Ectopic Tumor Research Articles

Surgical treatment for the patient with the neuroendocrine lung tumor associated with ectopic ACTH-secretion syndrome: case report

Among primary neoplasms of the lungs, the proportion of neuroendocrine tumors is up to 20–25%. According to the Surveillance Epidemiology and Results (SEER) database, since 1970 there has been a significant increase in primary carcinoid tumors of various organs with the highest incidence of morbidity in the broncho-pulmonary localization group. Carcinoid tumors of the lung level are approximately 2% of all primary lung tumors, but only 5% of these formations are associated with ACTH production.
 Despite the severity of clinical manifestations and the acceleration of hypercorticoid processes, the topical diagnosis of ACTH-ectopic syndrome often causes certain difficulties. The difficulty in diagnosing ectopic tumors is due to the fact that most of these formations are so small that they can be missed by standard methods of examination (X-ray, ultrasound, MSCT or MRI). After the diagnosis of ACTH-ectopic syndrome is completely optimal method of pathogenetic therapy is a radical removal of the tumor, which leads to the normalization of the functions of the adrenal cortex and the elimination of hypercorticoidism. However, the achievement of this goal in many cases is difficult due to late topical diagnosis, the prevalence of the tumor process and extensive metastasis.
 There is a clinical observation of a patient with a diagnosis of a neuroendocrine lung tumor with ACTH-ectopic syndrome. The importance of the multidisciplinary approach, the methods of differential diagnosis, the features of the perioperative period and the long-term results of treatment are described.

Abstract C47: Obesity enhances tumor colonization and promotes expression of the spliced variant MBD2_v2 and splicing factor SRSF2 in triple-negative breast cancer

Abstract Introduction: Triple-negative breast cancer (TNBC) is a molecular subtype that accounts for approximately 15-20% of invasive breast cancer diagnoses in the United States. TNBC occurs more prevalently in premenopausal African American women (AAW), and obesity coupled with chronic inflammation is a risk factor for both incidence and recurrence. TNBC is lacking therapeutic options and persons diagnosed with TNBC have the lowest 5-year survival rates among all breast cancer patients, with AAW disproportionally bearing this burden. In order to improve TNBC patient outcomes, etiologic factors driving disparities in disease progression need to be better understood. We previously showed in TNBC cell cultures that intrinsic oncogenic redox signaling promotes self-renewal capacity of cancer stem-like cells (CSC) by maintaining expression of an epigenetic reader protein and mRNA spliced variant, methyl binding domain protein 2 variant 2 (MBD2_v2). We hypothesized that obesity-induced chronic inflammation in vivo could promote a similar cancer phenotype. Methods: To study this potential phenomenon, we developed and characterized a diet-induced obesity mouse tumor xenograft (MTX) model of TNBC in C57BL/6J Rag1 (-/-) mice, a strain that maintains a functional innate immune system, to recapitulate the proinflammatory environment induced by increased adiposity. TNBC cell lines MDA-MB-468 and MDA-MB-231 were used to generate ectopic tumor xenografts in this model. Tumor colonization and growth were measured over 150 days. RT-PCR and microarray based assays were used to detect transcriptional changes in TNBC MTXs. Results: At 5 weeks post-diet consumption, mice fed a high-fat diet displayed a 5-10% increase in body weight and increased blood glucose concentration. By the end of the study high-fat fed mice displayed a 33-43% increase in body weight and appeared to have greater central adiposity post-mortem. In overweight mice relative to healthy weight controls, TNBC cell line tumor colonization rate and MBD2_v2 expression were significantly higher. Furthermore, staining for CSC marker EpCam was greater in overweight versus control MTXs, indicating that CSC self-renewal capacity could have been enhanced. Upon global gene expression analysis of overweight MTXs, we observed a significant upregulation of stem-related factors based on fold-change expression, as well as functional enrichment for RNA splicing, including SRSF2, a splicing factor that has been shown to drive MBD2_v2 expression in pluripotent stem cells. Knockdown of SRSF2 led to decreased expression of MBD2_v2 in MDA-MB-468 cells, and decreased CSC self-renewal capacity in vitro. Conclusions: Altogether these data support MBD2_v2 regulation by redox signaling, which may be driven by a proinflammatory environment in the presence of increased adiposity. Future studies will be conducted to confirm MBD2_v2's importance in obesity-related tumor colonization in in vivo overexpression and knockdown studies, and to determine its transcriptional regulatory functions in TNBC. Citation Format: Emily A. Girsch, Gregory Dyson, Cristina Mitrea, Bin Bao, Lisa Polin, Aliccia Bollig-Fischer. Obesity enhances tumor colonization and promotes expression of the spliced variant MBD2_v2 and splicing factor SRSF2 in triple-negative breast cancer [abstract]. In: Proceedings of the Tenth AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2017 Sep 25-28; Atlanta, GA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2018;27(7 Suppl):Abstract nr C47.

Abstract 1025: Clinical outcomes and differential tumor immune microenvironment in patients with small cell lung cancer and paraneoplastic syndromes

Abstract Background Among patients with small cell lung cancer (SCLC), approximately 20% develop a paraneoplastic syndrome (PNS). Neurologic PNS are immune-mediated phenomena predicated on host recognition of an onconeural antigen, while endocrinologic PNS are attributed to ectopic tumor secretion of normal hormones. The presence of neurologic PNS improves prognosis and endocrinologic PNS worsens prognosis in patients with SCLC. We hypothesized that tumors from patients with neurologic PNS may have increased TILs and PD-1/PD-L1 expression compared to tumors from patients with endocrinologic PNS and this improved immune recognition accounts for prognostic differences. Methods We searched electronic medical record text stored in the Vanderbilt University Medical Center (VUMC) clinical data warehouse to identify SCLC patients with and without a PNS. We obtained clinical information through manual, retrospective chart review using an IRB-approved protocol. Overall survival (OS) and progression free survival (PFS) were compared using a log rank test. Archived formalin fixed, paraffin embedded samples were obtained from the Vanderbilt University Pathology Tissue Repository. We performed multiplexed fluorescence immunohistochemistry combined with automated quantitative analysis (AQUA® Technoloy; Navigate BioPharma Services, Inc.) to assess PD-1, PD-L1, CD4, and CD8 expression. A PD-1/PD-L1 interaction score was calculated by measuring the total area of PD-1 positive cells within the proximity of PD-L1 positive cells. This area was then divided by the total area of all non-tumor nucleated cells in the image and multiplied by a factor of 10,000. CD4, CD8, and PD-1/PD-L1 interaction scores were compared using a two sample t-test or Wilcoxon Rank Sum test. Results A total of 145 SCLC patients were identified, 55 with a PNS (25 neurologic and 30 endocrinologic) and 90 control patients. Patients with neurologic PNS exhibited significantly improved OS and PFS compared to patients with endocrinologic PNS and control patients (median OS 24mo, 95% CI 16.4mo-not reached (NR), vs 12mo, 95% CI 8.3mo-15.5mo, vs 13mo, 95% CI 12.2mo-16mo; median PFS 14mo, 95% CI 9.3mo-NR vs 6mo, 95% CI 4.6mo-9.5mo, vs 7mo, 95% CI 6.6mo-8.1mo, respectively). Tumors from patients with neurologic PNS (n=9) had statistically significantly higher PD-1/PD-L1 interaction scores (p=0.02), and increased CD4 (p=0.01) and CD8 (p=0.003) T cell infiltrates compared to tumors from patients with endocrinologic PNS (n=11). Conclusion Our study of tumor tissue from patients with SCLC and PNS demonstrated a statistically significant increase in immune modulation markers' expression in patients with neurologic PNS. Tumor immunomodulation may be the driver of the improved prognosis that has been observed in ours and other retrospective cohorts of patients with SCLC and neurologic PNS. Citation Format: Wade T. Iams, Eileen Shiuan, Catherine B. Meador, Marc Roth, Jennifer Bordeaux, Christine Vaupel, Lucy L. Wang, Joseph T. Schneider, Jeremy L. Warner, Zhiguo Zhao, Christine M. Lovly. Clinical outcomes and differential tumor immune microenvironment in patients with small cell lung cancer and paraneoplastic syndromes [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 1025.

G‑protein‑coupled receptor120 regulates the development and progression of human esophageal cancer.

The aim of the present study was to investigate the role of G‑protein coupled receptor120 (GPR120) in esophageal cancer and explore the related mechanisms. The expression of GPR120 in esophageal cancer tissues was examined by immunohistochemistry. Correlation analysis was performed to investigate the association between the level of GPR120 and clinical parameters. The expression of GPR120 was evaluated in esophageal cancer cell lines and the effects of GPR120 on cell proliferation, clone formation, migration and invasion were evaluated in an invitro cell model and an invivo ectopic tumor nude mice model. In addition, the effect of GPR120 on epithelial‑mesenchymal transition (EMT), PI3K and I‑κB pathway, as well as angiogenesis and inflammation‑related cytokines was explored in order to elucidate the underlying mechanisms. Significantly increased expression of GPR120 was observed in esophageal cancer tissues compared to normal tissues. The expression of GPR120 was significantly related with histological grade, TNM stage and lymph node metastasis. GPR120 knockdown significantly decreased cell proliferation, clone formation, migration and invasion invitro and decreased tumor growth invivo. Furthermore significantly increased levels of E‑cadherin and decreased levels of N‑cadherin and vimentin, decreased level of Akt phosphorylation and I‑κB phosphorylation, as well as decreased levels of vascular endothelial growth factor (VEGF), interleukin‑8 (IL‑8) and cyclooxygenase‑2 (Cox‑2) and its corresponding protein PGE2 were observed as the underlying mechanisms. In conclusion, we observed an increased level of GPR120 in esophageal cancer tissues, which served as a positive regulator of the development and progression of esophageal cancer. Multiple mechanisms including EMT, PI3K and I‑κB pathway, as well as angiogenesis and inflammation‑related cytokines were involved.

Ultrasound Doppler as an Imaging Modality for Selection of Murine 4T1 Breast Tumors for Combination Radiofrequency Hyperthermia and Chemotherapy

Noninvasive radiofrequency-induced (RF) hyperthermia has been shown to increase the perfusion of chemotherapeutics and nanomaterials through cancer tissue in ectopic and orthotopic murine tumor models. Additionally, mild hyperthermia (37°C-45°C) has previously shown a synergistic anticancer effect when used with standard-of-care chemotherapeutics such as gemcitabine and Abraxane. However, RF hyperthermia treatment schedules remain unoptimized, and the mechanisms of action of hyperthermia and how they change when treating various tumor phenotypes are poorly understood. Therefore, pretreatment screening of tumor phenotypes to identify key tumors that are predicted to respond more favorably to hyperthermia will provide useful mechanistic data and may improve therapeutic outcomes. Herein, we identify key biophysical tumor characteristics in order to predict the outcome of combinational RF and chemotherapy treatment. We demonstrate that ultrasound imaging using Doppler mode can be utilized to predict the response of combinational RF and chemotherapeutic therapy in a murine 4T1 breast cancer model.

Effects of HPV Pseudotype Virus in Cutting E6 Gene Selectively in SiHa Cells.

The objectives of this study were to investigate the effects of the CRISPR/Cas9 system mediated by the HPV pseudotype virus on SiHa cytobiology behavior by cutting the HPV16 E6 gene selectively and to explore the role of this system in the treatment of cervical cancer. After designing specific gRNA sequences targeting HPV16 E6, generating hCas9-EGFP and E6-gRNA-RFP plasmids, and preparing the pseudovirus of HPV16 carrying E6-gRNA and Cas9 plasmids, we determined the titer of the pseudotype virus using the TCID50 method. We obtained the pseudotype virus of HPV16 carrying E6-gRNA and Cas9 plasmids to transfect cervical cancer SiHa cells. Experimental subjects were divided into control group, empty virus group, E6-gRNA transfected group, Cas9 transfected group and Cas9+E6-gRNA transfected group. The molecular size of the cutting sequence was detected using the T7E1 enzyme digestion method and agarose gel electrophoresis, and the cleavage function of CRISPR/Cas9 on the E6 gene was determined at the same time. RT-PCR and Western blotting were performed to detect the mRNA and protein expression levels of E6 in all the groups; the Transwell cell migration assay was performed to detect the cell migration ability and metastasis in all groups. Heterotopic transplantation tumors were incorporated into mice and were used to investigate the effects of the CRISPR/Cas9 system mediated by the HPV pseudovirus on the tumorigenic ability of SiHa cells by selectively cutting HPV16 E6. The HPV16 pseudotype virus carrying E6-gRNA and Cas9 plasmids could successfully infect SiHa cells, and there were two cutting zones in the Cas9+E6-gRNA transfected group. However, the empty virus group, E6-gRNA transfected group and Cas9 transfected group had no corresponding zone. Compared with those in the control group, the empty virus group, E6-gRNA transfected group and Cas9 transfected group, the mRNA and protein expression levels of E6 in SiHa cells were downregulated in the Cas9+E6-gRNA transfected group (P<0.01). In addition, the proliferation and migration abilities of SiHa cells were significantly inhibited (P<0.01). There were no significant differences among the other groups. In contrast to the control group, the HPV pseudotype virus carrying E6-gRNA and Cas9 plasmids could significantly delay the growth of tumor cells of the ectopic tumor transplantation model (P<0.01). The CRISPR/Cas9 system mediated by the HPV pseudotype virus to knockout E6 gene expression exhibited a clear inhibitory effect on the biological function of SiHa cells, which indicated that knocking out the E6 gene using the CRISPR/Cas9 system mediated by the HPV pseudotype virus had a potential effect of eliminating HPV infection and inhibiting the growth of HPV-related tumors. Taken together, these findings provide insight into a new treatment strategy for the prevention and treatment of hr-HPV infected disease, particularly in HPV-related tumors.

Spontaneous adrenocorticotropic hormone (ACTH) normalisation due to tumour regression induced by metyrapone in a patient with ectopic ACTH syndrome: case report and literature review

BackgroundEctopic adrenocorticotropic hormone (ACTH) syndrome (EAS) is caused by tumours releasing ACTH. Ectopic ACTH-producing tumour regression is rarely induced using steroidogenesis inhibitors. We presented a case of EAS in which ACTH production by a lung tumour was reduced by metyrapone (MTP) and also reviewed previous cases of ectopic ACTH production suppressed via steroidogenesis inhibition.Case presentationA 71-year-old female with general fatigue, central obesity and impaired glucose tolerance was diagnosed with Cushing’s syndrome due to elevated ACTH (192.9 pg/mL; normal range, 7.2–63.3 pg/mL), cortisol (73.1 μg/dL; 6.4–21.0 μg/dL) and 24-h urinary free cortisol (UFC) (6160 μg/day; 11.2–80.3 μg/day) levels. Chest computed tomography identified a solid 26.6 × 22.9 × 30.0 mm tumour with a cavity in the upper lobe of the left lung. There was no adrenal gland enlargement. Tumour markers were not significantly elevated; ACTH levels were not suppressed by 8-mg dexamethasone. A corticotropin-releasing hormone stimulation test revealed blunted ACTH response (basal ACTH, 204.6 pg/mL; highest ACTH level during the 120-min stimulation test, 214.0 pg/mL). She was diagnosed with EAS due to a lung lesion. MTP treatment was started to reduce cortisol production. ACTH levels and cortisol and UFC levels were normalised and the ACTH-producing lung tumour was ablated after MTP treatment. In several reported cases, plasma ACTH levels reduced during steroidogenesis inhibitor treatment for EAS. Among the 10 patients, three cases of pheochromocytoma, one of thymic carcinoid and one of islet cell carcinoma were reported. In four cases, the tumour was not detected. In our case, the pathology of the lung tumour was unknown because of lack of tumour cells in biopsy. The patients were treated with ketoconazole (KTZ) and/or MTP and exhibited ACTH and cortisol/UFC suppression, but tumour regression was observed only in our case.ConclusionMTP and/or KTZ may reduce ACTH and cortisol production. The tumour spontaneously regressed after MTP treatment, indicating that MTP may reduce the tumour size without surgery. The mechanisms of therapeutic effects of steroidogenesis inhibitors and prognosis of spontaneous remission should be elucidated further via molecular biology studies.

In the nose, not the sella: Case report of an ectopic pituitary adenoma

Abstract Background and importance Ectopic pituitary adenomas presenting in the nasal and paranasal sinuses are very rare. We report a patient with a large ectopic nonfunctioning pituitary macroadenoma involving the nasal passageways, ethmoid sinus, and extending through the cribriform plate extradurally to the anterior fossa with no involvement of the sella turcica. The patient presented with epistaxis and denied any visual disturbances or other associated clinical presentations that would demonstrate extensive cranial nerve involvement. Ectopic pituitary adenomas should be considered in the differential diagnosis of a nasal mass involving the nasal sinuses, even if there is no communication with the sella. Clinical presentation The patient is a 46-year-old male who presented with complete nasal obstruction. MRI revealed a large bilateral nasal mass extending into the anterior fossa through the cribriform plate. On nasal endoscopy, the patient was found to have a large mass occluding the entire nasal cavity. Subsequently, an endoscopic endonasal resection was performed. Conclusion This case represents a rare presentation of an ectopic pituitary adenoma. The extensive tumor involvement of the nasal passageways, ethmoid sinuses, and the anterior fossa with the absence of cranial nerve involvement are notable. Histopathologic confirmation of the ectopic pituitary tumor was made after complete endoscopic endonasal removal.

The effect of 2-methoxyestradiol liposome on growth inhibition, angiogenesis and expression of VEGF and Ki67 in mice bearing H22 hepatocellular carcinoma.

2-methoxyestradiol (2-ME), an endogenous metabolite of estrogen, has very low water solubility. It is currently in phase II clinical trials as both a chemopreventive and chemotherapeutic agent and has been orally administered to cancer patients. However, the poor oral absorption of the compound is one of the major obstacles for 2-ME development. Based on the molecular features of 2-ME, liposome can be considered an attractive formulation approach. Our purpose in this study is to research the antitumor efficacy of 2-methoxyestradiol liposome (2-ME-L) in mice bearing H 22 tumors. Murine H22 hepatocarcinoma served as an ectopic solid tumor model. The effects of antitumor therapy were evaluated by testing tumor growth, measuring the tumor inhibition rates in terms of weight and volume, and staining the tissues by hematoxylin and eosin. The synergistic mechanism of 2-ME-L therapy was elucidated by detecting changes in the expression of pathognostic factors in the tumor microenvironment. 2-ME-L significantly suppressed tumor growth. The morphological changes in the tumors indicated that the tumors in the treatment groups were effectively confined with little surrounding angiogenesis. Tumor cells of the treatment groups had abundant areas of necrosis with few nuclei in the mitotic phase. It was found that there was less immunohistochemical expression of vascular endothelial growth factor (VEGF), Ki67 and CD31 in the treatment groups and the efficacy of 2-ME-L was better than that of 2-ME solution (2-ME-S). This research demonstrated that 2-ME-L inhibited the growth of H 22 tumors in a concentration-dependent manner and was more effective than 2-ME-S. 2-ME-L can suppress the growth of H22 solid tumors and has antiproliferative, proapoptotic and antiangiogenic activity. 2-ME-L could be of potential use in the treatment of hepatocellular carcinoma.

Separate roles of IL-6 and oncostatin M in mouse macrophage polarization in vitro and in vivo.

Arginase-1 (Arg-1)-expressing M2-like macrophages are associated with Th2-skewed immune responses, allergic airway pathology, ectopic B16 melanoma cancer growth in murine models, and can be induced by Oncostatin M (OSM) transient overexpression in vivo. Here, we compare OSM to the gp130-cytokine IL-6 in mediating macrophage polarization, and find that IL-6 overexpression alone (Ad vector, AdIL-6) did not induce Arg-1 protein in mouse lungs at day 7, nor ectopic melanoma tumor growth at day 14, in contrast to overexpression of OSM (AdOSM). AdOSM elevated levels of IL-4, IL-5 and IL-13 in bronchoalveolar lavage fluid, whereas AdIL-6 did not. Bone marrow-derived macrophages respond with Arg-1 enzymatic activity to M2 stimuli (IL-4/IL-13), which was further elevated in combination with IL-6 stimulation; however, OSM or LIF had no detectable activity in vitro. Arg-1 mRNA expression induced by AdOSM was attenuated in IL-6-/- and STAT6-/- mice, suggesting requirements for both IL-6 and IL-4/IL-13 signaling in vivo. Ectopic B16 tumor burden was also reduced in IL-6-/- mice. Thus, OSM induces Arg-1+ macrophage accumulation indirectly through elevation of Th2 cytokines and IL-6 in vivo, whereas IL-6 acts directly on macrophages but requires a Th2 microenvironment, demonstrating distinct roles for OSM and IL-6 in M2 macrophage polarization.

The anti-cancer effect of E. coli in coordination with sorafenib

Objective To explore a new anticancer strategy by combination of E. coli and sorafenib in a rat Walker 256 tumor model. Methods Walker 256 cells (rat breast carcinoma cell line) were inoculated in liver of Wistar rats to establish an ectopic tumor model. Then intratumoral injection of E. coli 43013, which is tropic for tumor tissue other than normal tissue, was combined with or without sorafenib to treat the ectopic tumor. E. coli 43013 showed robust anticancer effects with the tumor volume decreased drastically and extensive necrosis and severe inflammation in peritumor tissue. Results E. coli 43013 in coordination with sorafenib have a better anticancer effects with smaller tumor volume and lower level of inflammation (P=0.015). Pathology analysis shows residual cancer absolutely disapears in coordination group when the E. coli group exist 40% of residual cancer rate. This therapy was safe according to blood cultures and gram stain assay. Immunostaining results showed that tumor cells underwent epithelial to mesenchymal transition (EMT) with epithelial marker cytokeratin (CK) upregulated and vascular endothelial growth factor (VEGF), Vimentin, matrix metalloproteinase (MMP)-9 and proliferating cell nuclear antigen (PCNA) downregulated by inflammation followed by E. coli 43013 intratumoral injection. Further research showed sorafenib could inhibit the cancer progression mediated by inflammatory associated pathway, which may be induced by necrotic debris of tumor cells, and tumor associated inflammatory cells. Conclusion Our results provide a potent new strategy by combination of microbe and multikinase inhibitor sorafenib for cancer therapy in clinical cancer management. Key words: E. coli; Sorafenib; Carcinoma, hepatocellular; Targeted therapy

Comparison of video-assisted thoracic surgery with open surgery in the treatment of ectopic mediastinal parathyroid tumors.

To investigate the therapeutic effect of different surgical treatment for ectopic mediastinal parathyroid tumors and pathological features. A total of 21 patients who were diagnosed with ectopic mediastinal parathyroid tumors and underwent surgeries in our department from May 1995 to May 2015 were collected and retrospectively analyzed. Twenty-one patients including 8 female (36.4%) and 13 male (63.6%) were collected. Among these patients, 9 cases were treated with video-assisted thoracic surgery (VATS), while 13 cases were treated with open surgery (including one secondary open operation after thoracoscopic operation). The average size of mediastinal tumors was 2.17±1.22 cm. For the post-operational pathology, 16 cases (76.2%) were diagnosed as ectopic parathyroid adenoma; 4 cases (19.0%) were diagnosed as parathyroid hyperplasia, while only 1 case (4.8%) was diagnosed as parathyroid adenocarcinoma. Parathyroid adenoma accounts for the major pathological type of ectopic mediastinal parathyroid tumors. In addition, the correct diagnosis with precise preoperative location was the key for the treatment of ectopic mediastinal parathyroid tumors accompanied with hyperparathyroidism. Surgical intervention was demonstrated to be an effective way for the treatment of ectopic mediastinal parathyroid tumors with satisfied therapeutic outcome, especially for the VATS due to its unique clinical advantages. However, there may some difficulties when locating ectopic mediastinal parathyroid tumor less than 1 cm and the operators should be very cautious when performing thoracoscopic operations.

MicroRNA-378 enhances radiation response in ectopic and orthotopic implantation models of glioblastoma.

Glioblastoma multiforme (GBM) is the most common and highly malignant primary brain tumor, which is virtually incurable due to its therapeutic resistance to radiation and chemotherapy. To develop novel therapeutic approaches for treatment of GBM, we examined the role of miR-378 on tumor growth, angiogenesis, and radiation response in ectopic and orthotopic U87 glioblastoma models. Cell and tumor growth rates, in vitro and in vivo radiation sensitivities, and tumor vascular density were evaluated in U87-GFP and U87-miR-378 tumor lines. Ectopic tumor response to radiation was evaluated under normal blood flow and clamp hypoxic conditions. Results show that in vitro, miR-378 expression moderately increased cell growth rate and plating efficiency, but did not alter radiation sensitivity. U87-miR-378 tumors exhibited a higher transplantation take rate than U87-GFP tumors. In vivo, under oxygenated condition, subcutaneous U87-miR-378 tumors receiving 25Gy showed a tendency for longer tumor growth delay (TGD) than control U87-GFP tumors. In contrast, under hypoxic condition, U87-miR-378 xenografts exhibited substantially shorter TGD than U87-GFP tumors, indicating that under normal blood flow conditions, U87-miR-378 tumors were substantially more oxygenated than U87-GFP tumors. Intracranial multi-photon laser-scanning microscopy demonstrated increased vascular density of U87-miR-378 versus control U87-GFP tumors. Finally, miR-378 increased TGD following 12Gy irradiation in U87 intracranial xenografts, and significantly prolonged survival of U87-miR-378 tumor-bearing mice (P = 0.04). In conclusion, higher miR-378 expression in U87-miR-378 cells promotes tumor growth, angiogenesis, radiation-induced TGD, and prolongs survival of orthotopic tumor-bearing hosts. Regulation of VEGFR2 by miR-378 significantly increased vascular density and oxygenation in U87 xenografts.

Chemoradiation Combined with Phosphatidylserine-Targeting Antibody Enhances Systemic Anti-tumor Immune Responses

Recent advances in cancer immunotherapy have sparked significant interest in harnessing the body’s immune system to fight cancer. However, the response rates of cancer immunotherapies, including immune checkpoint blockade, remain low. Strategies to enhance anti-tumor immune responses by targeting different steps along the immune activation cascade that can complement T cell-based immunotherapies are clearly needed. Here, we examine whether the addition of an antibody that targets extracellular phosphatidylserine (PS), a molecule that is recognized by myeloid derived cells, can enhance anti-tumor immune responses of chemoradiation for non-small cell lung cancer (NSCLC). Chemoradiation (CRT) was combined with a murine monoclonal antibody (mch1N11). PS is highly expressed in both orthotopically and ectopically implanted 393P murine NSCLC models. Radiation dose was 2 Gy/day, given for 5 days. Chemotherapy consists of carboplatin and paclitaxel at a dose of 30 mg/kg and 10 mg/kg, respectively. PS-targeting antibody (mch1N11) was given at 3mg/kg for 2 weekly doses. For ectopic tumor models, bilateral tumors were established in the legs. Radiation was directed to the R leg and tumor on the L side was shielded. Tumor growth was measured either with CT imaging or digital caliper. Tumor infiltrating immune cell profiles were analyzed using immunohistochemistry. Survival analyses were performed using Kaplan-Meier method and compared using Log-rank test. CRT + mch1N11 treatment significantly inhibited growth and improved survival in mice implanted with orthotopic 393P tumors as compared to CRT alone (median: 21 vs. 15 days, HR: 2.77, P = 0.006). Tissue analyses showed that CRT significantly increased the expression of PD-L1 within the tumor and drastically reduced the number of tumor infiltrating CD8 T cells. For the CRT + mch1N11 group, a similar up-regulation of PD-L1 expression was noted. However, the addition of mch1N11 re-populated the infiltrating CD8 T cells within the tumor. In the bilateral tumor model, the addition of mch1N11 antibody to CRT resulted in tumor regression in ∼40% of the non-irradiated tumors in the contralateral side as compared to 0% in the CRT alone or chemotherapy + mch1N11 groups. We showed that PS-targeting combined with standard CRT can significantly prolong survival in preclinical models of NSCLC and generate enhanced systemic anti-tumor immunity against lesions outside of the irradiated field. CRT up-regulated PD-L1 expression within the tumor and depleted tumor infiltrating cytotoxic CD8 T cells. The addition of mch1N11 antibody to CRT, however, was able to re-populate this critical immune effector cell population. Together, our results demonstrate that PS-targeting antibodies may be combined with CRT to enhance intrinsic tumor immunogenicity, activate systemic anti-tumor immune responses, and act as a priming strategy to sensitize the tumor to immune checkpoint inhibition with PD-1 or PD-L1 antibodies.

Effects of tumor microenvironments on targeted delivery of glycol chitosan nanoparticles

In cancer theranostics, the main strategy of nanoparticle-based targeted delivery system has been understood by enhanced permeability and retention (EPR) effect of macromolecules. Studies on diverse nanoparticles provide a better understanding of different EPR effects depending on their structure, physicochemical properties, and chemical modifications. Recently the tumor microenvironment has been considered as another important factor for determining tumor-targeted delivery of nanoparticles, but the correlation between EPR effects and tumor microenvironment has not yet been fully elucidated. Herein, ectopic subcutaneous tumor models presenting different tumor microenvironments were established by inoculation of SCC7, U87, HT29, PC3, and A549 cancer cell lines into athymic nude mice, respectively. In the five different types of tumor-bearing mice, tumor-targeted delivery of self-assembled glycol chitosan nanoparticles (CNPs) were comparatively evaluated to identify the correlation between the tumor microenvironments and targeted delivery of CNPs. As a result, neovascularization and extents of intratumoral extracellular matrix (ECM) were both important in determining the tumor targeted delivery of CNPs. The EPR effect was maximized in the tumors which include large extent of angiogenic blood vessels and low intratumoral ECM content. This comprehensive study provides substantial evidence that the EPR effects based tumor-targeted delivery of nanoparticles can be different depending on the tumor microenvironment in individual tumors. To overcome current limitations in clinical nanomedicine, the tumor microenvironment of the patients and EPR effects in clinical tumors should also be carefully studied.

Oxygen generating nanoparticles for improved photodynamic therapy of hypoxic tumours

Photodynamic therapy (PDT) is a clinically approved anti-cancer treatment that involves the activation of an otherwise inactive sensitiser drug with light, which in the presence of molecular oxygen, generates cytotoxic reactive oxygen species (ROS). As oxygen is a key requirement for the generation of ROS in PDT and given the fact that hypoxia is a characteristic of most solid cancerous tumours, treating hypoxic tumours using PDT can be a challenge. In this manuscript, we have prepared a CaO2 nanoparticle (NP) formulation coated with a pH-sensitive polymer to enable the controlled generation of molecular oxygen as a function of pH. The polymer coat was designed to protect the particles from decomposition while in circulation but enable their activation at lower pH values in hypoxic regions of solid tumours. The oxygen generating capability of the polymer coated NPs was demonstrated in aqueous solution with minimal oxygen produced at pH7.4, whereas it increased significantly when the pH was reduced to 6.2. The polymer coated CaO2 NPs were also observed to significantly increase tumour pO2 levels (p<0.05) in mice bearing ectopic human xenograft MIA PaCa-2 pancreatic tumours with an average increase in tumour pO2 of 6.5mmHg in the period 10–30min following administration. A statistically significant improvement in PDT mediated efficacy (p<0.001) was also observed when the particles were administered to mice bearing the same tumours 20min prior to PDT treatment. These results suggest that the polymer coated CaO2 NP formulation offers significant potential as an in situ method for oxygen generation to enhance the efficacy of treatments that depend on the presence of oxygen to elicit a cytotoxic effect.

Catching the Zebra: Clinical Pearls and Pitfalls for the Successful Diagnosis of Zollinger-Ellison Syndrome.

Zollinger-Ellison syndrome (ZES) results from an ectopic gastrin-secreting tumor leading to peptic ulcer disease, reflux, and chronic diarrhea. While early recognition portends an excellent prognosis with >80% survival at 15years, symptoms are often nonspecific making the diagnosis difficult to establish. Diagnosis involves a series of tests, including fasting gastrin, gastric pH, chromogranin A, and secretin stimulation. Performing these tests in the correct sequence and at the proper time is essential to avoid inaccurate results. Tumor localization is equally nuanced. Although providers have classically used 111indium-radiolabeled octreotide with somatostatin receptor scintigraphy to evaluate tumor size and metastases, recent studies have shown superior results with newer imaging modalities. In particular, 68gallium (68Ga)-labeled somatostatin radiotracers (i.e., 68Ga-DOTATOC, 68Ga-DOTANOC and 68Ga-DOTATATE) used with positron emission tomography/computed tomography can provide excellent results. Endoscopic ultrasound is another useful modality, particularly in patients with ZES in the setting of multiple endocrine neoplasia type 1. This review aims to provide clinicians with an overview of ZES with a focus on both clinical presentation and the proper utilization of the various biochemical and imaging tests available.

Platelet microparticles infiltrating solid tumors transfer miRNAs that suppress tumor growth

AbstractPlatelet-derived microparticles (PMPs) are associated with enhancement of metastasis and poor cancer outcomes. Circulating PMPs transfer platelet microRNAs (miRNAs) to vascular cells. Solid tumor vasculature is highly permeable, allowing the possibility of PMP–tumor cell interaction. Here, we show that PMPs infiltrate solid tumors in humans and mice and transfer platelet-derived RNA, including miRNAs, to tumor cells in vivo and in vitro, resulting in tumor cell apoptosis. MiR-24 was a major species in this transfer. PMP transfusion inhibited growth of both lung and colon carcinoma ectopic tumors, whereas blockade of miR-24 in tumor cells accelerated tumor growth in vivo, and prevented tumor growth inhibition by PMPs. Conversely, Par4-deleted mice, which had reduced circulating microparticles (MPs), supported accelerated tumor growth which was halted by PMP transfusion. PMP targeting was associated with tumor cell apoptosis in vivo. We identified direct RNA targets of platelet-derived miR-24 in tumor cells, which included mitochondrial mt-Nd2, and Snora75, a noncoding small nucleolar RNA. These RNAs were suppressed in PMP-treated tumor cells, resulting in mitochondrial dysfunction and growth inhibition, in an miR-24–dependent manner. Thus, platelet-derived miRNAs transfer in vivo to tumor cells in solid tumors via infiltrating MPs, regulate tumor cell gene expression, and modulate tumor progression. These findings provide novel insight into mechanisms of horizontal RNA transfer and add multiple layers to the regulatory roles of miRNAs and PMPs in tumor progression. Plasma MP-mediated transfer of regulatory RNAs and modulation of gene expression may be a common feature with important outcomes in contexts of enhanced vascular permeability.

Abstract 5138: Development of a gold nanoparticle platform capable of selective homing to tumor tissues for intravenous antitumor therapy

Abstract Intratumoral administration of high mobility group nucleosome-binding protein 1 (HMGN1) and R848/resiquimod together with a checkpoint inhibitor can eradicate large established tumors. However, the intratumoral route of administration is difficult to use on cancers of internal organs. To improve potential clinical use of HMGN1 and R848, it is necessary to deliver HMGN1 and R848 systemically to the tumor tissue. Gold nanoparticles (AuNP) have been extensively studied as a vehicle for various types of drugs, including anticancer compounds. In this study, we established a means of generating PEGylated Au-HMGN1-R848 (Au-PEG-HMGN1-R848) complexes, which, upon intravenous IV administration, preferentially target ectopic hepatoma or colon tumor tissue and avoids uptake of the particles by liver and spleen of tumor-bearing mice. Additionally, Au-PEG-HMGN1-R848 complexes were stable and preserved the capacity of HMGN1 and R848 to stimulate the maturation of dendritic cells. IV administration of Au-PEG-HMGN1-R848 complexes plus a checkpoint inhibitor had effective antitumor activity in curing 80% of C57BL/6 mice bearing large (1 cm in diameter) Hepa1-6 hepatoma and Balb/c mice bearing CT26 colon tumors. Therefore, an Au nanoparticle platform capable of selectively delivering HMGN1 and R848 systemically to tumor tissues can be used to effectively to cure ectopic Hepa1-6 hepatoma and CT26 colon carcinoma. Note: This abstract was not presented at the meeting. Citation Format: Zhen Han. Development of a gold nanoparticle platform capable of selective homing to tumor tissues for intravenous antitumor therapy [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 5138. doi:10.1158/1538-7445.AM2017-5138

Abstract 1867: Near infrared fluorescent imaging of brain tumor with IR780 dye incorporated phospholipid nanoparticles

Abstract Background: Near-IR fluorescence (NIRF) imaging is becoming a promising approach in preclinical tumor detection and clinical image-guided oncological surgery. While heptamethine cyanine dye IR780 has excellent tumor targeting and imaging potential, its hydrophobic properties limit its clinical use. In this study, we developed nanoparticle formulations to facilitate the use of IR780 for fluorescent imaging of malignant brain tumor. Methods: Self-assembled IR780-liposomes and IR780-phospholipid micelles were prepared and their NIRF properties were characterized. The intracellular accumulation of IR780-nanoparticles in glioma cells were determined using confocal microscopy. The in vivo brain tumor targeting and NIRF imaging capacity of IR780-nanoparticles were evaluated using U87MG glioma ectopic and orthotopic xenograft models and a spontaneous glioma mouse model driven by RAS/RTK activation. Results: The loading of IR780 into liposomes or phospholipid micelles was efficient. The particle diameter of IR780-liposomes and IR780-phospholipid micelles were 95 nm and 26 nm, respectively. While stock solutions of each preparation were maintained at ready-to-use condition, the IR780-phospholipid micelles were more stable. In tissue culture cells, IR780-nanoparticles prepared by either method accumulated in mitochondria, however, in animals the IR780-phospholipid micelles showed enhanced intra-tumoral accumulation in U87MG ectopic tumors. Moreover, IR780-phospholipid micelles also showed preferred intracranial tumor accumulation and potent NIRF signal intensity in glioma orthotopic models at a real-time, non-invasive manner. Conclusion: The IR780-phospholipid micelles demonstrated tumor-specific NIRF imaging capacity in glioma preclinical mouse models, providing great potential for clinical imaging and image-guided surgery of brain tumors. Citation Format: Shihong Li, Jennifer Johnson, Anderson Peck, Qian Xie. Near infrared fluorescent imaging of brain tumor with IR780 dye incorporated phospholipid nanoparticles [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 1867. doi:10.1158/1538-7445.AM2017-1867