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Background: Cancer patients who self-report tobacco use within the past 30 days are at high risk for relapse and benefit from cessation support. However, there is little data on whether assessment of tobacco use within the past 30 days increases referral rates to a cessation program across cancer disease sites. Methods: Structured tobacco assessments were administered to 426 new adult patients in a Radiation Oncology clinic at a single institution. Multiple logistic regression was used to assess if clinical aspects of cancer diagnosis (age, gender, race, disease site [head/neck or thoracic {HNT} vs. other], stage [advanced vs. early], and purpose [curative vs. palliative]) affected cessation service referral rates identified using assessment of tobacco use within the past 30 days. Results: Complete medical and tobacco use data were compiled for 387 adult cancer patients (91%). Of these, 147 (38%) were women, 259 (67%) were Caucasian, 115 (30%) were black, 177 (46%) were diagnosed with HNT cancer, and 289 (74%) were being considered for curative cancer treatment. Ever smoking was reported in 273 patients (71%), and 83 (21%) reported current smoking everyday or some days. In 180 former smokers who reported time since last cigarette, 21 (12%) reported smoking within with past 30 days and 9 (7%) of these reported smoking within the past day or week. In 21 patients who were identified for cessation support through evaluation of smoking within the past 30 days, 6 (29%) were women, 7 (33%) were black, 15 (71%) were being considered for curative treatment, and 15 (71%) were diagnosed with a HNT cancer. Multiple logistic regression identified that younger age (10 yr OR 2.41, 95% CI 1.47 - 3.98) and HNT cancer (OR 8.13, 95% CI 2.2-30.3) but not gender, race, stage, and purpose were significantly associated with referral based on assessment of tobacco use within the past 30 days. Identification of smoking within the past 30 days increased cessation referrals by 25% from 83 to 114 patients. Conclusions: Assessment of tobacco use within the past 30 days increased referrals to a smoking cessation program from a Radiation Oncology clinic using a standardized tobacco assessment. In clinical trials and practice, including this question has particular benefit for younger patients and patients with head/neck or thoracic tumors. Citation Format: Samuel Lewis Cooper, Katherine Hoover, Elizabeth Garrett-Mayer, Kenneth Michael Cummings, Matthew J. Carpenter, Kelly Crowley, David T. Marshall, Graham W. Warren. Assessment of tobacco use within 30 days increases referral for cessation support. [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 3736. doi:10.1158/1538-7445.AM2015-3736

Tobacco use is the single largest preventable risk factor for developing cancer. Furthermore, tobacco is associated with poor therapeutic outcomes in part because its continued use has been shown to decrease the efficacy of conventional chemotherapy and/or radiotherapy (RT). Preclinical data suggests that nicotine administration decreases RT efficacy by activating the alpha-7-nicotinic acetylcholine receptor (a7-nAChR). Our study evaluated the effects of nicotine and selective a7-nAChR agonists (LY 2087101, PHA 543613, PNU 282987, and TC 1698) administration on radiotherapeutic efficacy. H460 and A549 human lung cancer cell lines were dose escalated with nicotine and the a7-nAChR agonists alone or in combination with single fraction X-ray radiation (2, 6 Gy) and cell survival was assessed by clonogenic assay. Nicotine administration and activation of a7-nAChR by LY 2087101 or PHA 543613 was found to have a radioprotective effect in both cell lines by increasing cell survival following RT. Conversely, activation of a7-nAChR by PNU 282987 or TC 1698 did not modulate radiotherapeutic response. Western blot analysis demonstrated that although all four tested a7-nAChR agonists increased phosphorylation of AKT (S473), only PHA 543613 and LY 2087101 were found to also activate mTOR (S2448). Additional clonogenic studies were performed to further examine the role of mTORC1 in governing response to RT. Inhibition of mTORC1 by rapamycin alone sensitized both A549 and H460 cells to the effect of RT. Interestingly, rapamycin treatment was sufficient to block the radioprotective effects of nicotine, PHA 543613, and LY 2087101 in A549 cells, however, in H460 cells rapamycin did not restore radiotherapeutic efficacy. Moreover, examination of Deptor, a key component of mTOR complexes shown to be associated with RT response, demonstrated that a7-nAChR activation by nicotine, PHA 543613, or LY 2087101 increased Deptor degradation in A549 cells, while a7-nAChR activation decreased Deptor degradation in H460 cells. These observations suggest that differences in the regulation of Deptor level may contribute to the variation in both rapamycin sensitivity and restoration of therapeutic efficacy to RT between H460 and A549 cell lines. Additional studies will be necessary to further elucidate the role of the mTOR complexes in mediating a7-nAChR induced modulation of response to RT. Citation Format: Samantha L. Sobus, Sundaravadivel Balasubramanian, Michelle A. Romano, Graham W. Warren. The role of mTOR in mediating nicotine induced alpha-7 nicotinic acetylcholine receptor activation and the regulation of radiotherapeutic response. [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 3305. doi:10.1158/1538-7445.AM2015-3305

Purpose Tobacco use is not only associated with the development of cancer, but after a patient develops cancer, its continued use is associated with poorer cancer treatment outcomes. Despite this fact, most oncologists do not regularly provide tobacco cessation support. The purpose of this study was to implement an expedited process of patient identification and tobacco cessation for cancer patients. Methods The Michigan Oncology Quality Consortium (MOQC) collaborated with the Michigan Department of Community Health, Blue Cross and Blue Shield of Michigan, and the Michigan Cancer Consortium to enhance tobacco cessation support for cancer patients. A statewide tobacco cessation demonstration project using a lean engineering approach was developed to provide structured tobacco assessments, referral to state quitline resources, and provide tailored cessation support to cancer patients consisting of behavioral counseling and free pharmacotherapy where appropriate. These steps were rapidly deployed in over 30 locations including those in private oncology practices and cancer centers in the first year. Data were analyzed to evaluate participation, retention, and quit rates for cancer patients enrolled in the program. Results Previous reports have described high rates of patient contact and participation in the cessation program. In the 690 patients with at least 12 months since program enrollment, 200 (29%) participated in only 1 call, 313 (45%) participated in 1-2 follow-up calls, and 177 (26%) participated in 3 or more follow-up calls. Follow-up 30 day quit rate data was obtained in 198, 138, and 199 patients at 3, 6, and 12 months respectively. Self-reported quit rates remained largely consistent with 29.8% at 3 months, 27.5% at 6 months, and 30.2% at 12 months. Conclusions The MOQC demonstration project represents a model partnership between payors, clinicians, and statewide quitlines that maintained consistent and moderate quit rates in cancer patients. Large-scale deployment of this model program may provide a clinically efficient and effective mechanism to enhance widespread tobacco cessation efforts for cancer patients. Citation Format: Laura Petersen, Hilary Baca, Karen Brown, Jamie Lindsay, Sarah Mott, Jane Severson, Anna Schulze, Graham Warren. Rapid deployment of statewide tobacco cessation program for cancer patients. [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 3728. doi:10.1158/1538-7445.AM2015-3728

Background: The 2014 Surgeon General's Report concludes that cigarette smoke (CS) causes increased cancer specific mortality. Clinical data support that the effects of CS are reversible, but there are no extant biologic models that have shown therapeutic response profiles for cancer cells exposed to CS. Methods: An acute, subacute, and chronic dose escalated CS exposure model was developed to test the hypothesis that CS decreases therapeutic response to chemotherapy (CT, cisplatin) and radiotherapy (RT, 0, 2, 4, and 6 Gy) in a reversible manner in cancer cells. Results: H460, A549, and FaDu cancer cells were treated with single and multiple exposure CS for up to 12 weeks. Subacute and prolonged exposure to CS significantly decreased response to CT and RT. CS increased survival fraction following CT or RT between 30-157% across all cell lines. Exposure of cancer cells to CS for 6 or 9 weeks followed by removal of CS for 3 and 6 weeks resulted in a partial to complete restoration of baseline therapeutic response in a cell specific manner; however, all cell lines required several weeks to achieve a partial or full restoration of therapeutic response. Dose response curves for CS exposure demonstrate that H460 and A549 cells had a decreased therapeutic response profile at higher CS concentrations while FaDu cells were more sensitive to lower concentrations to CS. Dose response curves further demonstrated that high dose CS exposure was cytotoxic in all cell lines. Results were verified in all cell lines using 2 additional repeated experiments with freshly thawed cancer cells purchased from ATCC over 10 months. Conclusions: CS exposure reproducibly decreases therapeutic response to CT and RT in a dose dependent and reversible manner, reflective of clinical observations. Dose specific effects may vary between cancer cells. Data suggest that the biologic effects of CS removal, such as with acute smoking cessation in cancer patients, may not manifest in an immediate restoration of cancer cell response to CT or RT. Citation Format: Graham W. Warren, Michelle Romano, Samantha Sobus, Sundaravadivel Balasubramanian. Cigarette smoke and therapeutic response to chemotherapy and radiotherapy in cancer cells. [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 5593. doi:10.1158/1538-7445.AM2015-5593

Purpose: The purpose of this study was to evaluate oncologists' preferences for methods to increase delivery of tobacco cessation support to cancer patients. Methods: An email list of 6844 physicians at NCI Designated Cancer Centers was generated using publically available online resources. An online survey was conducted in the fall of 2013 to assess tobacco assessment and cessation practices by physicians, barriers to providing cessation support, and methods to improve access to cessation services for cancer patients. Results of this survey are compared to similar survey findings by the American Society of Clinical Oncology (ASCO) and the International Association for the Study of Lung Cancer (IASLC). Results: Of 6844 electronically delivered survey invitations, we received 887 responses (13%). Consistent with previously published ASCO and IASLC surveys, 90% of physicians agreed that tobacco use by patients affects cancer treatment outcomes (vs. 87-92% in ASCO/IASLC), 90% of respondents reported that they ask patients about their tobacco use (vs. 90% in ASCO/IASLC), and 83% routinely advise tobacco users to quit (vs. 80-82% in ASCO/IASLC). However, only 37% of respondents regularly discussed medications, and 35% actively treated patients (vs. 39-44% in ASCO/IASLC). Most (71%) responding oncologists preferred cessation support be provided at the initial patient consult, and few (11%) were concerned about the use of tobacco cessation medications. Lack of time and lack of available resources were the dominant barriers to the administration of tobacco cessation support to cancer patients by oncologists. Only 10% of oncologists believed they were adequately trained to counsel patients on smoking cessation, but 55% reported that training for someone else in their clinic would increase tobacco cessation support. When asked what provider they prefer to administer cessation support to cancer patients, 16% reported a primary care physician, 18% reported a mid-level (nurse practitioner or physician assistant), 50% reported any other clinical staff, and only 1% preferred to personally provide cessation support. Whereas most (58%) oncologists preferred to have dedicated cessation services within their institution, 38% reported they would use dedicated services either within, or outside their institution. Conclusions: Most oncologists at NCI Designated Cancer Centers recognize the importance of tobacco cessation, but prefer to have tobacco cessation support for cancer patients provided by another trained professional. Citation Format: Alix Pommerenke, Anthony Alberg, Thomas H. Brandon, Ivana Croghan, Michael K. Cummings, Carolyn Dresler, Ellen R. Gritz, Roy Herbst, Scott Leischow, James R. Marshall, Benjamin Toll, Graham Warren. Physician preferences in tobacco cessation support for cancer patients: A survey of physicians at National Cancer Institute Designated Cancer Centers. [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 5049. doi:10.1158/1538-7445.AM2014-5049

Purpose: Tobacco use is associated with poor therapeutic outcome because it in part decreases the efficacy of conventional cancer treatments including chemotherapy and/or radiotherapy (RT). However, preclinical data suggests that nicotine replacement decreases effectiveness of cancer treatment potentially through action of the α7-nicotinic acetylcholine receptor(nAChR). The purpose of this study is to examine the role of the α7-nAChR in mediating response to RT, evaluate the effect of nAChR and non-nAChR based pharmacologic tobacco cessation agents on response to RT, and identify critical downstream signal transduction pathways associated with changes in response. Methods: α4, α7 and β2-nAChR subunit expression was verified in H460, A549 (lung cancer) and FaDu (oropharyngeal cancer) using western blot. All three cell lines were dose-escalated with nicotine, α7-nAChR selective agonists (TC 1698, PNU 282987, PHA 543613, LY 2087101), or pharmacologic tobacco cessation agents (varenicline, bupropion, hydroxybupropion, cytisine) alone or in combination with single fraction 2Gy X-ray radiation treatment. Clonogenic survival assays were used to assess cell survival and downstream activation of radioprotective pathways (Akt, ERK, mTOR, MMP2, and hypoxia inducible factor 1-alpha [HIF-1α]) were analyzed by western blot. Results: Nicotine administration and activation of the α7- nAChR by PHA 543613 or LY 2087101 were found to have a radioprotective effect in the three tested cancer cell lines as indicated by increased cell survival. Conversely, TC 1698 and PNU 282987 treatment did not modulate cell survival when in combination with radiation. Nicotine, PHA 543613, LY 2087101, TC 1698 and PNU 282987 were all shown to induce a time and dose dependent increase in p-AKT, p-ERK, and HIF-1α. However, treatment with PHA 543613 or LY 2087101 induced an increase in mTOR phosphorylation at S2448 with no change in phosphorylation with TC 1698 or PNU282987. Furthermore, other nAChR-involved agents, cytisine and varenicline, showed mixed results with varenicline inducing radioprotection while cytisine did not alter response to RT. The non-nAChR involved agents, bupropion and hydroxybupropion, did not modulate response to radiotherapy and dose escalated bupropion did not change activation of p-Akt, p-ERK, or HIF-1α. Conclusions: Collectively, these data suggests that the non-nAChR based agent bupropion and its metabolite hydroxybupropion may be the ideal tobacco cessation strategy for cancer patients in order to maintain the biologic effectiveness of radiation treatment. Data further suggests that downstream activation of mTOR may be critical to modulating α7- nAChR based changes in response to RT. Citation Format: Samantha L. Sobus, Michelle A. Romano, Graham W. Warren. The role of α7 nicotinic acetylcholine receptor activation and pharmacologic tobacco cessation agents on response to radiotherapy. [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 3948. doi:10.1158/1538-7445.AM2014-3948

Purpose: Tobacco use is associated with poor therapeutic outcomes in cancer patients in part due to decreased therapeutic response to conventional cancer treatment such as chemotherapy and/or radiotherapy (RT). Tobacco cessation is warranted to improve cancer treatment outcomes, but preclinical data suggest that nicotine may decrease the biologic effectiveness of cancer treatment. The purpose of this study is to evaluate if nicotinic acetylcholine receptor (nAChR) and non-nAChR based tobacco cessation pharmacotherapy agents alter response to radiotherapy. Methods: Human lung cancer (A549, H460) and oropharyngeal cancer (FaDu) cell lines were treated with nicotine, a7-nAChR agonists, varenicline, or bupropion alone or in combination with RT. Mice xenografts were used to assess the effect of nicotine administration during RT. Cell survival and downstream activation of radioprotective pathways (Akt, ERK, mTOR, MMP, and hypoxia inducible factor 1-alpha [HIF-1a]) were analyzed. Results: Nicotine administration increased survival in all cell lines. Similarly, administration of the selective a7-nAChR agonists LY 2087101 and PHA 54613 was also found to increase cell survival with RT. Nicotine, LY 2087101, and PHA 54613 were all found to activate p-ERK 1/2, p-AKT (Ser-473), mTOR (S2448), and hypoxia inducible factor 1-alpha (HIF-1a) in a dose and time dependent manner. Tumor xenografts receiving nicotine administration during RT showed evidence of increased tumor regrowth in comparison non-nicotine treated controls. Treatment with varenicline also increased cell survival following RT at doses lower than with nicotine treatment; however, treatment with bupropion (an FDA approved non-nAChR based tobacco cessation agent) did not increase cell survival and exhibited a moderate cytotoxic effect following RT at high doses. Dose escalated bupropion did not increase activation of radioprotective pathways. Conclusions: These are the first data to suggest that bupropion may be an ideal FDA approved tobacco cessation agent in cancer patients to prevent potential biologic changes in therapeutic response associated with nAChR based cessation strategies. Citation Format: Samantha L. Sobus, Michelle A. Romano, Janelle Mackowiak, Krista Pundt, Graham W. Warren. Pharmacologic tobacco cessation agents and response to radiotherapy. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 66. doi:10.1158/1538-7445.AM2013-66

Purpose: Smoking has been associated with poor outcomes in cancer patients, but a detailed review of the effects of smoking on cancer treatment outcomes is necessary to effectively implement structured tobacco assessments and cessation into clinical trials design. Methods: We searched Pubmed citations between 1950 and 2011 with the objective of identifying any reports relating smoking with cancer and non-cancer related outcomes in cancer patients. Our search yielded 34,000 pertinent citations of on smoking and cancer. Over 5000 of these citations appeared to include data on smoking as it might be related to cancer treatment outcomes and of these more than 800 were felt to have sufficient information to examine how smoking affects cancer treatment outcomes. Results were cataloged in tables and cross-references according to cancer disease site. Results: The vast majority of pertinent reports provide evidence showing that smoking increases the risk of cancer and non-cancer related mortality as well as treatment related adverse events. The adverse effects of smoking and treatment outcomes are observed in all major cancer disease sites (lung, head/neck, breast, gastrointestinal, gynecologic, leukemia/lymphoma, prostate, and non-prostate genitourinary). A limited number of studies examine how smoking cessation affects treatment outcomes and generally these studies show that cessation of tobacco use is associated with fewer adverse treatment outcomes. Measurement of smoking behavior is a problem due to the reliance on self-reported behavior and lack of standardization in classifying smoking status. In most studies a patients’ cigarette use is categorized into ever smoking or current/former/never smoking; however, there is a broad diversity of definitions used making it less clear exactly how smoking affects treatment outcomes. Despite methodologic issues in published reports, results demonstrate a consistent pattern that smoking increases the risk of adverse outcomes across a broad spectrum of cancer disease sites. Conclusions: This review provides evidence that smoking adversely affects cancer related and non-cancer related outcomes in patients. What is less certain is how past and/or current smoking might differentially modify treatment outcomes for patients. Clinical trials must give strong consideration for prior tobacco history, current tobacco use, and efforts to track tobacco cessation during and following cancer treatment. Citation Format: Graham W. Warren, James R. Marshall, K. Michael Cummings. Smoking, cancer treatment, and design of clinical trials. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2421. doi:10.1158/1538-7445.AM2013-2421

Purpose: The lack of accurate prospective tobacco assessments in cancer patients limits the ability to provide tobacco cessation and to accurately evaluate the true impact of tobacco use on cancer treatment outcomes. The purpose of this study was to evaluate tobacco use assessment in patients enrolled in actively accruing cancer clinical trials. Methods: Protocols and forms for 155 actively accruing trials in the NCI's Clinical Trials Cooperative Group program were evaluated for detailed tobacco use assessment at enrollment and through follow-up using a structured coding instrument. Results: Of the 155 clinical trials reviewed, the mean accrual start date was 2008.6 (range 2003-2011), 74.1% were phase III trials, and the total potential accrual population was 106,985 patients. Any form of tobacco use (i.e., cigarette, pipe, cigar, smokeless tobacco, or unspecified “tobacco”) was assessed in 29% of trials (46% of total patient accrual) at enrollment. Current cigarette use at enrollment was assessed in 21.9% of trials and former cigarette use was assessed in 21.3% of trials. Only 4.5% of trials assessed any form of tobacco use during follow-up. Secondhand smoke exposure was captured in 2.6% of trials at enrollment and only 1 trial during follow-up. None of the trials assessed nicotine dependence or interest in quitting at any point during enrollment or treatment. Tobacco assessment varied by type of cancer with a higher proportion of patients being assessed in lung or head and neck cancer trials. Conclusion: Most actively accruing cooperative group clinical trials do not assess tobacco use. Failure to incorporate standardized tobacco assessments into funded national clinical trials will continue to limit the ability to provide evidence based cessation support and will limit the ability to accurately understand the precise impact of tobacco use on cancer treatment outcomes. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 648. doi:1538-7445.AM2012-648

Purpose: Nicotine is a systemically available component of tobacco known to decrease the effectiveness of chemotherapy and/or radiotherapy (RT) however, the mechanisms of decreased RT response are not well characterized. The purpose of this study is to evaluate the effects of nicotine and activation of the alpha-7 nicotinic acetylcholine receptor (a7-nAChR) on regulating cell survival following RT and to identify potential mechanisms of regulation. Methods: Human lung cancer (A549, H460) and oropharyngeal cancer (FaDu) cell lines were treated with nicotine alone or in combination with nAChR inhibitors, MEK inhibitors, PI3K inhibitors, or mTOR inhibitors followed by treatment with RT to evaluate cell survival using clonogenic assays. The effects of a7-nAChR agonists on RT were also evaluated. Western blots were performed to evaluate nAChR subunit expression profiles in a broad spectrum of cancer cell lines and to evaluate the effects of nicotine (alone or with inhibitors listed above) or agonists on downstream signal transduction. Results: Nicotine administration 2 hours prior to RT increased survival in all cell lines. nAChR expression was confirmed in several cell lines including A549 and H460 cells. Inhibition of specific nAChRs in A549 and H460 cells demonstrated that the a7-nAChR prevented the effects of nicotine on RT in both cell lines. Nicotine administration increased p-ERK 1/2, p-Akt (Ser-473), mTOR (S2448), activated matrix metalloproteinase 2 (MMP2), and hypoxia inducible factor 1-alpha (HIF-1a). Inhibition of MEK prevented ERK 1/2 phosphorylation, but did not prevent the effects of nicotine on RT. Inhibition of PI3K prevented Akt phosphorylation, MMP2 activation, and HIF-1a induction with concomitant inhibition of the effects of nicotine on RT. Administration of an mTOR inhibitor (rapamycin) significantly sensitized cells to RT, but did not prevent the pro-survival effects of nicotine. Administration of selective a7-nAChR agonists (LY 2087101, PHA 54613, PNU 282987, TC 1698) demonstrated that all increased p-ERK 1/2 and p-Akt; however, only LY 2087101 and PHA 54613 increased cell survival with RT. Further analysis demonstrates that only LY 2087101 and PHA 54613 increased mTOR (S2448) while there was no change in mTOR (S2448) with PNU 282987 or TC 1698. Conclusions: Data demonstrates that activation of the a7-nAChR can decrease the effectiveness of RT. Results suggest that PI3K and mTOR may play a central role in conferring resistance to radiation by nicotine or during a7-nAChR activation. Further analyses of critical mechanisms are required to understand the adverse systemic effects of tobacco on cancer treatment. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3456. doi:1538-7445.AM2012-3456

Introduction: Selective serotonergic reuptake inhibitors (SSRIs) are well-established antidepressants commonly used in cancer patients. Studies suggest that SSRIs may play a role in increased cell death in cancer cells; however, the use of SSRIs as adjunctive agents to be used in combination with conventional cancer treatment is not well established. The purpose of this study was to evaluate the effect of SSRIs on cell survival in combination with radiation therapy (XRT). Methods: Human lung cancer lines (A-549 and H-460) and squamous oropharyngeal cancer cells (FaDu) were treated with several dose escalated SSRIs (fluoxetine, sertraline, paroxetine) alone or in combination with dose escalated XRT (0,2,4,6 Gy) to evaluate the cytotoxic effects of SSRIs using clonogenic survival assays. Western blot analysis was used to evaluate apoptosis (cleaved Caspase-3 and cleaved PARP) and autophagy (LC3-II) to assess potential mechanisms of the effects of SSRIs on XRT. Results: Fluoxetine, paroxetine, or sertraline alone had a cytotoxic effect on cancer cells in a concentration dependent manner. Sertraline and paroxetine at 2.5-5uM appeared to have an additive effect on XRT with severe cytotoxicity noted at higher concentrations. In contrast, fluoxetine appeared to have a sensitizing effect on cells at 1-10uM in a tumor cell specific manor. Western blot analysis demonstrated that fluoxetine administration decreased cleaved Caspase-3 and cleaved PARP expression at 24 or 48 hours. Evaluation of autophagy using LC3-II demonstrated no change with XRT; however, treatment with fluoxetine increased the expression of LC3-II at 24 and 48 hours in the presence or absence of XRT. Conclusions: Most SSRIs appear to have a cytotoxic effect alone or in combination with XRT. Fluoxetine appears to have a unique sensitizing effect on XRT in cancerous cells possibly through an increased induction of autophagy. Data supports further examination of fluoxetine as an adjunctive agent in combination with XRT. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2652. doi:10.1158/1538-7445.AM2011-2652

Background: Current cancer diagnosis and treatment increasingly incorporates biomarker expression in the prognostication and therapeutic stratification of cancer patients; however, there is no accurate assessment of exposure to environmental agents such as nicotine that may significantly impact biomarker expression patterns. Methods: Athymic nude mice with human H460 lung cancer cell xenografts were stratified into three treatment groups (n=5 per group) beginning when tumors reached 5 mm in maximal dimension: control (CON), short term nicotine (STN, 60 ug subcutaneously every other day x 6 days), or long term nicotine (LTN, 60 ug SC every other day until tumor explant). All tumors were explanted at endpoint defined when tumors reached 15 mm in maximal dimension. Tumors were analyzed immunohistochemically for hypoxia (using CA-IX expression) and hypoxia-inducible factor 1-alpha (Hif-1) expression. This protocol was approved by the Institutional Animal Care and Use Committee at the University of Kentucky. Results: Nicotine administration had no effect on tumor growth rate and all tumors reached endpoint within a median of 9 days following treatment initiation (range 8-11 days). Areas of hypoxia were defined using CA-IX expression. Tumor hypoxia was not significantly affected by nicotine administration. Moreover, Hif-1 expression was only minimally increased in areas of hypoxia in CON animals. In STN animals, Hif-1 expression was very similar to CON animals. In contrast, LTN animals demonstrated widespread marked increases in Hif-1 expression at the viable: necrotic tumor interface. Separately, xenografts removed at day 6 demonstrated similar widespread Hif-1 expression confirming that Hif-1 expression is significantly increased after 6 days of nicotine administration. Conclusions: This study demonstrates that widespread Hif-1 expression may be dominated by the presence of nicotine rather than hypoxia. Furthermore, acute removal of nicotine results in the restoration of basal low-level Hif-1 expression within 3 days of nicotine removal. This data may significantly limit the prognostic utility of clinical Hif-1 expression in the absence of simultaneous nicotine assessment. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3729.

Background: Continued tobacco use during cancer treatment has been associated with poor outcome. Nicotine replacement is currently used as a standard of care for smoking cessation in cancer patients; however, prior data demonstrates that nicotine can promote tumor growth and decrease the effectiveness of cancer treatment in vitro. Methods: Athymic nude mice with human H460 lung cancer cell xenografts were stratified into three Groups beginning when tumors reached 5 mm in maximal dimension: control (CON), short term nicotine (STN, 60 ug subcutaneously every other day x 6 days), or long term nicotine (LTN, 60 ug SC every other day until tumor endpoint). Within each Group, mice were further stratified into three Treatments consisting of saline (SAL), radiotherapy (RT, 3 Gy daily x 5 days), or chemoradiotherapy (CRT, RT + concurrent cisplatin intraperitoneally 3 mg/kg daily x 5 days). Tumor volumes were measured and compared between Treatments and Groups (n=10 per Treatment within each Group). This protocol was approved by the Institutional Animal Care and Use Committee at the University of Kentucky. Results: In CON animals, RT (199 uL at day 30) or CRT (100 uL at day 30) significantly prevented tumor regrowth as compared with control SAL animals (864 uL at day 10 requiring euthanasia as per protocol). As expected, CRT was more effective than RT (p Conclusions: Results demonstrate that nicotine significantly reduces the efficacy of RT and CRT in vivo. Moreover, data suggest that nicotine use during treatment is the primary determinant of decreased therapeutic response. These results may significantly alter current recommendations for smoking cessation in cancer treatment populations. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 1408.