of materials specifically targeting known barriers to participation in clinical trials successfully improved accrual of minority women, and has the potential to improve the cure rate, decrease disparities in breast cancer care, and eventually decrease the breast cancer burden particularly in underserved minority groups in Inland Empire, California.