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INTRODUCTION: Adjuvant therapy trials required completion lymph node dissection (CLND) for sentinel lymph node (SLN)-positive melanoma prior to systemic treatment, but nodal surveillance without CLND is now common. For patients receiving adjuvant therapy without CLND, patterns of recurrence are unknown and the value of regional nodal ultrasound alongside cross-sectional imaging is not well-defined. METHODS: In a retrospective cohort of SLN-positive melanoma patients managed with nodal surveillance from June 2014-June 2019, we evaluated the association between adjuvant treatment and location of first recurrence (loco-regional, nodal, distant, or multi-site) using chi-squared tests. We compared methods of recurrence detection and cost by surveillance intensity using chi-squared and Dunn’s tests. RESULTS: Among 177 nodal surveillance patients, 66 (37%) received adjuvant therapy. Median follow-up was 24 months, during which 48 patients (27%) recurred. Adjuvant treatment did not alter patterns of initial recurrence (p=0.76). Adjuvant therapy recipients more often had both nodal ultrasound and cross-sectional imaging surveillance (p

The National Lung Screening Trial (NLST) demonstrated that screening with low-dose computed tomography (LDCT) is associated with a 20% reduction in lung cancer mortality. One potential limitation of LDCT screening is overdiagnosis of slow growing and indolent cancers. In this study, peritumoral and intratumoral radiomics was used to identify a vulnerable subset of lung patients associated with poor survival outcomes. Incident lung cancer patients from the NLST were split into training and test cohorts and an external cohort of non-screen detected adenocarcinomas was used for further validation. After removing redundant and non-reproducible radiomics features, backward elimination analyses identified a single model which was subjected to Classification and Regression Tree to stratify patients into three risk-groups based on two radiomics features (NGTDM Busyness and Statistical Root Mean Square [RMS]). The final model was validated in the test cohort and the cohort of non-screen detected adenocarcinomas. Using a radio-genomics dataset, Statistical RMS was significantly associated with FOXF2 gene by both correlation and two-group analyses. Our rigorous approach generated a novel radiomics model that identified a vulnerable high-risk group of early stage patients associated with poor outcomes. These patients may require aggressive follow-up and/or adjuvant therapy to mitigate their poor outcomes.

Cancer is the leading cause of death in Puerto Rico (PR). Hurricane Maria (HM) and its aftermath lead to widespread devastation on the island, including the collapse of the healthcare system. Medically fragile populations, such as cancer survivors, were significantly affected. The goal of this study was to assess the impact of HM on barriers to care, emotional distress, and inflammatory biomarkers among cancer survivors in PR. This exploratory longitudinal study was conducted in health care facilities and community support groups from PR. Cancer survivors (n = 50) and non-cancer participants (n = 50) completed psychosocial questionnaires and provided blood samples that were used to assess inflammatory cytokines levels. Among this cohort, we identified 41 matched cancer survivors/non-cancer participants pairs. Data were analyzed through descriptive, frequencies, correlational, and regression analyses. Cancer survivors that were affected by HM reported increased barriers in accessing medical care, which were directly associated with anxiety, perceived stress, and post-traumatic symptomatology. Moreover, being a cancer survivor, predicted more barriers to receiving health care, especially in the first six weeks after the event, after which the effect was attenuated. Several inflammatory cytokines, such as CD31, BDNF, TFF3, Serpin E-1, VCAM-1, Vitamin D BP, and PDGF-AA, were significantly upregulated in cancer survivors while MMP9 and Osteopontin both had significant positive correlations with barriers to care. HM significantly impacted Puerto Ricans psychosocial well-being. Cancer survivors had significant barriers to care and showed increased serum inflammatory cytokines but did not show differences in anxiety, stress, and post-traumatic symptoms compared to non-cancer participants.

Background: Over the last several decades there have been concerted efforts to reduce health disparities among intersecting underserved populations including racial and ethnic, social class, disability, and geographic location. However, there have been limited efforts to identify and address cancer disparities by sexual orientation and gender identity (SOGI). We sought to identify differences in demographics, risk factors, clinical characteristics, and outcomes between sexual and gender minority (SGM) and non-SGM cancer patients. Methods: Analysis include over 70,000 cancer patients at the Moffitt Cancer Center (Tampa, FL) who completed our Electronic Patient Questionnaire (EPQ) at their initial appointment. Moffitt’s EPQ is a standard-of-care (SOC) questionnaire instrument used to collect self-reported demographics, prior medical history, history of cancer, utilization of cancer screening services, and lifestyle risk factors. The SOGI items in the demographics section were used to categorize persons as a SGM or non-SGM. Cancer registry data were used to obtain clinical covariates on the patients that completed the EPQ. Results: SGM patients represented 2.3% of the patient population. The mean age of SGM patients was significantly lower (mean=56) compared to non-SGM patients (mean=62.3, p= Discussion: The Moffitt Cancer Center began collecting SOGI information as SOC information on all new patients since 2016. In just 5 years of data collection, our institution has identified numerous real-world disparities among SGMs patients. These findings will be critical to develop tailored interventions to improve equality care across the cancer care continuum for SGM populations. Citation Format: Shahrzad A. Zamani, Jaileene Perez-Morales, Jarred D. Miller, Matthew B. Schabath. Cancer disparities among sexual and gender minority patients at a National Cancer Institute-Designated Comprehensive Cancer Center [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1916.

Prostate cancer (PCa) ranks as the second leading cause of cancer-related deaths in American men. Most PCa deaths occur due to aggressive, fast-growing tumors that develop into metastasis when cancer cells spread to other body parts, including the lymph nodes, bones, etc. However, men could also be affected by indolent, slow-growing tumors, which may exist for an extended period without causing any symptoms or death. Current prognostic methods, such as Prolaris, or Oncotype, do not precisely specify how aggressive cancer might be, resulting in overtreating patients with indolent PCa. Therefore, improved ways to differentiate indolent from aggressive tumors are needed. In the presented study, we have validated the clinical potential of phospho-Rb S249, N-cadherin, B-catenin, and E-cadherin to discriminate between indolent and aggressive PCa tumors. We hypothesize that the biomarkers phospho-Rb S249, N-cadherin, E-cadherin, and B-catenin expression in PCa will be a clinical tool for the risk classification of PCa tumors when used as part of a prostate proteomic (PPS) that can predict the risk of the tumor of evolving into an aggressive phenotype. We conducted immunohistochemistry staining of these biomarkers on PCa tumor microarrays and correlated it with the patient’s clinical parameters, such as tumor size, lymph node invasion, metastasis, etc. Also, we correlated it with the PSA levels and created a PPS. The combination of our proteomic biomarkers predicts patient staging by 53.83%. Adding clinical data such as the tumor size, lymph node invasion, and metastasis (TNM) increases the precision of the prediction to 83.21%. In addition, the PSA levels increased as the phospho-Rb S249 expression and N-cadherin rise, accompanied by up-regulation of E-cadherin. Also, the PPS showed to distinguish between normal prostate tissue, hyperplasia, and PCa. Results suggest that these biomarkers could serve as a tool for the risk classification of indolent and aggressive PCa tumors. Citation Format: Sheila M. Valle Cortes, Pedro Santiago Cardona, Carlos Diaz Osterman, Jaileene Perez Morales, Darielys Maldonado Maldonado, Gilberto Ruiz Deya. The expression of phospho-Rb S249, N-cadherin, E-cadherin, and B-catenin as a prostate proteomic score for the risk classification of prostate cancer tumors. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5313.

Background: Our group developed and piloted the Curriculum for Oncologists on LGBT populations to Optimize Relevance and Skills (COLORS), an interactive web-based LGBT cultural sensitivity training for oncologists. Building on our formative pilot work, in February 2021 we launched a randomized trial to compare the effectiveness of the oncology-specific COLORS training to a general LGBT healthcare cultural sensitivity training. We report here on pre- and post-assessments of this trial. Methods: From the AMA Masterfile we obtained names and mailing addresses for 5,000 practicing oncologists that were randomly selected for equal distributions across the 9 U.S. Census Divisions. From February to May 2021, we mailed FedEx envelopes to these oncologists that included a invitation letter and information to register for the trial. Using REDCap, oncologists registered, completed a pre-training assessment, were randomized and emailed instructions on how to complete their assigned training (COLORS training or a publicly available web-based LGBT comparison training), and then completed a post-training assessment. The pre- and post-training assessments include measures for knowledge, attitudes (Modern Homonegativity Scale [MHS] and Modified Attitudes Toward LGBT Patients Scale [ATLPS-M], and practice behaviors (Gay Affirmative Practice [GAP] Scale) Results: As of October 2021, 224 oncologists were enrolled, 201 completed the pre-training and were randomized, and 138 completed the post-training assessment. Among those randomized to COLORS, there were significant decreases from pre- to post-training (median: 24 [IQR 19-30] vs. 22.5 [16-28.5]; p Conclusion: Results from two prior single-arm trials and now this RCT demonstrate that the COLORS training is effective in improving knowledge, attitudes, and affirming clinical practices related to the care of LGBT patients. This work is generously funded by the Bristol Myers Squibb (BMS) Foundation and BMS Company. Citation Format: Matthew B. Schabath, Jaileene Perez-Morales, Eryk N. Hernandez, Jarred D. Miller, Gwendolyn P. Quinn, Julia Seay. A randomized trial to examine the effectiveness of an LGBT cultural sensitivity training for oncologists: The COLORS trial [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3635.

Background: There are two first-line treatment recommendations for advanced ovarian cancer: i) upfront cytoreductive debulking followed by chemotherapy and ii) neoadjuvant chemotherapy prior to surgical debulking. The choice between these two treatment strategies is controversial as there are no standardized guidelines for clinical decision support. As such, there remains a critical unmet need to identity biomarkers to personalize the most effective treatment strategies. The primary objective of this study is to identify and validate radiomic biomarkers that predict treatment response among patients with high-grade serous ovarian cancer treated with upfront surgical debulking. Methods: Intratumoral radiomic features (n=308) were extracted from pre-treatment contrast-enhanced CT images; analyses were conducted to remove correlated, non-stable, and non-reproducible features. Patients treated with upfront surgery (N=182) were split into training (N=91) and test (N=91) cohorts and a cohort of patients treated with upfront neoadjuvant (N=116) was used to determine if the radiomic features were prognostic or predictive. Overall (OS) and progression-free survival (PFS) were the main endpoints. Classification and Regression Tree analysis was used to identity the most informative radiomic features in the training cohort, which were then analyzed in the test cohort and the upfront neoadjuvant cohort. Results: Decision tree analysis identified a volumetric feature, ROI volume center of mass (CoM) in X direction, as the most informative radiomic feature which stratified upfront surgery patients into high- and low-risk. In the training cohort, high-risk patients were associated with significantly worse OS versus low-risk patients (HR=2.01; 95% CI 1.07-3.77 vs. 1.00 and 5-year OS=39.9% vs. 56.1%, respectively; log-rank p-value=0.03). In the test cohort, the high- vs. low-risk patients were also associated with poor OS (HR=2.23; 95% CI 1.19-4.17 vs. 1.00 and 5-year OS=23.3% vs. 51.3%, respectively; log-rank p-value=0.01). This radiomic feature was not associated with OS among patients with upfront neoadjuvant (HR=1.23; 95% CI 0.73-2.07 vs. 1.00 and 5-year OS=23.9% vs. 36.6%, respectively; log-rank p-value=0.44). Similar findings were observed for PFS. Conclusion: Utilizing standard-of-care imaging, we identified and validated a predictive radiomic feature associated with outcomes among patients with high-grade serous ovarian cancer treated with upfront surgical debulking but not among patients treated with neoadjuvant chemotherapy. This radiomic biomarker, which describes the location of center of mass inside the tumor in pixels in the x-direction, could be potentially utilized as clinical decision support to guide first-line treatment options. This study was generously funded by a Miles for Moffitt pilot grant. Citation Format: Jaileene Perez-Morales, Christelle M. Colin Leitzinger, Sweta K. Sinha, Melissa J. McGettigan, Daniel K. Jeong, Olya Stringfield, Mahmoud Abdala, Natarajan Raghunand, Robert J. Gillies, Jing-Yi Chern, Lauren C. Peres, Matthew B. Schabath. Radiomic biomarkers to optimize treatment decision and predict patient outcomes in serous ovarian carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3218.

Prostate cancer (PCa) is the second most frequent malignancy in men worldwide. This disease is the cause of most cancer death among Puerto Rican men, representing 18.9% of overall cancer deaths. However, a patient outcome depends on the type of tumor, which can be indolent or aggressive. The indolent PCa is of slow tumor growth and favorable prognosis, whereas the aggressive form is of rapid tumor spread and high mortality. The discrimination between the indolent and aggressive forms of PCa has resulted in a significant problem of overtreating those patients with the indolent form of the disease. Thus, biomarkers that aid in the decision-making process regarding active surveillance in very low-, low-, and favorable intermediate-risk PCa face a large unmet need. For this purpose, we evaluate the expression of phospho-Rb S249 and epithelial to mesenchymal transition markers (E-cadherin, N-cadherin, B-catenin) to create a combination that can help distinguish between the indolent and aggressive forms of PCa. We hypothesize that the biomarkers E-cadherin, and B-catenin could be useful to identify indolence among PCa patients, whereas phospho-Rb S249 and N-cadherin could identify the aggressive form of the disease. As part of our methodology, we have conducted immunohistochemistry (N=182) against the expression of these biomarkers on PCa tumor microarrays (TMA’s). Correlation coefficient analyses were conducted to assess if the expression of these biomarkers could correlate with any of the patient’s clinical parameters (stage, grade, tumor size, lymph nodes metastasis, metastasis to distant sites, Gleason score, and Gleason grade). In addition, we created a linear regression analysis to determine if the combined proteins could precisely predict the staging, grade, or Gleason score of the patients. Results have shown that the combination of phospho-Rb S249, N-cadherin, E-cadherin, and B-catenin had a precision of 68% correctly predicting the staging of the patients, and adding the tumor size, lymph nodes metastasis, and metastasis to distant sites to the equation increased the prediction to a 96%. Additionally, we identified that E-cadherin and B-catenin negatively correlated with the tumor size, grade, stage, Gleason grade, and Gleason score of the patients. Whereas N-cadherin and phospho-Rb S249 positively correlate with the tumor size, stage, and Gleason grade. These results suggest that the expression of phospho-Rb S249, N-cadherin, E-cadherin, and B-catenin could be used as a potential tool to discriminate between the indolent and aggressive forms of PCa. This will contribute to enhancing the clinical decision of which patients should be treated or undergo active surveillance. Citation Format: Sheila M. Valle Cortes, Pedro Santiago Cardona, Carlos Diaz Osterman, Harold Saavedra, Shannalee R. Martinez, Gilberto Ruiz Deya, Jaileene Perez Morales, Adam B. Murphy. Phosphorylation of the retinoblastoma protein at Serine 249 and the cadherin switch as clinically informative biomarkers for the discrimination between indolent and aggressive forms of prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 6001.

11000 Background: Published studies report oncologists have limited knowledge about LGBT health and cancer needs but also report high interest in receiving education regarding the unique health needs of LGBT patients with cancer. As such, our group developed the Curriculum for Oncologists on LGBT populations to Optimize Relevance and Skills (COLORS), an interactive web-based LGBT cultural sensitivity training for oncologists that focused on focused on improving knowledge and communication skill-building. We report here on the results from two single-arm feasibility/acceptability trials and one randomized pragmatic trial (RPT) demonstrating effectiveness of the COLORS training. Methods: The first single-arm trial included 44 oncologists from three cancer centers in Florida and the second single-arm trial included 50 oncology fellows recruited through ASCO. The RPT included 225 oncologists randomly selected for equal distribution across the U.S., and participants were randomly assigned to either the COLORS training or a publicly available, web-based LGBT comparison training. Each of these studies included pre- and post-training assessments of LGBT-related knowledge, attitudes, and affirming practices. The RCT included a 3-month follow-up assessment to gauge retention of LGBT-affirming knowledge, attitudes, and practices over time. Results: Both single arm trials demonstrated that the COLORS training was feasible, acceptable, and results from the pre- and post-training assessments suggested the training improved knowledge, attitudes, and affirming practice behaviors. In the RPT, those randomized to COLORS demonstrated significant improvements in LGBT-related attitudes pre- to post-training (p < 0.01), and no significant changes in attitudes were observed among those randomized to the comparison training (p = 0.98). Both trainings yielded significant (p < 0.01) increases from pre- to post-training in LGBT-affirming practice behaviors. Likewise, both trainings yielded significant (p < 0.01) improvements in knowledge from pre- to post-training. Similar significant findings were observed when we analyzed the 3-month follow-up assessment. Conclusions: Results from the prior single-arm trials and the current RCT demonstrate that the COLORS training is effective in improving oncologist knowledge, attitudes, and affirming clinical practices related to the care of LGBT patients with cancer. Part of this work was funded by the Bristol Myers Squibb (BMS) Foundation and BMS Company.

The cancer phenotype is usually characterized by de-regulated activity of a variety of cellular kinases, with consequent abnormal hyper-phosphorylation of their target proteins. Therefore, antibodies that allow the detection of phosphorylated versions of proteins have become important tools both pre-clinically in molecular cancer research, and at the clinical level by serving as tools in pathological analyses of tumors. In order to ensure reliable results, validation of the phospho-specificity of these antibodies is extremely important, since this ensures that they are indeed able to discriminate between the phosphorylated and un-phosphorylated versions of the protein of interest, specifically recognizing the phosphorylated variant. A recommended validation approach consists in de-phosphorylating the target protein and assessing if such de-phosphorylation abrogates antigen immunoreactivity when using the phospho-specific antibody. In this chapter, we describe a protocol to validate the specificity of a phospho-specific antibody that recognizes a phosphorylated variant of the Retinoblastoma (Rb) protein in lung cancer cell lines. The protocol consists in the de-phosphorylation of the Rb-containing protein lysates by treating them with bovine intestinal phosphatase, followed by assessment of the de-phosphorylation by immunoblot.

Simple Summary Biobehavioral disorders can negatively impact patients with ovarian cancer. Growing evidence suggests that chronic stress can promote tumor progression, the release of inflammatory mediators, and macrophage infiltration into the tumor. However, the role of stress hormones in regulating cancer cell/macrophage crosstalk remains unclear. This study aimed to assess the role of stress hormone-stimulated macrophages in modulating inflammatory networks and ovarian cancer biology. Our data show that stress hormones induced secretion of inflammatory proteins in ovarian cancer cell/macrophage co-cultures. Furthermore, we show that restraint stress leads to cancer growth, macrophage infiltration, and PDGF-AA protein expression in animal models of ovarian cancer. Conversely, zoledronic acid was able to prevent the effects of restraint stress on ovarian cancer growth. Overall, our data suggest a role for stress hormone-stimulated macrophages in ovarian cancer progression and suggest the involvement of PDGF-AA as a key mediator of this process. Abstract Multiple studies suggest that chronic stress accelerates the growth of existing tumors by activating the sympathetic nervous system. Data suggest that sustained adrenergic signaling can induce tumor growth, secretion of pro-inflammatory cytokines, and macrophage infiltration. Our goal was to study the role of adrenergic-stimulated macrophages in ovarian cancer biology. Cytokine arrays were used to assess the effect of adrenergic stimulation in pro-tumoral cytokine networks. An orthotopic model of ovarian cancer was used to assess the in vivo effect of daily restraint stress on tumor growth and adrenergic-induced macrophages. Cytokine analyses showed that adrenergic stimulation modulated pro-inflammatory cytokine secretion in a SKOV3ip1 ovarian cancer cell/U937 macrophage co-culture system. Among these, platelet-derived growth factor AA (PDGF-AA), epithelial cell-derived neutrophil-activating peptide (ENA-78), Angiogenin, vascular endothelial growth factor (VEGF), granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin-5 (IL-5), Lipocalin-2, macrophage migration inhibitory factor (MIF), and transferrin receptor (TfR) were upregulated. Enriched biological processes included cytokine-mediated signaling pathways and positive regulation of cell proliferation. In addition, daily restraint stress increased ovarian cancer growth, infiltration of CD68+ macrophages, and expression of PDGF-AA in orthotopic models of ovarian cancer (SKOV3ip1 and HeyT30), while zoledronic acid, a macrophage-depleting agent, abrogated this effect. Furthermore, in ovarian cancer patients, high PDGFA expression correlated with worse outcomes. Here, it is shown that the adrenergic regulation of macrophages and PDGFA might play a role in ovarian cancer progression.

In 2017, approximately 5 million Americans were living with Alzheimer’s disease (AD), and it is estimated that by 2050 this number could increase to 16 million. In this study, we apply mathematical optimization to approach microarray analysis to detect differentially expressed genes and determine the most correlated structure among their expression changes. The analysis of {"type":"entrez-geo","attrs":{"text":"GSE4757","term_id":"4757"}}GSE4757 microarray dataset, which compares expression between AD neurons without neurofibrillary tangles (controls) and with neurofibrillary tangles (cases), was casted as a multiple criteria optimization (MCO) problem. Through the analysis it was possible to determine a series of Pareto efficient frontiers to find the most differentially expressed genes, which are here proposed as potential AD biomarkers. The Traveling Sales Problem (TSP) model was used to find the cyclical path of maximal correlation between the expression changes among the genes deemed important from the previous stage. This leads to a structure capable of guiding biological exploration with enhanced precision and repeatability. Ten genes were selected (FTL, GFAP, HNRNPA3, COX1, ND2, ND3, ND4, NUCKS1, RPL41, and RPS10) and their most correlated cyclic structure was found in our analyses. The biological functions of their products were found to be linked to inflammation and neurodegenerative diseases and some of them had not been reported for AD before. The TSP path connects genes coding for mitochondrial electron transfer proteins. Some of these proteins are closely related to other electron transport proteins already reported as important for AD.

The breast cancer mortality rate in Puerto Rican women is higher when compared to Hispanic women in the United States, despite having similar incidence. While disparities are well documented between Non-Hispanic Blacks and Non-Hispanic Whites, Hispanic populations have not been explored as thoroughly. Previous studies have identified high Body Mass Index (BMI) as a risk factor in postmenopausal breast cancer. Breast cancer incidence tends to increase with age, but younger age of diagnosis may be related to more aggressive disease. Studies have demonstrated that mammogram and continuous breast cancer screening can reduce mortality through early detection. Our study focuses on characterizing BMI, age at diagnosis, and screening information in Puerto Rican women obtained from a patient survey administered by the Puerto Rico BioBank (PRBB). These factors may contribute to the observed disparity in breast cancer mortality in these patients. Data abstracted from the PRBB self-reported questionnaire identified 260 females who had been diagnosed with breast cancer between the ages of 25 and 86. The questionnaire includes demographic, family history, lifestyle, and cancer-related questions. BMI was calculated from the self-reported height and weight values, then classified according to the Center for Disease Control (CDC) categories: underweight ( Citation Format: Abigail E. Lantz, William D. Cress, Allison Bahr, Edna Gordián, Jaileene Perez-Morales, Idhaliz Flores, María Rojas. Body mass index, age at diagnosis, and mammography screening behaviors in women diagnosed with breast cancer in Puerto Rico [abstract]. In: Proceedings of the AACR Virtual Conference: 14th AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2021 Oct 6-8. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2022;31(1 Suppl):Abstract nr PO-218.

Cancer is the leading cause of death in Puerto Rico (PR). Hurricane Maria (HM) and its aftermath lead to widespread devastation in the island, including the collapse of the healthcare system. Medically fragile populations, such as cancer survivors, were significantly affected. The goal of this study was to assess the impact of HM on barriers to care, emotional distress, and inflammatory biomarkers among cancer survivors in PR. This exploratory longitudinal study was conducted in health care facilities and community support groups from PR. Cancer survivors (n=50) and non-cancer participants (n=50) completed psychosocial questionnaires and provided blood samples that were used to assess inflammatory cytokines levels. Data were analyzed through descriptive, frequencies, correlational, and linear regression analyses. Cancer survivors that were affected by HM reported increased barriers in accessing medical care, which were directly associated with anxiety, perceived stress, and post-traumatic symptomatology. Moreover, being a cancer survivor, along with closeness in time from HM predicted more barriers to receiving health care. Several inflammatory cytokines, such as CD31, BDNF, TFF3, Serpin E-1, Vitamin D BP, VCAM-1, Osteopontin, Chitinase 3 like 1, MMP-9 and MIF were significantly upregulated in cancer survivors while BDNF, MMP9 and Osteopontin had significant positive correlations with barriers to care. HM significantly impacted Puerto Ricans psychosocial well-being. Cancer survivors had significant barriers to care and showed increased serum inflammatory cytokines, but didn’t show differences in anxiety, stress and post-traumatic symptoms compared to non-cancer participants.

Background: Most lung cancer screening trials to date and current screening guidelines overlook vulnerable populations with disproportionate lung cancer burden such as racial and ethnic minorities. Given the barriers, perceptions, and socioeconomic burden of these underserved populations, uptake of lung cancer screening has minimal. To develop and implement better methods of recruitment underserved populations, concerted efforts to understand the barriers and perceptions of these population are needed. Objective: Conduct a systematic review of methods for recruitment of racial and ethnic minorities into lung cancer screening trials or programs. Methods: We searched for studies that reported methods of recruitment and racial and ethnic minorities into lung cancer screening from 2010 to June 2020, using Ovid MEDLINE and Embase databases. Full-text screening and data extraction were completed independently by two reviewers and adjudication was performed by a third, independent reviewer. We compared referral rates of the recruitment modalities by racial/ethnic subgroups and by sex. Results: We identified 525 studies of which 25 were included in the systematic review corresponding to 291,329 patients. The most frequent methods of recruitment were referral, physical mail, community-based strategies, targeted strategies, phone, advertisement, and social media. Referral (n=7; 2,510 patients) was the most effective method of recruitment with response rates ranging from 31% to 97%. Direct mail (n=8 studies; 189,799 patients) was the second most extensively used recruitment method with response rates of 28% to 79%. Direct mail to Hispanic patients (2,995 total) for lung cancer screening programs was more effective than other methods of recruitment. Direct mail was more effective in recruiting African American patients (8,331 total) compared to referral (1,083 total). Conclusion: This review identified recruitment modalities that were more successful and less unsuccessful in recruiting racial and ethnic minorities into lung cancer screening trials and programs. This work highlights priorities and next steps required to improve recruitment of racial and ethnic minorities in lung cancer screening. There is a need to develop culturally relevant lung cancer screening interventions for Hispanics/Latino patients and to create targeted intervention for African American patients. Citation Format: Tiffany J. Gilliard, Jaileene Perez-Morales, Matthew Schabath, Mary Katherine Haver. Methods to improve representation of racial and ethnic minorities in lung cancer screening programs: A systematic review [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr LB219.

BACKGROUND: The gold standard treatment for urinary bladder cancer is cystectomy, which can result in postoperative complications, patient morbidities, and lengthy hospital visits post-surgery. At present, there are limited models that can predict length of stay for patients post cystectomy. Identifying risk factors associated with increased length of stay could have translational implications in mitigating such risk factors. As such, the goal of this study was to generate parsimonious models to predict length of hospital stay (LOHS) following cystectomy in bladder cancer patients. METHODS: Data from the American College of Surgeons (ACS) National Surgical Quality Improvement Program (NSQIP) was used to analyze patients who underwent cystectomy between 2014 and 2016. Modeling building approaches were used to identify the most informative preoperative and postoperative covariates to predict LOHS. Separate models were identified for preoperative covariates and for postoperative covariates and the most informative pre- and postoperative covariates were combined into a single model. RESULTS: This analysis includes 6,074 patients (mean LOHS = 9.4 days). The preoperative (5 covariates; R squared= 0.245 and Adj R-squared = 0.240) and postoperative models (13 covariates; R squared= 0.237 and Adj R-squared = 0.236) were significantly better predictors compared to the ACS risk calculator model (19 covariates; R-square= 0.026 and Adj R-squared = 0.025). Likewise, the combined model (15 covariates) was a significantly better predictor (R squared= 0.241 & Adj R-squared = 0.239) when compared to the ACS risk calculator model. The combined model was subjected into CART analysis to stratify patients into five risk-groups based on ventilator over 48 hours, organ/space surgical site infection, wound disruption, and pneumonia. Patients in the high-risk group had significantly longer mean LOHS of 26.29 days compared to low-risk group of 8.4 days. CONCLUSIONS: This study identified and validated a clinically important combined model that is to the ACS NSQIP surgical risk calculator to predict LOHS in patients who underwent cystectomy. Patients at high-risk may benefit from allocation of additional follow-up to prevent longer LOHS. Citation Format: Rashmi Pathak, Jaileene Perez-Morales, Sephalie Y. Patel, Michael A. Poch, Matthew B. Schabath. Parsimonious model to predict length of stay following cystectomy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 883.

Background: In 2014, Moffitt Cancer Center initiated a lung cancer screening program for high-risk individuals who meet USPSTF inclusion criteria. As of January 2020, 576 patients have completed one or more lung cancer screenings. However, approximately 40% of lung cancer screening patients were noncompliant with their recommended follow-up. To understand reasons for lack of follow-up screening and to determine patient satisfaction and experiences, we conducted a survey with open-ended items to assess patient experiences and satisfaction among lung cancer screening patients. Methods: In this analysis, we report on findings from the qualitative portion of the survey. Six open-ended items elicited barriers related to returning for screening, experience with other cancer screenings, positive and negative experiences with the LDCT visit, as well as suggestions for improving the process of LDCT screening visits. Content analysis using the constant comparison method was applied to the text and coded based on the a priori codes of the open-ended questions. Results: As of November 2020, 190 people responded to the quantitative survey and 77 provided responses to one or more open-ended questions. Thirty-five respondents provided general positive comments about their lung cancer screening experience with typical responses of “positive” and “good experience”. Twenty respondents mentioned that the staff was “caring”, “knowledgeable”, and “great staff”. Ten respondents mentioned that they were “always treated well”, “treated with respect”, “high quality care”.. Negative comments were provided by 32 respondents in the areas of: “wanting more information on results” or “waiting too long for results” (N=7), “inefficient scheduling issues” (N=6), and billing issues (N=5). Additionally, six patients reported COVID-related issues impacted their lung cancer screening experience. Suggestion for improvements included: on-line appointments or text or email reminders (N = 10), lower costs (N=6), and uncertainty about remaining qualified for lung cancer screening due to age or current/prior medical conditions (N=5). Conclusion: Overall, findings provide insights about patient experiences and satisfaction for lung cancer screening. Suggestions for improvement centered on better scheduling systems and appointment reminders as well as the ability to speak to a clinician to better understand results and future eligibility. Ongoing patient-centered feedback may improve the lung cancer screening experience and increase follow-up screening rates. Citation Format: Jaileene Perez-Morales, Rashmi Pathak, Monica Reyes, Ada Sulbaran, Tiffany Gilliard, Haley Tolbert, Rajwantee Tirbene, Jhanelle Gray, Vani N. Simmons, Matthew B. Schabath, Gwendolyn P. Quinn. Qualitative findings from a survey on patient experiences and satisfaction with lung cancer screening [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 889.

Background: In 2014, Moffitt Cancer Center (MCC) in Tampa, Florida launched a lung cancer screening program for high-risk individuals meeting USPSTF inclusion criteria. To reveal successes and potential limitations of this program, we conducted a survey to measure lung cancer screening patient experiences and satisfaction. Methods: In August 2020, a survey was mailed to 576 patients who completed one or more lung cancer screenings at MCC. The survey collected demographics and smoking history and measured patient satisfaction and experiences in 6 domains: the appointment process, clinical staff interactions, communication, visit with the provider, screening results and cost, and the clinic facility/overall satisfaction. There were also questions on the impact of the COVID-19 pandemic. A 5-point Likert scale was used to measure satisfaction with clinical staff and clinic facility/overall satisfaction (1= excellent to 5= poor) and other domains (1=strongly agree to 5=strongly disagree). Results were quantified using descriptive statistics. Results: Among the 160 patients that completed the survey, 96.3% were White, 93.1% were non-Hispanic, 46.9% were female, 78.1% were former smokers, and the mean age was 69 years. For the appointment process, 91.2% “strongly agreed/agreed” it was easy to make an appointment and 92.5% “strongly agreed/agreed” the clinic check-in process was efficient. Next, nearly 93% reported the courtesy of the clinical staff was “excellent/very good”. For communication, while 90% “strongly agreed/agreed” that the lung cancer screening process was clearly explained, 63.2% “strongly agreed/agreed” that the potential harms and limitations were clearly explained and 71.2% “strongly agreed/agreed” that the process for follow-up screening was clearly explained. For the provider questions, 71.3% “strongly agreed/agreed” that the provider was willing to listen carefully and 66.3% “strongly agreed/agreed” that the instructions regarding follow-up were easy to understand. For results and costs, 79.4% “strongly agreed/agreed” the screening results were clearly explained and 70% “strongly agreed/agreed” that the cost of the screening was justified. Regarding overall satisfaction, 87.6% “strongly agreed/agreed” they would recommend lung cancer screening at MCC. Finally, 10.6% of patients were concerned about getting a lung cancer screening during the COVID-19 pandemic and only 8.1% reported that the COVID-19 pandemic prevented them from getting a lung cancer screening. Conclusion: Generally, for all 6 domains, patients reported favorable satisfaction and experience with lung cancer screening at MCC. However, higher rates of “neither agree/disagree” and “prefer not to answer” were observed for items related to communication and visit with the provider items suggesting improvements may be warranted, particularly related to future visits. Citation Format: Monica E. Reyes, Jaileene Perez-Morales, Rashmi Pathak, Ada Sulbaran, Tiffany J. Gillard, Haley Tolbert, Rajwantee Tirbene, Jhanelle Gray, Vani N. Simmons, Gwendolyn P. Quinn, Matthew B. Schabath. Survey results of lung cancer screening patient experiences and satisfaction [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 890.

INTRODUCTION: The lesbian, gay, bisexual, transgender, and queer/questioning (LGBTQ) community is a medically underserved population that suffers from cancer disparities. To identify potential gaps in attitudes, knowledge, and institutional practices toward LGBTQ patients, we conducted a mixed-methods survey of healthcare providers in the ECOG-ACRIN Cancer Research Group. METHODS: A validated, web-based survey was administered to members of the ECOG-ACRIN Cancer Research Group in late 2019. The survey was completed by 490 healthcare providers measuring attitudes and knowledge about LGBTQ health and institutional practices regarding collection of sexual orientation and gender identity data. Results were quantified using descriptive and stratified analyses. RESULTS: Among the 490 healthcare providers that completed the survey, 77% were White, 86% were non- Hispanic, 75% were female, 81% were heterosexual, and the mean age was 46 years. Approximately 47% of the respondents were medical oncologists, 39% practiced in academic medical centers (AMCs), and 49% were from the Midwest/Northeast. As reported by prior institutional and national survey studies, there was high interest (77%) in receiving education regarding the unique health needs of LGBTQ patients, an overall limited knowledge about LGBTQ health and cancer needs, and a significant decrease from survey assessment to postsurvey assessment for confidence in knowledge for LGBT health needs. There was high agreement (71%) regarding the importance of knowing gender identity, which was contrasted with a low agreement (48%) regarding the importance of knowing sexual orientation. Stratified analyses revealed significant differences of some attitude and knowledge items. For example, when stratified by licensure/terminal degree, MD/DO vs. RN/NP/PA were significantly more willing to be listed as a LGBTQ-friendly provider (81% vs. 50%) and reported to be knowledgeable about LGB health needs (69% vs. 53%). Providers at AMCs were significantly more willing to be listed as a LGBTQ-friendly provider (71% vs.53%) believe there should be mandatory education (72% vs. 60%) regarding care of LGBQ patients versus those at non-AMCs. Having family and/or friends who are LGBTQ (vs. none) and political affiliation (conservative vs. liberal) were strong effect modifiers resulting in significant differences for 9 and 8 items, respectively. CONCLUSION: To our knowledge, this was the first study of cancer care providers in a large cancer clinical trials research group that assessed attitudes, knowledge, and institutional practices of LGBTQ patients with cancer. Consistent with prior studies, there was limited knowledge about LGBTQ health and cancer needs, but high interest in receiving education regarding this community. New and intriguing differences for many attitude and knowledge items were revealed when we stratified the responses by licensure/terminal degree, practice setting, LGBTQ friends and/or family members, and political affiliation. Citation Format: Matthew B. Schabath, Jaileene Perez-Morales, Megan Sutter, Lynne Wagner, Melissa Simon, Ruth C. Carlos, Bruce J. Giantonio, Gwendolyn P. Quinn, Edith P. Mitchell. Survey of healthcare providers in the ECOG-ACRIN cancer research group: Attitudes, knowledge, and practice behaviors about LGBTQ patients with cancer [abstract]. In: Proceedings of the AACR Virtual Conference: Thirteenth AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2020 Oct 2-4. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2020;29(12 Suppl):Abstract nr PO-068.

Background: Cancer is the leading cause of death in Puerto Rico (PR). Hurricane Maria (HM) and its aftermath lead to widespread devastation in the island, including the collapse of the healthcare system. Medically fragile populations, such as cancer patients and survivors, were significantly affected. The goal of the current study was to assess the impact of HM on barriers to care, emotional distress and inflammatory biomarkers among cancer patients in PR. Methods: This exploratory longitudinal study was conducted in health care facilities and community support groups from PR. Cancer patients (n=50) and non-cancer patients (n=50) completed a battery of psychosocial questionnaires and provided blood samples that were utilized to assess inflammatory cytokines levels. Data were analyzed through descriptive, frequencies, correlational and linear regression analyses. Results: Cancer patients that were affected by HM reported increased barriers in accessing medical care, which were positively associated with anxiety, perceived stress, and post-traumatic symptomatology. Moreover, being a cancer patient or survivor, along with closeness in time from HM predicted more barriers to receiving health care. Several inflammatory cytokines were significantly upregulated in cancer patients and positively correlated with barriers to care. Conclusions: HM significantly impacted Puerto Ricans psychosocial well-being. Cancer patients had significant barriers to care and increased serum inflammatory cytokines, but similar anxiety, stress and post-traumatic symptoms compared to non-cancer controls. Impact: These findings demonstrate the urgency of delineating a plan for providing cancer care to patients in the aftermath of a natural disaster while promoting and influencing resilience and well-being. Citation Format: Mary Rodriguez-Rabassa, Ruthmarie Hernandez, Zindie Rodriguez, Claudia B Colon-Echevarria, Lizette Maldonado, Nelmit Tollinchi, Estefania Torres, Adnil Mulero, Daniela Albors, Jaileene Perez-Morales, Idhaliz Flores, Heather Jim, Eida M Castro, Guillermo N Armaiz-Pena. Impact of a natural disaster on access to care and biopsychosocial outcomes among Hispanic/Latino cancer patients [abstract]. In: Proceedings of the Twelfth AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2019 Sep 20-23; San Francisco, CA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2020;29(6 Suppl_2):Abstract nr A042.

Prediction of lung cancer metastasis relies on post-resection assessment of tumor histology, which is a severe limitation since only a minority of lung cancer patients are diagnosed with resectable disease. Therefore, characterization of metastasis-predicting biomarkers in pre-resection small biopsy specimens is urgently needed. Here we report a biomarker consisting of the phosphorylation of the retinoblastoma protein (Rb) on serine 249 combined with elevated p39 expression. This biomarker correlates with epithelial-to-mesenchymal transition traits in non-small cell lung carcinoma (NSCLC) cells. Immunohistochemistry staining of NSCLC tumor microarrays showed that strong phospho-Rb S249 staining positively correlated with tumor grade specifically in the squamous cell carcinoma (SCC) subtype. Strong immunoreactivity for p39 positively correlated with tumor stage, lymph node invasion, and distant metastases, also in SCC. Linear regression analyses showed that the combined scoring for phospho-Rb S249, p39 and E-cadherin in SCC is even more accurate at predicting tumor staging, relative to each score individually. We propose that combined immunohistochemistry staining of NSCLC samples for Rb phosphorylation on S249, p39, and E-cadherin protein expression could aid in the assessment of tumor staging and metastatic potential when tested in small primary tumor biopsies. The intense staining for phospho-Rb S249 that we observed in high grade SCC could also aid in the precise sub-classification of poorly differentiated SCCs.

The retinoblastoma protein (pRb) is an important tumor suppressor and cell cycle repressor. pRb is a phosho-protein whose function is regulated primarily at the level of phosphorylation, and therefore, detecting pRb´s phosphorylation status in human tissue samples can be clinically informative. Unfortunately, detection of phosphorylated pRb residues can be technically challenging, as these residues can often be weak antigens. In this chapter, we describe an enhanced sensitivity immunohistochemistry protocol for the staining of phosphorylated serine 249 in pRb, in human lung tumor samples.

The retinoblastoma tumor suppressor protein (pRb) is a preeminent tumor suppressor that acts as a cell cycle repressor, specifically as an inhibitor of the G1-S transition of the cell cycle . pRb is a phosphoprotein whose function is repressed by extensive phosphorylation in several key residues, and therefore, pRb's phosphorylation status has become a surrogate for pRb activity. In particular, hyperphosphorylation of pRb has been associated with pathological states such as cancer, and therefore, assessing pRb's phosphorylation status is increasingly gaining diagnostic and prognostic value, may be used to inform therapeutic decisions, and is also an important tool for the cancer biologists seeking an understanding of the molecular etiology of cancer. In this chapter, we discuss an immunoblot protocol to detect pRb phosphorylation in two residues, serine 612 and threonine 821, in protein extracts from cancer cells.

Microarray experiments are capable of determining the relative expression of tens of thousands of genes simultaneously, thus resulting in very large databases. The analysis of these databases and the extraction of biologically relevant knowledge from them are challenging tasks. The identification of potential cancer biomarker genes is one of the most important aims for microarray analysis and, as such, has been widely targeted in the literature. However, identifying a set of these genes consistently across different experiments, researches, microarray platforms, or cancer types is still an elusive endeavor. Besides the inherent difficulty of the large and nonconstant variability in these experiments and the incommensurability between different microarray technologies, there is the issue of the users having to adjust a series of parameters that significantly affect the outcome of the analyses and that do not have a biological or medical meaning. In this study, the identification of potential cancer biomarkers from microarray data is casted as a multiple criteria optimization (MCO) problem. The efficient solutions to this problem, found here through data envelopment analysis (DEA), are associated to genes that are proposed as potential cancer biomarkers. The method does not require any parameter adjustment by the user, and thus fosters repeatability. The approach also allows the analysis of different microarray experiments, microarray platforms, and cancer types simultaneously. The results include the analysis of three publicly available microarray databases related to cervix cancer. This study points to the feasibility of modeling the selection of potential cancer biomarkers from microarray data as an MCO problem and solve it using DEA. Using MCO entails a new optic to the identification of potential cancer biomarkers as it does not require the definition of a threshold value to establish significance for a particular gene and the selection of a normalization procedure to compare different experiments is no longer necessary.

Lung cancer is characterized by its aggressiveness and proclivity for early metastasis. A predictor of metastasis of NSCLC rely on post-resection evaluation of tumor histology, which is a severe limitation since only 15% of the patients are diagnosed with resectable disease. Hence, there is a need to characterize biomarkers with metastasis-predicting value in pre-resection small biopsy specimens. Rb is a tumor suppressor inactivated due to hyper-phosphorylation in NSCLC. Our preliminary data relates the Rb S249 phosphorylation to an epithelial-to-mesenchymal transition (EMT), a trait strongly associated to metastasis and increase in CDK5 activator p39. Here, we examined the prognostic significance of Rb S249 and p39 as biomarkers of metastasis staging in non-small cell lung cancer. Our hypothesis is that Rb hyper-phosphorylation in S249 can have prognostic value by being associated with a metastatic phenotype and EMT. To determine p39 and Rb phosphorylation in lung cancer, immunohistochemistry analysis was performed on two lung cancer microarrays. Three pathologists observed the microarrays and scored them using the all-red scoring system. P39 and Rb phosphorylation in S249 were more present in lung tumors than adjacent normal tissue. P39 positive expression had a positive correlation with staging, lymph node involvement, and metastasis. Rb phosphorylation in residue S249 had a positive correlation with histological grading. Using general linear model, we determined that staging can be predicted when combining phosphorylation of Rb S249 and p39. Our analysis suggests that p39 and Rb S249 are likely to be diagnostic biomarkers for lung cancer metastasis. Citation Format: Jaileene Perez-Morales, Stephanie Rivera, Bryan Torres, Xavier Rodriguez, Angel Isidro, Pedro Santiago-Cardona. Expression of p39 and Rb in lung cancer and its correlation with clinicopathologic parameters [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1074.

Lung cancer is characterized by its poor prognosis, aggressiveness, and proclivity for early metastasis. Non-small cell lung cancer (NSCLC) it’s the most common type of lung cancer with a five-year survival rate of 18%. Most predictors of metastasis and recurrence of NSCLC rely on post-resection evaluation of tumor histology, which is a severe limitation since only 15% of the patients are diagnosed with resectable disease. Hence, there is a need to characterize biomarkers with metastasis-predicting value in pre-resection small biopsy specimens. In addition to the proclivity for metastasis, another feature of lung cancers is the inactivation of the retinoblastoma protein (Rb). The retinoblastoma protein (Rb) is a tumor suppressor inactivated due to hyper-phosphorylation in most human cancers, including NSCLC. Our preliminary data relates the Rb S249/T821 phosphorylation signature to an epithelial-to-mesenchymal transition (EMT), a trait strongly associated to metastasis. We found that cells that have hyperphosphorylated Rb in residues S249/T821 express EMT markers such as decreased expression of E-cadherin and integrin α5 as well as increased expression of N-cadherin, Vimentin, and p-FAK. We propose to focus on the cell adhesion and migration-related kinase Cdk5 with its activator p39 as the kinase responsible for engendering the Rb S249/T821 phosphorylation signature. Our hypothesis is that Rb phosphorylation in S249 and T821 due to CDK5 activity can have prognostic value by being associated with a metastatic phenotype and EMT. Studies performed in a tissue microarray in a population of NSCLC adenocarcinoma that had undergone EMT found an up-regulation in p39 and that its expression is correlated with metastasis. Knockdown of CDK5 induced a decrease in phosphorylation of Rb S249/T821 with a decrease in E-cadherin expression. These data suggest that CDK5-dependent phosphorylation of Rb can affect cell adhesion by regulating E-cadherin. Introducing p39 into Rb hypo-phosphorylated cell line induced an increase in phosphorylation of Rb residues. Furthermore, the CDK5-p39-Rb axis might point to the explanation and prediction of the metastatic phenotype in lung cancer. Citation Format: Jaileene Perez-Morales, Darielys Mejias-Morales, Bryan Torres-Collazo, Harry Negron-Pagan, Pedro Santiago-Cardona. A Rb phosphorylation code associated with lung cancer metastasis [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 2845. doi:10.1158/1538-7445.AM2017-2845

In spite of recent therapeutics advances and early detection, lung cancer is still the leading cause of cancer-associated deaths worldwide. The five-year survival rates for its two major subtypes, small-cell lung cancer (SCLC) and non-small cell lung cancer (NSCLC) are estimated to be 6% and 18%, respectively. This high mortality is due to its aggressive nature even when detected at an early stage. Besides its aggressive nature and tendency for early metastasis, another feature of lung cancer is the inactivation of the retinoblastoma protein (Rb) that is observed in both lung cancer subtypes. NSCLC exhibits Rb inactivation by hyper-phosphorylation. Rb is phosphorylated by cyclin dependent kinases (CDKs) and their cyclin regulatory partners. CDK5 is a unique CDK because its regulatory partners, p35 and p39, are not cyclins, hence its function is not cell cycle related. This work aims to employ empirical modeling and statistical experimental design techniques to gain a comprehensive mechanistic understanding of metastasis. Previous mass spectroscopy experiments performed in our laboratory on a panel of NSCLC cell lines identified five-residues (S249, S612, S807, S811, and T821) whose phosphorylation is associated with lack of E-cadherin expression, and therefore may be associated with metastasis. Our hypothesis is that Rb inactivation via phosphorylation due to CDK5 leads to the acquisition of the epithelial-to-mesenchymal transition (EMT) phenotype and the disruption of cell-cell adhesion that characterizes metastatic cells. As a first step to establish a mathematical model the dependency of the controllable variables, Rb residues and CDK5 expression was assessed. To this end, we used siRNA to knockdown CDK5 expression, which caused a decrease in phosphorylation of Rb in the five residues mentioned above and a subsequent decrease in E-cadherin expression. These data suggest that Rb phosphorylation in the five residues is dependent on CDK5 expression. This dependency in our controllable variables points to a nested experimental design. Our next step is to perform invasion assays and lung acini formation to quantify the invasion capacity and the malignant transformation. In the long term, at a clinical setting, a lung cancer tumor may be extracted from a biopsy and immunohistochemistry staining will be performed with antibodies of interest such as Rb phosphorylation residues, E-cadherin, and CDK5/p35 or CDK5/p39. This mathematical model would help provide a prognosis about the invasion capacity and a malignancy prediction of the lung tumor. Citation Format: Jaileene Perez-Morales, Mauricio Cabrera-Rios, Jonathan Gonzalez-Flores, Pedro Santiago-Cardona. Characterizing the non-linear dependency of the CDK5-Rb axis in non-small cell lung cancer. [abstract]. In: Proceedings of the AACR Special Conference on Engineering and Physical Sciences in Oncology; 2016 Jun 25-28; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2017;77(2 Suppl):Abstract nr A30.

Lung cancer remains the leading cause of cancer deaths among men and women worldwide. Its five-year survival rate (17.8%) is by far the lowest among the most common cancers such as colon (65.4%), breast (90.5%), and prostate (99.6%). Early detection and screening still remain challenging, and despite the many recent breakthroughs in lung cancer treatment, improvement in the long-term survival of lung cancer patients is still limited. The inactivation of the retinoblastoma tumor suppressor (Rb) is a major driving force behind tumorigenesis in most cancer types, including lung cancer. Rb is a phosphoprotein traditionally known as a cell cycle repressor. While Rb inactivation in small cell lung carcinomas, osteosarcomas and retinoblastomas occurs by mutational inactivation of the RB1 gene, most other human cancers show Rb inactivation by hyperphosphorylation. The effect of Rb hyperphosphorylation in cell cycle control has been characterized and shown to consist in the abrogation of Rb’s capacity to bind and repress the EF2 transcription factors responsible for triggering S-phase related gene expression. However, our data show that Rb phosphorylation can have novel consequences beyond cell cycle control, and that certain phosphorylations in Rb can impair the cell’s capacity to engage in cell-to-cell interactions. In order to search for phosphorylation patterns that may be associated with cell adhesion defects, we conducted a mass spectroscopy analysis on a panel of E-cadherin-expressing and E-cadherin-deficient cell lines. This analysis identified a four-residues phosphorylation signature (S612, S249, S807 and S821) that is associated to lack of E-cadherin expression. These four residues are strongly phosphorylated in the E-cadherin null cell line H520, relative to the E-cadherin expressing cell line H1666. Consistent with these data, H520 cells overexpress the Cyclin-dependent kinases Cdk2 and Cdk6, both of them known to have Rb as one of its targets. Treatment with CDK inhibitors (Roscovitine, PD00332991, and GW8510) resulted in inhibition of Cdk2, Cdk5, Cdk7, and Cdk9 together with reduced the phosphorylation of Rb S807 and S612, and with the reestablishment of E-cadherin expression. Moreover, shRNA-mediated Rb knockdown caused a marked induction of EMT in our lung cancer cell lines, indicated by decreased E-cadherin expression and elevated N-cadherin levels. Taken together, our data show a Rb phosphorylation signature associated with a non-traditional role for Rb as a regulator of cell adhesion. Given that Rb phosphorylation in specific residues can lead to loss of E-cadherin expression and concomitant loss of cell adhesion, both of them landmarks of the metastatic phenotype, we postulate that this phosphorylation signature may have prognostic value in predicting a proclivity for metastasis of lung cancer cells. Citation Format: Jaileene Perez-Morales, Jonathan Gonzalez-Flores, Alexander De La Rosa, Elvin Estrada, Pedro Santiago-Cardona. A retinoblastoma protein phosphorylation code associated with cell adhesion and invasiveness. [abstract]. In: Proceedings of the AACR Special Conference: Developmental Biology and Cancer; Nov 30-Dec 3, 2015; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Res 2016;14(4_Suppl):Abstract nr A12.