bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 4300: Constitutive activation of Stat3 increases hair follicle progenitor population at the expense of bulge region keratinocyte stem cells
Authors: Rao Dharanija , Carbajal Steve , Kiguchi Kaoru , DiGiovanni John
Published: February 2011

Mouse skin epidermis is maintained throughout life by a highly proliferative compartment located in the outer root sheath called the hair follicle bulge. Stem cells located in the mouse hair follicle bulge region have self-renewing, slow cycling and quiescent characteristics and maintain hair and epidermal growth. In the present study, we examined the impact of Stat3 activation on the number and proliferative capacity of bulge region keratinocyte stem cells (KSCs). K5. Stat3C transgenic mice express a constitutively active/dimerized form of Stat3 called Stat3C via the bovine keratin 5 (K5) promoter. The K5 promoter targets expression of Stat3C in the basal layer of the epidermis as well as in the bulge region KSCs. We analyzed the number of KSCs using the bulge specific stem cell markers CD34 and α6 integrin. The number of CD34+ve/α6 +ve cells was significantly reduced in Stat3C mice as compared to the non-transgenic littermates. Sca-1 positive cells reside in the infundibulum region of the hair follicle. We analyzed the Sca-1 expressing population in the total hair follicle cells and found that the number of Sca-1 expressing cells was increased in the Stat3C mice. Activation of Stat3, in bulge region KSCs appeared to cause epidermal stem cells to exit the stem cell compartment. To understand how Stat3 exerts these effects we studied the expression of Stat3 target genes like Myc and cyclin D1. Both Myc and cyclin D1 were found to be upregulated in total hair follicle cells in the transgenic mice. In addition, the levels of both α6 and β1 integrin were reduced in total hair follicle cells of the Stat3C mice supporting the hypothesis that activation of Stat3 causes stem cell to exit from the bulge region. Overall, these studies suggest that Stat3 is critical for the behavior, growth properties and quiescent characteristics of bulge region KSCs. Supported by NIH Grant CA76520.

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 4300.
Abstract 4301: Isolation and expansion of human gastric cancer-initiating cells for drug development
Authors: Liu Yan , Lv Junjie , Ning Jinying
Published: February 2011

Gastric carcinoma is the second most common cause of death from cancer in the world. The isolation and characterization of tumorigenic gastric cancer cells may help to develop novel diagnostic and therapeutic methods. Although there is increasing evidence that cancer stem cell, which is responsible for tumor formation and maintenance, exists in a wide variety of solid tumors, this has not been extensively explored in gastric cancer. We report here the isolation and in vitro propagation of gastric cancer-initiating cells from human primary tumor xenografts. The isolated tumor cells can form spheroid colonies in defined serum free medium. Subcutaneous injection of as few as 100 sphere-forming cells readily reproduced the original tumor in immunodeficient mice. Such tumors were serially transplanted for several generations and the cultured cells grew exponentially for more than 40 passages in vitro as undifferentiated tumor spheres. Interestingly, CD44, recently reported as a gastric cancer stem cell surface marker, was positive in both cultured cells and in engrafted tumor tissues. In vitro and in vivo drug sensitivity assays on these sphere-forming cells are currently ongoing.

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 4301.
Abstract 4308: Dynamic regulation of CD24 and the invasive, CD44posCD24neg phenotype in breast cancer cell lines
Authors: Meyer Matthew J. , Fleming Jodie M. , Ali Mustapha A. , Pesesky Mitchell W. , Ginsburg Erika , Vonderhaar Barbara K.
Published: February 2011

Introduction

The invasive, mesenchymal phenotype of CD44posCD24neg breast cancer cells has made them a promising target for eliminating the metastatic capacity of primary tumors. It has been previously demonstrated that CD44neg/lowCD24pos breast cancer cells lack the ability to give rise to their invasive CD44posCD24neg counterpart. Here we demonstrate that noninvasive, epithelial-like CD44posCD24pos cells readily give rise to invasive, mesenchymal CD44posCD24neg progeny in vivo and in vitro. This interconversion was found to be dependent upon Activin/Nodal signaling.

Methods

Breast cancer cell lines were sorted into CD44posCD24pos and CD44posCD24neg populations to evaluate their progeny for the expression of CD44, CD24, and markers of a mesenchymal phenotype. The FACS sorted populations were injected into immunocompromised mice to evaluate their tumorigenicity and invasiveness of the resulting xenografts.

Results

CD24 expression was dynamically regulated in vitro in all evaluated breast cancer cell lines. Furthermore, a single noninvasive, epithelial-like CD44posCD24pos cell had the ability to give rise to invasive, mesenchymal CD44posCD24neg progeny. Importantly, this interconversion occurred in vivo as CD44posCD24pos cells gave rise to xenografts with locally invasive borders as seen in xenografts initiated with CD44posCD24neg cells. Lastly, the ability of CD44posCD24pos cells to give rise to mesenchymal progeny, and vice versa, was blocked upon ablation of Activin/Nodal signaling.

Conclusions

Our data demonstrate that the invasive, mesenchymal CD44posCD24neg phenotype is under dynamic control in breast cancer cell lines both in vivo and in vitro. Furthermore, our observations suggest that targeted therapy against CD44posCD24neg tumor cells may have limited success in preventing primary tumor metastasis unless Activin/Nodal signaling is arrested.

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 4308.
Abstract 431: Cigarette-smoke-induced expression of MUC4 in human pancreatic cancer cells: Potential pathological roles
Authors: Momi Navneet , Ponnusamy Moorthy Palanimuthu , Batra Surinder K.
Published: February 2011

Pancreatic cancer is a major problem worldwide and the fourth leading cause of cancer-related deaths in the United States. In the pancreatic malignancy, long term smoking is the most established risk factor. Cigarette smoke contains a variety of constituents of which nicotine is the major component present. Previously, we have shown that MUC4, a member of transmembrane mucin family, is aberrantly expressed in the pancreatic tumors compared to the normal pancreas and is functionally involved in the tumorigenecity and metastasis of pancreatic cancer. The aim of the study was to investigate the role of nicotine on the pathogenesis of pancreatic cancer and to check if MUC4 is the major regulator involved in the process. The western blot and confocal analysis showed a gradual increase in the expression of MUC4 with nicotine treatment of the cells in a dose-dependent manner. Further, it was seen that retinoic acid (RA), a known MUC4 regulator, plays a synergistic role along with the nicotine, in up-regulating MUC4 expression in CD18/HPAF-SF. Further, studies were performed in order to elucidate the possible regulatory mechanism behind the nicotine mediated MUC4 induction and it's downstream signaling. RT PCR and western blot analysis showed an over expression of the nicotine receptor subunit α7 in nicotine-treated CD-18/HPAF as compared to the untreated cells. In addition, the study revealed that the α7 nicotine receptor subunit mediated JAK2-STAT3 pathway and the FAK activation, are the main regulatory mechanisms involved. Furthermore, the role of the α7 nicotine receptor subunit was confirmed by treating the cells with an α7 receptor antagonist, DhβE, which reduced the MUC4 expression and the JAK2 mediated activation of STAT3. In addition, in vitro functional assays were performed to assess the effect of nicotine induced MUC4 on CD-18/HPAF cells. Interestingly, the wound healing assay showed an increase in the migration potential of nicotine treated CD-18/HPAF cells as compared to the untreated cells. The western blot analysis provides clear evidence that nicotine also facilitate the phenomenon of EMT in CD-18/HPAF cells as shown by the down-regulation of the epithleial marker, E-cadherin, and an up-regulation of the mesenchymal marker, N-cadherin. Overall, our study shows a malignant potential of nicotine in the pancreatic cancer pathogenesis through α7 receptor subunit mediated MUC4 induction, thereby, increasing the migration potential of pancreatic cancer cells.

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 431.
Abstract 4311: Identification of a subset of peripheral blood cells expressing ALDH1 with chromosomal abnormalities in lung cancer patients suggests these are putative cancer stem cells
Authors: Khanna Abha , Zaidi Tanweer M. , He Weigong , Fernandez Ricardo , Zhang Hua-Zhong , Caraway Nancy P. , Mehran Reza , El-Zein Randa , Blowers David P. , Jiang Feng , Katz Ruth L.
Published: February 2011

The concept that cancers originate from cytogenetically abnormal stem cells (SC) is becoming accepted. Mechanisms by which cancer stem cells (CSC) contribute to tumor initiation and progression are unknown, however CSCs are resistant to chemotherapy and have metastatic potential. ALDH1 is a marker for identifying SCs. Increased ALDH1 expression in lung cancer (LC) was shown to correlate with disease stage and poor survival. Recently we showed that cytogenetically abnormal cells (CACs) similar to those in the primary tumor are present in the peripheral blood (PB) of patients with LC and numbers of CACs correlated with disease stage. We wished to discover if there were PB cells that expressed ALDH1 and showed chromosomal abnormalities (CA), representing CSCs, and if their presence correlated with tumor subtype and survival.

Materials and Methods: Prior to resection of lung cancers, peripheral blood mononuclear cells (PBMNCs) were isolated from 5 controls and 11 cases of LC. Patients ranged in age from 41-83 years. There were 4 low stage (I-II) and 7 high stage (III-IV) patients. Patients were followed prospectively over a 20 month period. Immunostaining for ALDH1 (BD Pharmingen) was performed, following which FISH for CEP 3, CEP 7, CEP 17 and LSI 9p21 probe set (Abbott Molecular) was performed on relocated ALDH1+ cells. FISH signals were scored for any chromosomal abnormality; deletion or gain.

Results: A low percentage of ALDH1+ cells were present in PB of 7/7 adenocarcinoma (ADC), 2/2 non-small cell lung carcinoma (NSCLC) 1/2 squamous carcinoma (SQC) patients and in controls. In LC, 53 ALDH1+ cells were detected of which 18 (34%) showed CAs. Controls were negative for CAs.

Conclusions: The numbers of ALDH1+ cells with CA tended to increase with stage. Genetically abnormal ALDH1+ cells may represent CSCs and provide a therapeutic target for efficient treatment of lung cancer.Demographic data and chromosomal abnormalities in ALDH1 + cells# casesStageHistologic SubtypeMean/Range % ALDH1+ cellsMean/Range % ALDH1+ cells with chromosomal abnormalities# of Chromosomal Abnormalities in ALDH1+ cellsFollow up in monthsVitals5Controls1 (1-2)00N/AN/A 7I-II2 SQC 2 ADC2.57 (0-9)0.71 (0-3)5 Monosomy 17 (3) Trisomy 9p21 (2)17-207 alive7III-IV2 NSCLC 5 ADC5 (2-12)1.86 (0-4)13, Monosomy 17 (4), gain 9p21 (3) Monosomy 3 (2) trisomy 3, trisomy 17, monosomy 7, monosomy 9p211-202 alive 5 dead

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 4311.
Abstract 4316: A novel wide field-of-view imaging device for real-time, intra-operative tumor bed assessment
Authors: Mito Jeffrey K. , Ferrer Jorge M. , Brigman Brian E. , Lee Chang-Lung , Dodd Rebecca D. , Eward William C. , Marshall Lisa F. , Dankort David , McMahon Martin , Kim Yongbaek , Lee W. David , Griffith Linda G. , Bawendi Moungi G. , Kirsch David G.
Published: February 2011

Limb-sparing surgery for extremity soft tissue sarcoma removes all cancer cells at the primary tumor site in the majority of patients. However, without radiation therapy, microscopic residual sarcoma cells left behind in the tumor bed will cause a tumor recurrence in approximately one-third of patients. Therefore, adjuvant radiation therapy is delivered to most patients, even when the tumor bed lacks residual cancer. Here, we present an imaging system for residual cancer assessment, consisting of a novel wide field-of-view imaging device and a protease-activated fluorescent probe. We demonstrate that this system directly images microscopic residual sarcoma cells in the tumor bed of mice when primary soft tissue sarcomas are resected. Moreover, this system can detect single tumor cells that have activated the fluorescent probe in vivo. This technology has the potential to be used as an intra-operative tool to identify microscopic residual disease for soft tissue sarcoma and other cancers with the goal of reducing rates of local recurrence, re-operation for positive margins, and adjuvant radiation therapy to tumor beds that lack residual cancer.

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 4316.
Abstract 4322: Ligand-directed therapy and molecular imaging in a mouse model of androgen-independent osteogenic prostate cancer
Authors: Driessen Wouter HP , Mandelin Jami , Cardó-Vila Marina , Kim Jaesung , Mathew Paul , Navone Nora M. , Soghomonyan Suren , Flores Leo G. , Alauddin Mian M. , Gelovani Juri G. , Pasqualini Renata , Arap Wadih
Published: February 2011

In working towards a theranostic approach in human prostate cancer, we have used in vivo phage display in a unique animal model of androgen-independent human osteoblastic bone metastases (animal model described by Li et al. J. Clin. Invest. 2008 Aug;118(8):2697-710) to uncover accessible targets and peptide ligands for molecular imaging and treatment. This unbiased combinatorial screening identified a peptide targeting Glucose-regulated Protein 78kDa (GRP78), a protein previously implicated in human cancer, including prostate cancer.

By using a hybrid gene delivery vehicle consisting of phage capsid-proteins and AAV genomic elements (AAVP) we have successfully expressed thymidine kinase protein from Herpes Simplex Virus (HSVtk) for a gene-directed enzyme prodrug therapy (GDEPT) approach. In combination with 18FEAU as an imaging probe, the expression of this protein was monitored by Positron Emission Tomography (PET). After expression was confirmed, treatment with the prodrug gancyclovir was used to induce tumor-specific apoptosis. In a second approach, this ligand-receptor system was used to specifically and efficiently deliver small interfering RNA (siRNA) in vitro and in vivo. Both approaches clearly show that the newly identified peptide targeting GRP78 can be considered for targeted drug development against human metastatic androgen-independent osteogenic prostate cancer.

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 4322.
Abstract 4327: Development of pancreatic cancer specific targeting agents as imaging agents
Authors: Maeda Yoshiko , Xiao Wenwu , Liu Ruiwu , Uriyama Shiro , Maitra Anirban , Lam Kit , Kumaresan Pappanaicken R.
Published: February 2011

In the “one bead one compound” combinatorial library method, millions of In the “one bead one compound” combinatorial library method, millions of random peptides can be generated rapidly, such that each bead displays only one peptide entity. When live cancer cells are co-cultured with these peptide-bead libraries, beads with cell adhesion peptides will be covered entirely by the cells. Low passage human pancreatic cancer (PDAC cells;XPA1, XPA3) and immortalized normal pancreas cells were used for this study. Positive peptide-beads were then isolated for sequence analysis. Using this high throughput screening approach, we evaluated four different peptide libraries and identified several peptides that bind to pancreatic cancer cells. To test their binding specificity, these peptides were labeled with fluorophor or biotinylated, and used as reagents for in vivo imaging. Athymic nude mice were used to induce pancreatic xenografts and very low uptake in liver. One of the cyclic peptide (PCa-pep6) targets pancreatic xenografts selectively. Based on the antibody blocking assay, PCa-pep6 binds to alpha-2 and alpha-6 integrins which are over expressed in pancreatic cancer. Therefore it may potentially be used as lead compounds for the development of high affinity and high specificity therapeutic and imaging agents for pancreatic cancers.

This work is supported by grant RO3-CA129786

Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend.

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 4327.
Abstract 4332: Ultra-low background optical sentinel lymph node imaging using two unique nano-particles
Authors: Kosaka Nobuyuki , Mitsunaga Makoto , Ogawa Mikako , Choyke Peter L. , Kobayashi Hisataka
Published: February 2011

Word limit: 2600 characters, including abstract body and title.

Optical imaging has the potential to become a robust tool in intraoperative sentinel lymph node mapping because of its portability, high spatial resolution, and absence of radiation. Most preclinical and clinical studies have used conventional fluorescent agents, which need excitation light to emit fluorescence. However, excitation light also induces autofluorescence (background signal) from intrinsic fluorophores in the normal tissue, lowering the target-to-background ratio. We developed two new methods for optical sentinel lymph node mapping to eliminate background signals using two unique nano-particles; an upconverting nano-particle (UCNP) and a self-illuminating bioluminescence resonance energy transfer quantum-dot (BRET-QD) nano-particle. UCNPs are unique nano-sized particles that emit light at shorter wavelengths with excitation in the near infrared (NIR). Since fluorophores typically emit light at longer wavelengths than the excitation light, there is no autofluorescence. Moreover, by changing the composition of the doping metals, UCNPs can be designed to emit at specific wavelengths. When UCNPs were injected into the chin of anesthetized mice, and excited at 950-nm, superficial neck lymph nodes were successfully depicted with no background signals in the visible-range (peak 550 nm) or NIR-range (peak 800 nm) in all mice. Furthermore, with serial injection of the visible and NIR-range UCNPs, simultaneous lymphatic imaging was achieved in two colors. The BRET-QD is self-illuminating and employs the substrate, coelenterazine, which reacts with luciferase conjugated on the BRET-QD. The bioluminescent light in blue efficiently excites the NIR QD, and emits photons in the NIR, therefore, there is no autofluorophore excitation. Moreover, by changing the QDs within BRET-QD, it is possible to alter their emission wavelength. BRET-QD655, (QD655 in its core), was injected at different sites (chin, ear, forepaws, and hind paws) of anesthetized mice. After BRET-QD655 injection, coelenterazine was intravenously injected and imaging was performed without excitation light. In all mice, sentinel lymph nodes in each lymphatic basin were clearly visualized with ultra-low background signals, and lasted at least 30 min after coelenterazine injections. In conclusion, we demonstrate that optical imaging using two unique nano-particles were able to depict sentinel lymph nodes with minimal autofluorescence. This enables the direct, real time sentinel lymph node mapping without extensive post processing of the images. These two novel nano-particles have the potential to be robust tools for sentinel lymph node detection.

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 4332.
Abstract 4333: Optical diagnostic system with Raman spectroscopy for gastric cancer
Authors: Konno Hiroyuki , Kawabata Toshiki , Shigetoshi Okazaki , Ichirota Iino , Uehara Takeshi , Setoguchi Tomohiko , Kikuchi Hirotoshi , Yamamoto Masayoshi , Ohta Manabu , Kamiya Kinji
Published: February 2011

Background. Raman spectroscopy (RAS) is one of the available optical techniques, and the Raman spectrum for each molecule and tissue is characteristic and specific. We reported the usefulness of a Raman spectrum as a novel diagnostic modality for gastric cancer by using the biopsied specimens on 100th Annual Meeting. We present here the advanced data of RAS by using resected specimen of gastric cancers to clarify the possibility of endoscopic application of RAS for the real-time diagnosis of the gastric cancer.

Methods. Without any pretreatment, the fresh specimens were measured with a near-infrared multichannel Raman spectroscopic system with an excitation wavelength of 1064nm, and Raman spectra specific for the specimens were obtained. A principal component analysis (PCA) was performed to distinguish gastric cancer and non-neoplastic tissue, and a discriminant analysis was used to evaluate the accuracy of the gastric cancer diagnosis. We reported sensitivity of RAS was 66%, specificity was 73%, and accuracy was 70%, respectively, by using 251 fresh biopsy specimens of gastric carcinoma and non-neoplastic mucosa. In the present study, we examined the Raman spectrum of the resected specimen of gastric cancers to clarify the clinical usefulness of RAS during endoscopy, because Raman spectrum during endoscopy is obtained from whole layer of stomach. Raman spectrum from mucosal side were obtained at 213 points (81 cancer points and 132 non-cancer points) by using 10 resected specimen (12 lesions).

Results. Raman spectrum in the cancer points clearly differed from those for on-neoplastic points, especially at around 1644cm−1, as well as those observed in biopsied specimens. Analysis with PCA demonstrated that sensitivity was 72%, specificity was 78%, and accuracy was 76%. Interestingly, in the advanced gastric cancers, specificity was increased to 92% resulting that accuracy was also increased to 88%. Furthermore, the comparison between cancer points and non-cancer points without atrophy demonstrated that sensitivity was 89%, specificity was 86%, and accuracy was 88%. Conclusions. RAS with a 1064-nm excitation wavelength is useful for gastric cancer diagnosis. Raman diagnostic system may be improved according to the development of both systemic instruments and analytical methods. And in the near future, RAS makes possible real-time diagnosis of gastric cancer during endoscopy.

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 4333.
Abstract 4334: Tracking of leukemic stem cells from frozen and fresh samples in a mouse model of human AML using bioluminescent imaging
Authors: Graham Amanda , Stuart Robert , LaRue Amanda , Rosol Michael
Published: February 2011

The aim of this study was to develop and use in vivo bioluminescent imaging to track luciferase-labeled acute myelogenous leukemia (AML) stem cells in immunedeficient mice using AML blood samples collected from patients and subsequently frozen. The ability to utilize frozen pools as viable reservoirs for AML stem cells will allow the freezing back of samples from the clinic for later use as well as the use of previously frozen samples for engraftment and therapeutic studies. Using thawed, isolated cells, we assessed AML stem cell engraftment and quantitatively compared it to fresh AML stem cell engraftment in a xenogeneic transplant model using in vivo bioluminescent imaging. Frozen AML stem cell engraftment was also compared to both fresh and frozen normal human hematopoietic stem cell (HSC) engraftment. Human hematopoietic stem cells (CD34+CD38-) were isolated from both normal frozen cord blood and from peripheral blood of AML patients using modified Miltenyi Biotec (Auburn, CA) magnetic bead isolation kits. Isolated cells were transduced with a lentiviral vector (HIV-luc) pseudotyped with the VSV envelope able to express the firefly luciferase (fluc) gene in human HSC. Using a Caliper Life Sciences/Xenogen IVIS 200 optical imaging system, cells were assayed for fluc expression 48 hours after transduction. Transduced HSC and AML cells (6 × 104 each) were transplanted intravenously into sublethally irradiated adult NOD/SCID/IL2Rγnull mice and mice were imaged serially from day 1 up to day 160 post transplantation. Observed bioluminescent signal and patterns from thawed AML stem cell engraftment are similar to our results obtained using fresh AML stem cells in mice. Luminescent signal from peripheral blood and spleen engraftment of frozen AML stem cells was detectable within one day post-transplantation. Both fresh and frozen AML stem cell populations exhibited diffuse whole body signal, indicating luciferase-labeled stem cells in the peripheral blood, similar to a hallmark of the disease in humans. The presence of blast cells consistent with AML disease was observed by blood smears taken from mice engrafted with frozen AML stem cells. To our knowledge, we have demonstrated for the first time that frozen AML stem cells can be efficiently labeled with a luciferase enzyme for dynamic tracking in vivo. This work has resulted in a useful animal model that will enable us to elucidate the engraftment patterns of AML cells in living animals, learn the dynamics of the interplay of different stem cell populations, and provide information on the efficacy of new treatments in the future.

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 4334.
Abstract 4337: Tumor T1 is a non-invasive imaging marker of proliferation and response to the mTOR inhibitor everolimus
Authors: McSheehy Paul MJ , Allegrini Peter R. , Becquet Mike , Fux Caroline , Sticker-Jantscheff Melanie , Weidensteiner Claudia
Published: February 2011

Everolimus (RAD001) is an oral mTOR inhibitor recently approved for the treatment of 2nd-line renal cell carcinoma. Further development of everolimus will be aided by identification of suitable biomarkers for stratification and response. We have recently shown that a non-invasive imaging marker (T1) is a rapid and robust marker of tumor response for five compounds with different mechanisms of action (Clinical Cancer Res, in the press). Here we have correlated everolimus-induced changes in T1 (ΔT1) with other biomarkers in a murine tumor model.

Cultured RIF-1 tumor cells (5 × 106) were injected s.c. in the flank of C3H mice (n=14) and after 2 weeks were treated daily with everolimus (10 mg/kg p.o.) or vehicle. Magnetic resonance imaging and 1H-spectroscopy (MRI and MRS) were performed at baseline (day-0) and after 5 days treatment on a 2 mm central slice of the tumor. MRI used an IR-TrueFISP method to quantify the spin-lattice relaxation time of water protons (T1) and MRS the total levels of choline as the Cho/H20 ratio. On day-5, the tumors were ablated, paraffin-embedded and stained for the percentage of cells positive for cleaved caspase-3 (apoptosis) and the proliferation marker Ki67 using immunohistochemistry (IHC). Effects are summarized as mean T/C (ratio of treated divided by vehicle-control); relationships between variables were assessed using Pearson's correlation, with significance set at p<0.05.

Everolimus strongly inhibited tumor growth (T/C=0.05, p<0.001) without affecting body-weight. MRS signals for the proliferation marker choline were clearly detectable in all tumors at both baseline and day-5, and in 6/7 tumors everolimus reduced choline (T/C=0.73, p=0.1). Basal T1 was 2.3 sec and this was always reduced by everolimus (T/C=0.89, p<0.01). The ΔT1 correlated with changes in tumor volume, dTVol (r=0.81, p<0.001), and choline (r=0.59, p<0.05). Basal T1 did not correlate with dTVol, i.e. did not predict response to everolimus, but choline did (r= −0.81, p<0.05), so that higher levels of choline predicted a better response. IHC showed that the %-caspase and %-necrosis were unchanged by everolimus but the %Ki67 positive cells was reduced (T/C=0.55, p<0.001) and there was a significant correlation between %Ki67 and ΔT1 (r=0.61, p<0.05).

These results show that everolimus-induced changes in T1 correlate with known markers of proliferation. The speed and robustness of the method should facilitate its use in clinical trials as an early-response marker. In this model, basal levels of choline were a stratifier for response to everolimus.

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 4337.
Abstract 4339: Prostate adenocarcinoma and intensive bone remodeling in a new prostate cancer IGR-CaP1 preclinical model derived from a human primary tumor
Authors: NAKOUZI Nader AL , OPOLON Paule , BAWA Olivia , LERONDEL Stéphanie , LEPAPE Alain , GONIN Patrick , MASSARD Christophe , FIZAZI Karim , CHAUCHEREAU Anne
Published: February 2011

Background: Bone metastasis is a frequent and catastrophic consequence of prostate cancer for which new therapeutics are needed. Although animal models of bone metastatic prostate cancer are necessary for understanding disease mechanism, there is currently no animal model that recapitulates spontaneous clinical bone metastases. We recently obtained the new prostate cell line IGR-CaP1 derived from primary epithelial prostate cancer. In vitro characterization indicates that IGR-CaP1 corresponds to basal epithelial cells expressing cancer stem cell markers and showing TP53 and PTEN mutations.

Methods: We realized orthotopic xenografts in male nude mice and used bioluminescence imaging (BLI) that allows the sensitive detection of luciferase-expressing IGR-CaP1 cells to monitor tumor growth. Bone lesions were observed after direct intratibial injections of 5×105 luciferase-expressing IGR-CaP1 cells and were evaluated by both Computerized Tomography (CT) and micro Single-Photon Emission Computed Tomography (SPECT) measuring the bone fixation of 99Tc methylenediphosphonate (m99Tc-MDP).

Results: Orthotopic injection of IGR-CaP1 cells yielded an undifferentiated prostate adenocarcinoma capable of metastasizing mainly to the liver and lung. Contrasting with the widely used PC3 preclinical model showing only osteolytic activity after bone injection, high resolution CT scan showed both lytic lesions in the cortical bone and osteoblastic lesions in the trabecular bone in 86% of the IGR-CaP1-injected mice (6/7). The mixed nature of the lesions and the intensive bone remodeling were confirmed by bone scintigraphy with a ∼5 fold increase in m99Tc-MDP fixation in injected tibia versus control tibia (P=0.0013).

Conclusion: IGR-CaP1 is a rare model issued from a primary tumor able to reconstitute prostate adenocarcinoma in animals. Characterization of this new model of prostate cancer will provide investigators with an exceptional tool for developing approaches that prevent and treat prostate cancer bone metastases. In this way, micro SPECT/CT imaging provides a useful and quantitative tool for evaluating bone-targeting therapeutic strategies.

Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend.

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 4339.
Abstract 4340: Development of short peptide probe to detect ovarian tumors in mouse xenograph model
Authors: Lee Chung-Wein , Guo Lili , Stantz Keith M.
Published: February 2011

Ovarian cancer is the leading gynecological cancer in women with five year survival rate less than 45%. Ovarian epithelial cancer (OEC) is responsible for 80 to 90% of all ovarian cancers. Postmenopausal women have high level of gonadotropin in blood circulation and high occurrence of OEC. Therefore, the elevated gonadotropin, follicle-stimulating hormone (FSH) and luteinizing hormone (LH), was proposed to contribute to transforming normal ovarian surface epithelium to neoplastic ovarian surface epithelium. The studies on gonadotropin theory have shown both supporting and conflicting epidemiological and animal model data. Therefore an alternative marker for ovarian tumor detection is beneficial. Follicule Stimulating Hormone Receptor (FSHR) belongs to G protein-coupled receptor with substantial extracellular domains. FSHR normally is expressed on normal ovarian epithelium, fallopian tubal epithelium and similar to LHR. Recent studies showed that FSHR is not only highly expressed on many ovarian cancer cells but is involved in the development and progression of ovarian cancer influencing a variety of oncogenes (MAPK, EGFR, c-myc, and HER-2/neu). FSHR expression increases in the following order: ovarian epithelial inclusions (OEIs) to benigh ovarian epithelial tumor (OET), and to borderline OETs. Our objective is to develop a small molecular weight ligand to image in vivo the differential expression of FSHR associated with OEC, possibility to image OEC in different stages with associated binding molecules to FSHR. D.W. Lee et. al. identified a deca peptide (BI-10) from FSH binding inhibitor of bovine serum. We conjugated the BI-10 peptide with fluorescein to quantify the binding affinity of OVCAR3 cells in flow cytometry. The cells were incubated with BI-10 at various concentrations for 30 mins before removing from six-well culture plates and washing off free peptides. The EC50 of BI-10 binding peptides to OVCAR3 was measured to be 160 μM, with an average number of FSHR receptors per cell to be 1.7×107. After conjugating a NIR fluorochrome (Alexa Fluoro 750) to this peptide and and injection (i.v.) into a mouse xenograft model of ovarian cancer, OEC tumors were clearly detected by optical imaging. A tumor-to-reference tissue update ratio was measured to be 1.20, 1.26, and 1.43 with respect to injected dose 15, 30, and 120 μg probe, respectively. We are currently testing techniques to further increase the BI-10 peptide binding to various ovarian cancer cell lines both in vitro and in vivo by constructing multimeric forms including the BI-10 peptide with various flexible linkers.

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 4340.
Abstract 4341: Tumor imaging via β-galactosidase fragment complementation with a multifunctional targeted-reporter complex
Authors: Broome Ann-Marie , Lavik Kari , Ramamurthy Gopalakrishnan , Liggett Luther A. , Agnes Richard S. , Basilion James P.
Published: February 2011

Determining the status of cell surface receptors has become routine in the care of patients with cancer and has proven to be helpful in guiding treatment. Mutational events that drive a normal cell to become a cancer cell require the coordinated overexpression of not just one receptor at a time, but rather multiple biomarkers. A growing body of evidence from genomic and proteomic research asserts that several receptors contribute to tumor behavior and these expression patterns are referred to as the cancer signature. Many cancers are characterized by an abnormal increase in the activity of epidermal growth factor receptors (EGFR) and transferrin receptors (TfR). Our data of representative human cancer cell lines demonstrate unique, observable expression patterns for the two receptors. Targeted-reporter imaging agent platforms have real application for noninvasive imaging of the multi-step progression of cancer growth, creating the next frontier in in vivo imaging. To develop imaging tools that take advantage of the diagnostic molecular signature, new technologies must employ a contrast agent or signal-amplifying material conjugated to a targeting agent. By linking a reporter enzyme, such as β-gal, to a targeting moiety, either ligand or a short peptide, signal-amplification at the molecular level is achieved. We divided β-gal into unique, independent polypeptides that reassemble and complement enzymatic activity in bacteria and in mammalian cells. We created two sets of complementing pairs that individually have no enzymatic activity. However, when brought into close proximity, complementing pairs associate, resulting in detectable enzymatic activity. We then constructed a targeting complex composed of reporter fragment, linker, and targeting moiety. Our studies demonstrate a time course- and dose-dependent uptake in vitro. Further, we were able to simultaneously visualize the two cell surface receptors implicated in cancer development, EGFR and TfR, using complementing pairs of the targeted-reporter fragment complex. In addition, we are able to image tumor localization in vivo of the targeted-complexes when a fluorescence reporter is added to the complex, as well as immunohistochemical staining of the β-gal reporter complex ex vivo.

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 4341.
Abstract 4342: Multimodal contrast agent development using imprint lithography based techniques: New tools for examining biodistribution
Authors: Herlihy Kevin P. , Merkel Timothy , DeSimone Joseph M.
Published: February 2011

Modern medicine stands to benefit greatly from recent advances in polymer science. Many state of the art therapeutics suffer from a poor therapeutic index because of poor biodistribution characteristics. To solve this problem, researchers are turning to new methods for therapeutic delivery. Herein we employ a top-down particle fabrication technique called Particle Replication In Non-wetting Templates (PRINT®) to fabricate unique biocompatible particles with distinct therapeutic properties based on their composition, size and shape. The particles are easily tagged with a variety of imaging beacons and radioisotopes making them effective as contrast agents for magnetic resonance imaging, positron emission tomography and fluorescence imaging and as radiotherapeutics. We employ this technique to design particles that will target or de-target specific organs and tissues as a function of their size and shape and therefore more effectively deliver their cargo. To our knowledge this work represents the first example of imprint lithography-based multimodal contrast agents and the first in-depth study of biodistribution of nanometer scale shape and size specific particles.

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 4342.
Abstract 4346: Genome-wide analysis of alternative splicing in pediatric medulloblastoma
Authors: Menghi Francesca , Jacques Thomas S. , Barenco Martino , Hubank Mike , Clifford Steven C. , Ham Jonathan
Published: February 2011

Background

Alternative pre-mRNA splicing (AS) is a key post-transcriptional mechanism for the generation of protein diversity. Abnormal alternative splicing can occur in cancer, and it results in the production of novel transcript variants or in an imbalance between normal mRNA isoforms. In both cases it can affect the global pattern of protein expression within a cell in ways that have the potential to sustain tumour growth.

To investigate patterns of alternative splicing in human pediatric medulloblastoma, we used the Affymetrix Human Exon 1.0 ST array, and measured gene expression at the level of individual exons on a genome-wide scale for 14 medulloblastoma specimens and 5 samples of normal cerebellum.

Results

Through statistical analysis of differential splicing we identified 1260 genes potentially undergoing AS between normal cerebellum and medulloblastoma. We selected 9 examples of differential splicing of cassette exons and validated them by semi-quantitative RT-PCR and sequencing. Interestingly, 3 of the validated genes showed a pattern of AS specific for the molecular subgroup of medulloblastomas characterized by hyper-activation of the Sonic Hedgehog (Shh) signalling pathway. In an in vitro mouse granule cell progenitor (GCP) culture model in which cells depend on Shh for the maintenance of their progenitor state, the pattern of AS in the presence of Shh for 5 out of the 6 genes analyzed recapitulated the pattern observed in the tumor samples, suggesting that the preferential expression of specific transcript forms is regulated during normal cerebellar development. Similarly, by analyzing different stages of post-natal mouse cerebellar development, we observed that the switch between the two alternative transcript forms occurred between P7 and P14, at the time when GCPs mature into post-mitotic neurons.

Conclusions

We identified and validated AS events between normal cerebellum and medulloblastoma and showed that medulloblastomas display patterns of AS which are observed during the normal development of the mouse cerebellum. Further studies will investigate the regulation and biological role of AS events in cerebellar development and tumorigenesis.

Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend.

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 4346.
Abstract 4348: PDGF-Rα expression and activation of the MAPK pathway in disseminated pilocytic astrocytomas
Authors: Engels Alexander C. , Denkhaus Dorota , Gnekow Astrid , Pietsch Torsten
Published: February 2011

Background: Pilocytic astrocytomas (PA) are the most common pediatric tumors of the central nervous system. Approximately 5 % of patients with PA present a tumor dissemination. Although the disseminated growth is associated with severe clinical problems, the genetic alterations associated with this biological behaviour have never been investigated. As the histology of the disseminated PAs does not differ significantly to sporadic PAs, we analyzed the mitogen-activated protein kinase (MAPK) pathway for aberrations because genetic alterations of members of this pathway as BRAF and NF-1 have been found to be involved in the pathogenesis of non-disseminated PAs.

Methods: Immunohistochemical analysis of the platelet-derived growth factor receptors alpha (PDGF-Rα) and beta (PDGF-Rβ), the epidermal growth factor receptor (EGF-R), and the downstream activated (phosphorylated) extracellular signal-regulated kinase (p-ERK) was performed in a cohort of 18 patients suffering disseminated PAs. In three patients, material of the primary tumor and of relapses could be studied. The BRAF gene was analyzed for mutations by single strand conformation analysis (SSCP) and for gene copy numbers by semiquantitative PCR with fluorescence labelled primers. Mutational hotspots of the K-RAS, H-RAS and N-RAS genes were also analyzed by SSCP.

Results: Immunohistochemical analysis showed an increased PDGF-Rα expression in 71.4% of disseminated Pas, but no significant overexpression of EGF-R or PDGF-Rβ. A constitutive ERK phosphorylation was demonstrated in all disseminated PAs investigated. Molecular genetic analysis revealed V600E BRAF mutations in two tumors and an increased BRAF gene copy level in the majority of cases. No RAS mutations were found.

Conclusions: This is the first report of molecular genetic analysis of the MAPK pathway in disseminated pilocytic astrocytomas. It underlines the important role of the upstream PDGF-Rα receptor and subsequent MAPK pathway in these gliomas. Its activation represents a potential target for novel strategies in the therapy of disseminated PAs.

Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend.

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 4348.