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Simple Summary Glycogen synthase kinase-3 beta is a protein kinase implicated in the promotion and development of various cancers, including pancreatic cancer. In cell culture and animal studies, drugs targeting the inhibition of this protein show treatment potential in pancreatic cancer. Studies show targeting this protein for treatment may overcome resistance to conventional chemotherapy in pancreatic tumors. Early-stage clinical trials are currently studying small molecule inhibitors targeting glycogen synthase kinase-3 beta and interim results show favorable results. Recent studies also suggest that targeting this protein will create synergy with immunotherapy, such as checkpoint inhibitors. Future studies should aim to study new combination treatments involving glycogen synthase kinase-3 beta targeting drugs with chemotherapy and immunotherapy in pancreatic cancer. Abstract Glycogen synthase kinase-3 beta is a ubiquitously and constitutively expressed molecule with pleiotropic function. It acts as a protooncogene in the development of several solid tumors including pancreatic cancer through its involvement in various cellular processes including cell proliferation, survival, invasion and metastasis, as well as autophagy. Furthermore, the level of aberrant glycogen synthase kinase-3 beta expression in the nucleus is inversely correlated with tumor differentiation and survival in both in vitro and in vivo models of pancreatic cancer. Small molecule inhibitors of glycogen synthase kinase-3 beta have demonstrated therapeutic potential in pre-clinical models and are currently being evaluated in early phase clinical trials involving pancreatic cancer patients with interim results showing favorable results. Moreover, recent studies support a rationale for the combination of glycogen synthase kinase-3 beta inhibitors with chemotherapy and immunotherapy, warranting the evaluation of novel combination regimens in the future.

BACKGROUND GSK3β serine/threonine kinase regulates metabolism and glycogen biosynthesis. GSK3β overexpression promotes tumor progression and resistance through NF-κB and p53 apoptotic pathways. GSK3β inhibits immunomodulation by downregulating checkpoints, e.g. PD-L1 and LAG-3, and increasing NK and T-cell mediated tumor killing. 9-ING-41 is a small-molecule, potent, selective GSK3β inhibitor with preclinical activity. In chemoresistant PDX glioblastoma models, 9-ING-41 enhanced lomustine antitumor effect. METHODS Patients with refractory malignancies were treated with 9-ING-41 monotherapy (n=65) or combined with 8 cytotoxic regimens after prior exposure (n=162) in the first-in-human study (NCT03678883). The recurrent gliomas subset was treated with 9-ING-41 monotherapy IV TIW q21day cycles at 3.3, 5, 9.3, 15mg/kg, or combined with lomustine 30 mg/m² PO weekly q84day cycles. Primary objective was safety and tolerability. RESULTS An RP2D of 15mg/kg IV TIW was confirmed across all 9 regimens, no accentuation of chemotherapy toxicity noted. Of 18 glioma patients enrolled, 13 were glioblastoma, 2 anaplastic astrocytomas, 1 anaplastic oligodendroglioma, and 1 diffuse astrocytoma; 6 female, 12 male; median age 52 (30-69) years; median ECOG was 1 (0-2). All received initial radiation and temozolomide (18/18), prior salvage therapies included nitrosoureas (15/18), bevacizumab (8/18), TTFields (6/18), checkpoint inhibitor (4/18). Median recurrences 3 (1-6). NGS alterations included: IDH/wildtype (11), IDH/mutation(3); 1p19q/codeletion(10); MGMT/unmethylated(11), MGMT/methylated(1); EGFR/amplification(6), EGFR/v3mutation(3), TERT/mutation(6), PTEN/loss(3), NF1/rearrangement(2), ATRX/loss (2), TP53/mutation(4), CDKN2A/deletion(2), RB1/loss(1), PALB2/mutation(10). Four patients received 9-ING-41 monotherapy, 14 concurrently treated with lomustine. No SAEs or grade 3/4 AEs attributed to 9-ING-41 noted, only G1/2 vision changes (9/18, 50%), infusion reactions (4/18, 22%). Lomustine-related toxicities included G3/4 thrombocytopenia (3/14, 21%), and G1/2 fatigue (4/14, 28%). Median therapy duration was 55 (4-305); 1 partial response ( >50%) noted with 9-ING-41/lomustine. Median PFS and OS were 1.9 (0.3-11.1) and 6.0 (1.6-16.6) months, respectively. CONCLUSIONS 9-ING-41 plus/minus lomustine is safe and warrants further study in glioma patients.

The field of tumor immunology has faced many complex challenges over the last century, but the approval of immune checkpoint inhibitors (anti-cytotoxic T-lymphocyte-associated protein 4 [CTLA4] and anti-programmed cell death-1 [PD-1]/PD-ligand 1 [PD-L1]) and talimogene laherparepvec (T-VEC) for the treatment of metastatic melanoma have awakened a new wave of interest in cancer immunotherapy. Additionally, combinations of vaccines and oncolytic viral therapies with immune checkpoint inhibitors and other systemic agents seem to be promising synergistic strategies to further boost the immune response against cancer. These combinations are undergoing clinical investigation, and if successful, will hopefully soon become available to patients. Here, we review key basic concepts of tumor-induced immune suppression in malignant melanoma, the historical perspective around vaccine development in melanoma, and advances in oncolytic viral therapies. We also discuss the emerging role for combination approaches with different immunomodulatory agents as well as new developments in personalized immunization approaches.

BackgroundNeuroblastoma (NBL) is an embryonic tumor originating from neural crest precursor cells. It is one of the most prevalent pediatric solid tumors, with an estimated five-year progression free survival of less than 50% in stage-4 patients. While T-cell mediated therapies, including immune checkpoint inhibition, have marked activity against many cancers in adults, NBL is relative refractory to these approaches. One reason for the limited success of the existing immune therapies is the extremely low peptide/MHC-I complex expression of NBL cells. There is an unmet need to improve immunogenicity of NBL, and thus to enhance the cytotoxic T lymphocyte (CTL)-mediated anti-tumor response. The serine/threonine kinase GSK-3beta is overexpressed by NBL and has been proposed as a therapeutic target. This kinase is broadly involved in energy metabolism pathways and thus may regulate peptide/MHC-I complex display at the tumor cell surface. 9-ING-41 has broad spectrum pre-clinical anti-cancer activity, including against NBL. It has established efficacy and a favourable safety profile in adult patients with refractory cancers – studies in children are underway. IFN-y upregulates MHC-I expression by inducing the expression of multiple genes related to MHC-I antigen processing and presentation.MethodsWe assessed the potential of 9-ING-41 to increase immunogenicity, using four distinct NBL cell lines which we cultured for 24 or 72 hours in the continuous presence of IFN-y and/or 9-ING-41. We measured cell surface expression of MHC-I and PD-L1 using flow cytometry analyses.ResultsIn GIMEN, SH-SY5Y, SK-N-BE, and IMR32 NBL lines, we found that 9-ING-41 significantly increased IFN-y-mediated MHC-I surface expression on all 4 cell lines, markedly so in the cell lines without MYCN amplification. PD-L1 expression was also upregulated in two of the cell lines, suggesting that 9-ING-41 combined with PD-1/PD-L1 immune checkpoint blockade is worthy of further investigation.ConclusionsThese data indicate that 9-ING-41 may facilitate recognition and killing of NBL by CTL and support the further development of 9-ING-41 as a potential treatment for NBL.

Background: Studies have shown that implantation of cardiac resynchronization therapy (CRT) devices could reduce morbidity and mortality in heart failure patients. Aims of this study are to examine the temporal trends and outcomes in CRT device implanted cancer survivors. Method: This study was a retrospective analysis of Nationwide Inpatient Sample during the period 2005-2014. Patients ≥18 years and who underwent CRT device implantation were included in the study. CCS codes 11-44 were used to identify cancer survivors. The International Classification of Diseases-9th Revision-Clinical Modification (ICD-9-CM) primary procedure codes 00.50 and 00.51 were used to identify patients with device implantation. Association between cancer diagnosis and in-hospital mortality and complications were determined using multivariable analyses. Result: During 2005 to 2014, a total of 392,758 inpatient CRT implantations were performed in the United States. Nearly 8.3% of these procedures were performed among patients with history of cancer. Number of procedures among cancer survivors decreased from 3127 in 2005 to 2115 in 2015. However, proportion of cancer patients undergoing CRT implantations increased from 6.8% in 2005 to 10.3% in 2014 (relative increase: 51.4%, P Discussion: Our study found that proportion of CRT implantations among cancer survivors increased from 6.8% to 10.3% during the study period. In addition, CRT implantations among cancer survivors were associated higher in-hospital complications. Strategies like prehabilitation and close collaboration between cardiologists and oncologists could improve outcome and general well-being of cancer survivors undergoing CRT implantations.

2051 Background: GSK-3β, a serine/threonine kinase, is a key regulator of metabolism and glycogen biosynthesis. GSK-3β aberrant overexpression promotes tumor progression and chemotherapy resistance through NF-κB and p53-mediated apoptotic pathways. GSK-3β inhibition impacts immunomodulation through downregulation of checkpoints, such as PD-L1 and LAG-3, and increasing NK and T-cell mediated killing of tumor cells. 9-ING-41 is a small-molecule potent selective GSK-3β inhibitor with preclinical antitumor activity against several tumor types. In chemoresistant PDX models of glioblastoma (GBM), 9-ING-41 enhanced the antitumor effect of CCNU and CPT-11. Methods: In the first-in-human study (NCT03678883), patients (pts) with refractory malignancies received 9-ING-41 monotherapy (n = 65) or in combination with one of 8 cytotoxic regimens after prior treatment with the same chemotherapy (n = 162). We report the subset of pts with recurrent gliomas treated with 9-ING-41 monotherapy IV twice a week in 21-day cycles at different dose levels (3.3, 5, 9.3, 15mg/kg), or in combination with lomustine 30 mg/m² orally once weekly in 84-day cycles. Primary objective was safety and tolerability. Results: An RP2D of 15mg/kg IV was confirmed across all 9 regimens, no accentuation of chemotherapy-related toxicity noted. Of 18 glioma patients enrolled, 13 were GBM, 2 anaplastic astrocytomas, 1 diffuse astrocytoma, and 1 anaplastic oligodendroglioma. Four patients received single agent 9-ING-41, 14 treated concurrently with lomustine. Demographics: 6 female, 12 male; median age 52 (30-69) years; median ECOG was 1 (0-2). All received first-line radiation and temozolomide (18/18), prior therapies for recurrences included nitrosoureas (15/18), bevacizumab (8/18), TTFields (6/18), immune checkpoint inhibitor (4/18). Median recurrences 3 (1-6). Genomic alterations from available NGS reports included: IDH WT (11), IDH mutation (3), MGMT promoter unmethylated (11), MGMT promoter methylated (1), 1p19q co-deletion (10), EGFR amplification (6), EGFR v3 mutation (3), TERT promoter mutation (6), PTEN loss (3), NF1 rearrangement (2), ATRX loss (2), TP53 mutated (4), CDKN2A deletion (2), RB1 loss (1), PALB-2 mutation (10). No SAEs or grade 3/4 AEs attributed to 9-ING-41 were noted; AEs included G1/2 transient vision changes (9/18, 50%), infusion reactions (4/18, 22%). Side effects from lomustine included: G3/4 thrombocytopenia (3/14, 21%), and G1/2 fatigue (4/14, 28%). Best overall response: 1 minimal response (-43%) after 2 cycles of 9-ING-41 and lomustine. Median days on therapy was 55 (4-305), 4/18 (22%) were stable for 20 weeks or longer. Conclusions: These results show 9-ING-41 alone or in combination is safe and warrants further study in glioma patients. Clinical trial information: NCT03678883.

Immunotherapy has been one of the recent major breakthroughs in cancer therapy. The basic mechanism of immunotherapy agents is to facilitate the immune system to view cancer cells as a foreign presence. The recent success demonstrated by immune checkpoint inhibition in melanoma has launched a boom in immune checkpoint inhibitor trials in several different histologies, but these unfortunately have not shown the same outcome as melanoma. There still exists a significant gap to bridge in therapeutic improvement of these therapies, with patient selection still a major unresolved issue.

Due to its extremely high mortality rates, strong efforts continue to be made to develop new therapies in the treatment of pancreatic adenocarcinoma. The use of combined modality chemoradiotherapy for the treatment of pancreatic adenocarcinoma remains an approach with both promise and controversy. This article reviews the conflicting data with regards to role of combined modality therapy in pancreatic adenocarcinoma and provides an update on current studies in the field.

Background: We have previously identified Glycogen Synthase Kinase-3 beta (GSK-3β) nuclear accumulation as a hallmark of pancreatic ductal adenocarcinoma (PDAC) and demonstrated the antitumor effects of GSK-3β tool compound inhibitors in vivo (Ougolkov, Clin Cancer Res 2006). 9-ING-41 (Actuate Therapeutics Inc), a first-in-class, small-molecule potent selective GSK-3β inhibitor, has significant preclinical antitumor activity in PDAC. 9-ING-41 is synergistic with gemcitabine (GEM) in significantly reducing PDAC cell proliferation via impairment of ATR/Chk1 DNA damage response (Ding, Clin Cancer Res 2019). These data provided the rationale for inclusion of patients with PDAC in the 1801 phase 1/2 study evaluating 9-ING-41 alone and in combination with chemotherapy, including GEM and GEM plus nab-paclitaxel. Methods: Parts 1 and 2 of the 1801 phase 1/2 study evaluate the safety and tolerability of 9-ING-41 in patients with refractory malignancies. Part 1 establishes the recommended phase 2 study dose (RP2D) for 9-ING-41 monotherapy, while Part 2 assesses 9-ING-41 in combination with chemotherapies. For Part 1, patients received twice-weekly IV infusions of 9-ING-41 (21-day cycle). Based on prior treatment, Part 2 for those with PDAC consists of 9-ING-41 plus GEM (1000 to 1250 mg/m2 days 1 and 8; 21-day cycle) or 9-ING-41 plus GEM and nab-paclitaxel (125 mg/m2). Part 3 (Simon 2-stage phase 2 at the RP2D) will assess clinical benefit in patients with refractory PDAC treated with 9-ING-41-based combinations at the RP2D established in Part 2. Response is defined by response evaluation criteria in solid tumors (RECIST) criteria in evaluable lesions. The study is opening in 25 sites globally and will accrue approximately 300 patients. Results: To date, the study has accrued 34 patients with advanced solid tumors (n=33) or hematologic malignancies (n=1). Four dose levels (1, 2, 3.3, and 5 mg/kg) have been completed without dose-limiting toxicities (DLT). Part 2 of the study evaluating combination of 9-ING-41 with chemotherapy agents has been activated. Six patients with advanced PDAC have enrolled. One patient has completed 5 cycles of treatment with 9-ING-41 (1mg/kg) with stable disease (SD) as the best response. Another patient with SD as the best response has completed 6 cycles and now is joining the arm combining 9-ING-41 with GEM after disease progression. Four additional patients are awaiting first response assessment with enrollment ongoing. 9-ING-41-attributable AE to date are Grade 1 transient visual changes (color perception). Conclusions: 9-ING-41 is well tolerated. Preclinical data support the inclusion of patients with advanced PDAC on clinical studies of 9-ING-41. Citation Format: Benedito A. Carneiro, Malika Dhawan, Brittany Borden, Ludimila Cavalcante, Howard Safran, Andrey Ugolkov, Andrew P. Mazar, Daniel D. Billadeau, Francis J. Giles, Pamela Munster. GSK-3β blockade with 9-ING-41 in pancreas cancer: The 1801 phase 1/2 study [abstract]. In: Proceedings of the AACR Special Conference on Pancreatic Cancer: Advances in Science and Clinical Care; 2019 Sept 6-9; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2019;79(24 Suppl):Abstract nr B09.

96 Background: ACC is an exceedingly rare malignancy with an estimated incidence of 0.7 cases per million per year. Patients presenting with metastatic disease have an estimated 5-year survival rate of < 15%, without effective standard treatment options. Immunotherapy approaches such as checkpoint inhibitors have emerged as effective therapies for multiple malignancies. Nivolumab, a monoclonal IgG4 antibody against the programmed death-1 (PD-1) receptor on T-cells, acts by inhibiting the interaction with PD-L1 and PD-L2 and thus eliciting an increased anti-tumor immune response. This phase II study will assess the efficacy of nivolumab monotherapy according to objective response rate (ORR) in patients with advanced ACC – the study is currently in the patient enrollment phase. Methods: Patients with metastatic or locally advanced ACC with disease progression after treatment with ≥ 1 line of therapy (including mitotane and/or chemotherapy), or not eligible for first-line chemotherapy are currently being enrolled on this phase II study. Nivolumab is given at a fixed dose of 240mg IV q2 weeks until confirmed disease progression, unacceptable toxicity, or withdrawal. Planned accrual of up to 33 patients will follow a Simon two stage design with interim analysis for efficacy after 10 evaluable patients are enrolled. Results: This is an initial report of the first 7 evaluable patients enrolled on study. Median age 57 years; female (4/7); endocrine hyperfunction (2/7); prior chemotherapy with EDP +/- mitotane (5/7). Patients were treated with a median of 4 doses of nivolumab. Median time to progression of 8 weeks. Best overall response rate thus far has been progression of disease in 5 pts, with 2 pts pending evaluation. Grade 3 or 4 adverse events (AEs) at least possibly related to nivolumab: tremor, hypokalemia (1 pt each). Reported grade 1 and 2 AEs: LFT elevation (3 pts), lower extremity edema (2 pts), lymphopenia, urinary frequency and fatigue (1 pt each). Conclusions: Nivolumab has been well tolerated in this population of patients with metastatic ACC. Clinical trial information: NCT02720484.

Background: Axillary lymph node status at diagnosis remains the strongest predictor of long-term survival in breast cancer. Patients with more than ten axillary lymph nodes at diagnosis have a poor long-term survival. In this single institutional study, we set out to evaluate the prognosis of this high-risk group in the era of multimodality therapy. Materials and Methods: In this retrospective study, we looked at all breast cancer patients with greater than ten axillary lymph nodes diagnosed at Mount Sinai Medical Center (MSMC) from January 1st 1990 to December 31st 2007 (n=161). In the univariate analysis, descriptive frequencies, median survival, and 5- and 10-year survival rates were estimated for common prognostic factors. A multivariate prognostic analysis for time-to-event data, using the extended Cox regression model was carried out. Results: With a median and mean follow-up of 70 and 89.9 months, respectively, the overall median survival was estimated to be 99 months. The five-year disease-free survival (DFS) was 59.3% and the ten-year DFS was 37.9%, whereas the five- and ten-year overall survival (OS) was 66.6% and 43.9%, respectively. Multivariate analysis revealed a significant improvement in DFS among black patients compared to whites (p=0.05), improved DFS and OS among young patients (ages 21-45) compared to elderly patients (age greater than 70) (p=0.00176, p=0.0034, respectively), and improved DFS and OS among patients whose tumors were ER positive (p=0.049, p=0.0034). Conclusions: In this single institution study of patients with greater than 10 positive axillary nodes, black patients had a significantly improved DFS compared with white patients. Young age and ER tumor positivity was associated with improved outcomes. Using multivariate analysis, there were no other variables associated with statistically significant improvements in DFS or OS including date of diagnosis. Further work is needed to improve breast cancer survival in this subgroup of patients.

e11575 Background: Axillary lymph node status is one of the most important prognostic factors in patients with breast cancer, and those with more than ten metastatic lymph nodes at diagnosis have a decreased overall survival. The purpose of this study is to better characterize the clinical course of this high-risk, poorly described patient population and determine the factors associated with long-term survival. Methods: A retrospective cohort analysis of all breast cancer patients with greater than ten metastatic lymph nodes diagnosed at Mt. Sinai Medical Center from January 1990 to December 2007 (n= 175). Descriptive frequencies, overall median survival (OMS), 5- and 10-year survival were calculated for standard prognostic factors and treatment variables. Univariate statistical analysis was performed, followed by a multivariate prognostic analysis for time-to-event data, using the Cox and extended Cox regression model. Results: The majority of patients were non-Hispanic white females between the ages of 56-70, diagnosed between 1990-1999, with tumors between 2-5 cm and 10-15 metastatic lymph nodes. Most were ER/PR positive, HER2 negative, and treated with surgery, chemotherapy, radiation and hormonal therapy. The OMS was 94 months (CI = 69-114) with 5- and 10-year survival rates of 64.3 and 41.6%, respectively. Ages between 21-45 years (OMS of 187 months, p = 0.03), tumors less than 2 cm (146 months (95% CI = 85-198), p = 0.02), ER positivity (131 months (95% CI = 94-157) vs. 39 months (95% CI = 27-59), p = 0.0003) and treatment received between 2000-2003 (98 months (95% CI = 55-133), p = 0.02) were all associated with significantly improved survival. Conclusions: Over the past decade there were significant gains in the long-term survival of breast cancer patients with greater than ten positive nodes at diagnosis, possibly due to improvements in multimodality therapy, such as the introduction of taxanes, although stage migration may be another contributing factor. Our study further showed an encouraging survival for ER positive patients and a dismal one for ER negative patients, highlighting the need for new targeted therapies directed towards ER negative tumors.

Purpose: The aim of the current study is to assess the impact of the glycolytic inhibitor 2-deoxy-D-glucose (2-DG) on tumor burden and hypoxia in the LHBETATAG retinal tumors. Methods: Group A: 17-week-old LHBETATAG transgenic mice (n=30) received periocular injections of saline and 2-DG (62.5, 125, 250, and 500mg/kg). Injections were given 2 times a week for 3 weeks. Group B: 4-week-old mice received oral delivery of 2-DG in custom made food pellets and were treated for either 8 or 18 weeks. 17-week-old mice received oral 2-DG for 8 weeks. At the time of enucleation, all eye samples were snap frozen and analyzed for tumor burden and hypoxic regions using immunohistochemistry. Results: Following injections of 2-DG, tumor control was not different between the control and the lowest two doses (62.5 and 125mg/kg). However, the difference in tumor burden was significant from 250mg/kg dose (p Conclusions: This study exhibits the efficacy of focal 2-DG as potential therapy to decrease both intratumoral hypoxia and tumor burden. The current study also shows that the non-invasive oral delivery 2-DG treatment has similar or better effects on the tumor depending on the treatment schedule used. In advanced disease of LHBETATAG retinal tumors, hypoxia is increasingly present. The use of glycolytic inhibitors as a therapeutic strategy has the potential to enhance current retinoblastoma treatments. 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 2050. doi:10.1158/1538-7445.AM2011-2050

Purpose: The purpose of the current study is to establish the distribution of tumor associated macrophages (TAMs) during tumor development in a transgenic retinoblastoma mouse model, study the contribution of bone-marrow derived macrophages in these tumors, and assess the supportive role of TAMs in retinoblastoma tumor growth. Methods: Macrophage infiltration in transgenic retinoblastoma tumors was assessed by immunohistochemistry at different time points in tumorigenesis. To establish the origin of TAMs in transgenic retinoblastoma tumors, 107 bone marrow cells from green fluorescent protein (GFP) positive 16-week-old mice were transplanted into age-matched, irradiated LHBETATAG mice via tail vein injections. Macrophage depletion was performed by subconjuctival (SC) delivery of liposomal clodronate. Results: TAMs’ density increased from 4- to 12-weeks of age in mice with small to medium-sized tumors (p=0.037), and remained stable in the later stages of the disease (i.e., 16-week-old with large tumors; p=0.20). 38% of TAMs (2.5 ± 3.2 cells per 400X hpf) in 16-week old mice were GFP-positive, bone marrow-derived macrophages. Total TAM depletion was correlated with a significant decrease in the expression levels of MMP-9 (p=0.014) and mature vessels (p Conclusions: The current study expands our understanding of the complex role that macrophages play in retinoblastoma. Macrophage modulation in the tumor microenvironment is a critical factor in retinoblastoma tumor development. 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 1517. doi:10.1158/1538-7445.AM2011-1517

Purpose: The purpose of the current study is to examine vessel targeting, chemotherapy, and mammalian target of rapamycin (mTOR) inhibitor agents in LHBETATAG retinal tumors and their impact on tumor burden. Methods: Group A: Ten-week-old, LHBETATAG mice (n=30) received a single subconjunctival injection of anecortave acetate (AA; 1200, 600, 300, and 150 µg) delivered to right eyes only. Group B: Ten-week-old, LHBETATAG mice (n=30) received a single subconjunctival injection of AA (600, 300, and 150 µg) delivered to right eyes only, either during a cycle of carboplatin (six subconjunctival deliveries) or after the completed cycle. Carboplatin was delivered at the subtherapeutic concentration of 62.5 µg. All animals were euthanatized at 16 weeks of age, and the eyes were examined histopathologically. Group C: Eighteen-week-old, LHBETATAG mice received (n=30) subconjunctival injections of rapamycin once weekly for two consecutive weeks (0.00333, 0.167, 3.33, and 6.67 mg/kg). Tumor sections were analyzed for tumor burden with immunohistochemistry techniques. Results: A statistically significant reduction in tumor burden was detected after a single periocular injection of AA. The reduction of tumor burden followed a U-shaped dose-response curve. Tumor burden was significantly decreased when AA and carboplatin were combined. However, varying doses and delivery schedule of these agents had significant impact on the effectiveness of the combined treatment. The most effective scheme was delivering a low dose (150-300 µg) of AA after a complete cycle of carboplatin. Reduction in tumor burden were significantly different between rapamycin doses and control (p Conclusions: AA, as monotherapy or as adjuvant therapy, significantly controlled tumor burden in a murine model of retinoblastoma. Moreover, adjuvant therapy enabled the use of typically subtherapeutic carboplatin doses without decreasing efficacy of the therapy. Inhibition of mTOR reduced tumor burden during late disease in the LHBETATAG retinoblastoma tumor model. Rapamycin may have a role in combination with chemotherapy or other adjuvant therapies to enhance retinoblastoma tumor control. 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 5352. doi:10.1158/1538-7445.AM2011-5352

Purpose: The aims of this study were to evaluate the hypothesis that LHBETATAG retinoblastoma tumors exhibit regional and temporal variations in gene expression and vascular development in retinoblastoma. Methods: To evaluate intratumoral gene expression, LHBETATAG mice aged 12, 16, and 20 weeks were euthanized (n=9). Specimens were taken from 5 tumor areas (apex, anterior lateral, center, base, and posterior lateral). Samples were hybridized with an Affymetrix GeneChip Mouse Gene ST 1.0 arrays. To examine the spatial distribution of new and mature blood vessels, immunohistochemical analyses were conducted on enucleated human (n=10) and LHBETATAG retinal tumors from 16-week-old mice (n=11). Results: There were significant temporal differences in gene expression for LHBETATAG retinoblastoma tumors (p 2.5 there were significant changes in gene expression for 190 genes apically, 84 genes anterolaterally, 126 genes posteriorly, 56 genes centrally, and 134 genes at the base. Differentially expressed genes were analyzed with significant involvement in angiogenesis (p Conclusions: There are significant temporal and regional variations in the gene expression and vasculature of retinoblastoma. Differentially expressed genes overlap with angiogenesis, hypoxia, and cellular metabolism. There is a heterogeneous vessel population in advanced retinoblastoma disease. These findings provide further understanding of the mechanisms involved in tumor progression, and emphasize that tumor biopsies cannot be used for prognostication secondary to tumor regionalization. Finally, optimally-timed adjuvant therapies that target critical components of identified pathways may potentially enhance retinoblastoma tumor control. 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 3865. doi:10.1158/1538-7445.AM2011-3865