bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 2415: Evaluation of a DNA vaccine and adjuvant construct for producing anti tumor immunity targeting IGFBP2
Authors: Holt Gregory E. , Gad Ekram , Dang Yushe , Disis Mary
Published: February 2011

DNA based vaccines offer the advantage of stability, low cost, and longer duration of antigen exposure than protein based vaccines. We sought to evaluate strategies to augment the anti tumor potency. Using three immunogenic peptides from the ovarian cancer antigen, human insulin-like growth factor binding protein-2 (IGFBP2) we created a DNA epitope construct that encodes these three peptides in sequence and vaccinated mice with the epitope vaccine or an admixture of the peptides or plasmids expressing either the full length human or mouse genes. Mice received three vaccinations separated by two weeks followed fourteen days later by a lethal injection of syngeneic MMC tumor cells known to express IGFBP2. The DNA epitope vaccine produced a greater level of protection than any of the other constructs proving it to be the best formulation for inducing protective anti tumor responses. Full length human and mouse IGFBP2 DNA constructs induced a similar level of protection as the peptide based vaccine. This IGFBP2 epitope DNA vaccine was then tested in conjunction with plasmids encoding murine GM-CSF and IL-12 either alone or in combination in an identical vaccination strategy. Each genetic adjuvant either alone or in combination improved the level of protection induced by the IGFBP2 DNA epitope vaccine though no difference in tumor protection was seen between adjuvants used either alone or together. Evaluation of the inoculation site revealed a persistence of CD11c+ dendritic cells and CD3+ T cells fourteen days after final vaccination predominantly in mice who received plasmids containing the cytokines versus those who received soluble protein. These data support the use of epitope based DNA vaccines in conjunction with genetic adjuvants for the production of anti tumor efficacy.

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 2415.
Abstract 2420: Modified vaccinia Ankara (MVA) expressing survivin combined with gemcitabine generates specific antitumor effects in a murine pancreatic carcinoma model
Authors: Ishizaki Hidenobu , Manuel Edwin , Srivastava Tumul , Song Guang-Yun , Diamond Don J. , Ellenhorn Joshua D.I.
Published: February 2011

Survivin is overexpressed by 70-80% of pancreatic cancers, and is associated with resistance to chemotherapy and a poor prognosis. Gemcitabine has been a standard treatment for patients with advanced pancreatic cancer for a decade. Recent reports have demonstrated that gemcitabine treatment attenuates the tumor-suppressive environment by eliminating CD11b+/Gr-1+ myeloid derived suppressor cells (MDSCs). It may also enhance the effect of cancer vaccines. The highly attenuated strain of vaccinia, modified vaccinia Ankara (MVA) has been employed as a vaccine vector to deliver a tumor associated antigen, and MVA vaccines have demonstrated safety, immunogenicity, and evidence of clinical benefit in patients with cancer. We hypothesize that a cancer vaccine targeting survivin can achieve enhanced efficacy when use in combination with gemcitabine. In this study, we tested this hypothesis using an MVA expressing full-length murine survivin. The poorly immunogenic mouse pancreas adenocarcinoma cell line, Pan02, which express murine survivin and syngeneic to C57BL/6, was used for this study. To develop an established tumor model, mice were challenged with 2.5 × 105 Pan02 cells by subcutaneous injection. When the tumors were palpable and reached 4 to 5 mm in diameter, mice were immunized twice at a 1-week interval with 5 × 107 pfu of MVA-survivin or control MVA. Immunization with MVA-survivin resulted in a modest therapeutic antitumor effect on established Pan02 tumors. Gemcitabine (60mg/kg) was administrated 48 hours before first immunization. Mice treated with a control MVA or PBS with gemcitabine all died of progressive tumor by day 35. When administered with gemcitabine, MVA-survivin immunization resulted in significant tumor regression and prolonged survival. The enhanced vaccine efficacy is in part due to decreased CD11b+/Gr-1+ MDSCs. To analyze the survivin specific immune response to MVA-survivin immunization, we utilized a peptide library of 15mers with 11 residues overlapping from full-length murine survivin. Splencytes from mice immunized with MVA-survivin produced intracellular γ-interferon in response to in vitro stimulation with the overlapping peptide library. These data suggest that vaccination with MVA-survivin in combination with gemcitabine represents an attractive strategy to overcome tumor-induced peripheral immune tolerance, and this effect may be associated with a clinical benefit in pancreatic 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 2420.
Abstract 2426: Anti-Glypican3 antibody for treatment of human liver cancer
Authors: Ishiguro Takahiro , Kinoshita Yasuko , Sugimoto Masamichi , Miyazaki Yoko , Kato Atsuhiko , Nakano Kiyotaka , Takai Hirotake , Tsunoda Hiroyuki , Sugo Izumi , Ohizumi Iwao , Aburatani Hiroyuki , Hamakubo Takao , Kodama Tatsuhiko , Tsuchiya Masayuki , Okabe Hisafumi Yamada
Published: February 2011

Glypican-3 (GPC3) is a member of the glypican family of heparan sulfate proteoglycans, which are linked to the cell surface through a glycosyl phosphatidyl inositol anchor. GPC3 has been reported to be highly expressed in the majority (70-100%) of HCC, and considered to play a role in the tumorigenesis of HCC. Although the molecular mechanism by which GPC3 functions in tumorigenesis has not been fully elucidated, the high prevalence in HCC has led to considerable interest in GPC3 as a diagnostic marker and therapeutic target.

In this study, we obtained a monoclonal antibody (mAb) against the COOH-terminal part of GPC3, which induced antibody-dependent cellular cytotoxicity (ADCC). The mAb, designated mGC33, exhibited marked tumor growth inhibition of s.c. transplanted Hep G2 and HuH-7 xenografts that expressed GPC3 but did not inhibit growth of the SK-HEP-1 that was negative for GPC3. mGC33 was efficacious even in an orthotopic model; it markedly reduced the blood alpha-fetoprotein levels of mice intrahepatically transplanted with Hep G2 cells. To develop an antibody-based immunotherapy, we generated humanized GC33 (hGC33) by complementarily determining region (CDR) grafting. hGC33 was as efficacious as mGC33 against the Hep G2 xenograft, but hGC33 lacking carbohydrate moieties caused neither ADCC nor tumor growth inhibition. Depletion of CD56+ cells from human peripheral blood mononuclear cells markedly abrogated the ADCC caused by hGC33. The results show that the antitumor activity of hGC33 is mainly attributable to ADCC, and in human, natural killer cell-mediated ADCC is one possible mechanism of the antitumor effects by GC33. We also evaluated the antitumor activity of hGC33 combined with standard chemotherapy agent sorafenib. hGC33 and sorafenib combination was more potent in inhibiting tumor growth than sorafenib alone in the s.c. transplanted Hep G2 xenograft model. Administration of sorafenib alone did not change the GPC3 expression level in xenograft tumor. These suggest that this combination regimen may be clinically useful as an anti-liver cancer therapy.

In careful examination of the safety of hGC33 in nonclinical studies, specific adverse findings on GPC3 expressed tissue or organs were not observed after repeated administration. Therefore hGC33 will provide a novel treatment option for liver cancer patients with GPC3-positive tumors.

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 2426.
Abstract 2428: PPMX-2017: Antibody against CDH3 with potent efficacy against lung cancer
Authors: Zhang Lilin , Ishii Keisuke , Kayukawa Yoko , Yagami Shuing , Kouda Katsushi , Katsumi Keiko , Sakamoto Aya , Satoh Hirokazu , Onishi Hiroshi , Nomura Fumiko , Kotaka Romi , Matsuura Tadashi , Sudo Yukio , Aburatani Hiroyuki
Published: February 2011

Cadherin 3/P-cadherin (CDH3) is a member of cadherin family proteins involved in the cell-cell adhesion. Based on a genome-wide cDNA microarray analysis of pancreatic cancer, we found that CDH3 is overexpressed in pancreatic cancer. We also confirmed by our immunohistochemistry study that CDH3 protein was expressed highly in pancreatic, lung, colon and other types of cancer tissues but not in normal tissues. Since CDH3 is a transmembrane glycoprotein and overexpressed in cancer tissues, it is an attractive therapeutic target for various kinds of cancer. In this present study, we generated a series of monoclonal antibodies against CDH3 and evaluated their anti-tumor effect both in vitro and in vivo. PPMX2017, an antibody from the series, is an anti-human CDH3 specific mouse IgG2a antibody. The in vitro studies demonstrated that PPMX2017 elicited strong antibody dependent cellular cytotoxicity (ADCC) activity on CDH3-positive cancer cell lines (lung cancer: NCI-H358 and A431, pancreatic cancer: KLM1 and Bxpc3.). Furthermore, PPMX2017 showed a significant antitumor effect in a therapeutic xenograft model of lung cancer where NCI-H358 cell-generated tumors grew to 70∼90 mm3 before administration of this antibody. PPMX2017 suppressed tumor growth by 60∼80% compared with control IgG in a dose range of 0.3mg∼5mg/kg. These findings show that CDH3 is a potential target for cancer therapy and PPMX2017 is a candidate for the development of antibody therapeutics.

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 2428.
Abstract 2429: Evidence of a therapeutic window with a T cell-engaging BiTE antibody based on monoclonal antibody cetuximab
Authors: Lutterbuese Ralf , Thomas Oliver , Raum Tobias , Kischel Roman , Hoffmann Patrick , Friedrich Matthias , Lorenczweski Grit , Wolf Andreas , Urbig Thomas , Schaller Evelyn , Rau Doris , Mangold Susanne , Baeuerle Patrick A. , Kufer Peter , Rattel Benno
Published: February 2011

Treatment with chimeric monoclonal antibody cetuximab has become the standard of care for patients with advanced colorectal cancer (CRC). Cetuximab and other EGFR-specific monoclonal antibodies predominantly inhibit cancer growth by interfering with receptor signalling. Recent analyses have shown that CRC patients with mutated KRAS and BRAF oncogenes do not profit from treatment with such antibodies. Here we have used the binding domains of cetuximab to construct a BiTE antibody that can directly engage and activate T cells by transiently connecting T cells via the CD3 subunit with EGFR-expressing target cells.

The cetuximab-based BiTE antibody (C-BiTE) showed potent redirected lysis of KRAS- and BRAF-mutated human CRC lines HCT116 and HT-29, respectively, by unstimulated peripheral human T cells at half maximal concentrations of 10-40 pg/ml (ca. 0.2-0.8 picomolar). The BiTE antibody also completely prevented the outgrowth of tumors from HCT116 and HT-29 xenografts in mice at doses as low as 0.005 mg/kg when given daily i.v. for 10 days. The monoclonal antibody cetuximab was not effective in these models even at 50 mg/kg given twice weekly for 4 weeks.

As a prerequisite for nonclinical safety assessment of C-BiTE in non-human primates, crossreactive binding of C-BiTE to CD3 and EGFR antigens of human and Cynomolgus monkey (Macaca fascicularis) origin was demonstrated. This binding translated into a comparable potency of redirected lysis of human EGFR-expressing cancer cells by stimulated Cynomolgus and human PBMC. Continuous i.v. infusion of 6.2 or 12.4 µg/kg C-BiTE to Cynomolgus monkeys for 3 weeks was well tolerated and led to relatively constant serum concentrations of C-BiTE between 1 and 10 ng/ml. In vitro, these serum levels would support a >90% lysis of EGFR-overexpressing CRC cells within 24 h by both Cynomolgus and human effector cells. Of note, no skin toxicity was observed during the 3-week exposure to C-BiTE. The highest doses of 31 and 154 µg/kg caused lethal damage to EGFR-expressing cells in vital organs. Inflammatory cell infiltration into the EGFR-positive tissues was observed at all dose levels but its incidence and severity showed no correlation with clinical signs.

Our data suggest that C-BiTE can be safely administered to a relevant non-human primate species at dose levels similar to those of CD19-specific BiTE antibody blinatumumab, which has shown partial and complete responses in lymphoma patients. We anticipate that C-BiTE has a therapeutic window in humans.

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 2429.
Abstract 2433: Eradication of established pancreatic tumors in mice by engagement of extra-tumoral human T cells with BiTE antibody MT110
Authors: Friedrich Matthias , Drache Daniela , Amann Maria , Filusch Susanne , Voelkel Markus , Thomas Oliver , Baeuerle Patrick A. , Rattel Benno
Published: February 2011

MT110 is a T cell-engaging, EpCAM/CD3-bispecific BiTE antibody that is currently being tested in a dose-escalating clinical phase 1 study in gastrointestinal and lung cancer patients. MT110 has shown high potency of redirected lysis in cell-based assays at picomolar concentrations.

Previous studies with MT110 have shown that subcutaneous (s.c.) pancreatic tumor formation can be delayed in NOD/SCID mice when human tumor cells are mixed prior to inoculation with human peripheral blood mononuclear cells (PBMC) at a ratio of 1:2. Likewise, MT110 is able to eradicate s.c.-implanted human metastatic tissue (<80 mm3) freshly dissected from ovarian cancer patients. In this model, the only source of effector cells were tumor-resident T cells from the respective patient sample.

Here we have further explored in a new NOD/SCID mouse model the anti-tumor activity of MT110 on established s.c. tumors derived from the human pancreas cancer cell line Aspc1. Effector cells, human peripheral T cells, were expanded ex vivo in the presence of T cell mitogens. NOD/SCID mice were reconstituted with human T cells by intraperitoneal (i.p.) injection once the tumors had reached at least 150 mm3. Following adoptive transfer, human T cells were detected in the peripheral blood of mice for up to 6 days. Tumor-bearing mice were treated for 28 days with 350 µg MT110/kg (approx. 3 µg MT110/mouse) by either daily i.v. bolus injection or continuous i.p. infusion using Alzet osmotic pumps.

With the onset of MT110 treatment by either route, Aspc1 tumors of approx. 300 mm3 started to shrink. After two weeks, tumors in all MT110-treated mice were less than

50 mm3, and 3 out of animals 11animals were completely free of tumor. In contrast, all tumors in vehicle control mice had reached an average size of 500 mm3 during this period. At the end of the study after 8 weeks and 2.5 weeks after last dosing, 3 out of 11 animals remained tumor-free. Among the remaining eight tumor-bearing animals, all showed tumor regression 38-40 days after start of the study. Our data show that established tumors grown can be eradicated by BiTE antibody MT110 by engaging initially extra-tumoral human T cells. This indicates that cancer therapy with MT110 and other BiTE antibodies in development may not solely rely on tumor-resident T cells. Moreover, MT110 may have activity against pancreatic 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 2433.
Abstract 2436: AGS-16M8F is a novel antibody drug conjugate (ADC) for treating renal and liver cancers
Authors: Gudas Jean M. , Torgov Michael , An Zili , Jia Xiao- Chi , Morrison Karen J. , Morrison Robert K. , Yang Peng , Raitano Arthur B. , Kanner Steven B. , Jakobovits Aya
Published: February 2011

AGS-16 is a novel transmembrane antigen discovered by Agensys using transcription profiling and is expressed in kidney, liver and other cancers. A fully human IgG2k monoclonal antibody that binds to AGS-16 with high affinity was conjugated to the anti-microtubulin drug monomethyl auristatin F (MMAF) via a noncleavable maleimidocaproyl (mc) linker to yield the antibody drug conjugate, AGS-16M8F. AGS-16M8F was evaluated for binding affinity, species cross reactivity and cell cytotoxicity in vitro. Its’ anti-tumor activity was investigated using multiple established patient-derived and cell line models of renal clear cell carcinomas. The pharmacokinetics (PK) of AGS-16M8F was evaluated in mice. In addition, the expression of AGS-16 protein in patient kidney and liver cancer specimens was confirmed using IHC. AGS-16M8F bound with high affinity (Kd = 0.3 nM) to both human and cynomolgus monkey AGS-16 orthologs. Following cell surface binding, AGS-16M8F was internalized and trafficked to lysosomes leading to catabolism and release of active drug metabolite. AGS-16M8F mediated potent cell cytotoxicity in vitro and led to significant and prolonged growth inhibition or complete eradication of multiple established renal cancer xenograft models, including those grown orthotopically. Pharmacokinetic analysis of AGS-16M8F in mice demonstrated ADC stability. IHC analysis confirmed that AGS-16 was highly expressed in over 80% of renal clear cell cancers and 33% of hepatocellular carcinomas. When considered together these data indicate that AGS-16M8F is a promising therapeutic ADC for the treatment of renal and other AGS-16 expressing cancers.

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 2436.
Abstract 2439: Cross-linker evaluation in the design of antibody-SN-38 conjugates for cancer therapy
Authors: Moon Sung-Ju , Tat Fatma , Sheerin Agatha , Zalath Maria , Arrojo Roberto , Cardillo Thomas M. , Govindan Serengulam V. , Goldenberg David M.
Published: February 2011

Antibody (MAb) conjugates of the potent topoisomerase I inhibitor, SN-38, prepared using the bifunctional derivative, ‘CL2-SN-38’ (maleimido-[x]-Phe-Lys-PABOCO-20-O-SN-38), were previously shown to produce significant and selective therapeutic efficacies of human tumor xenografts in nude mice. With a view to examining alternative linkers for possible improvement, we prepared a variant, namely maleimido-[x]-Lys-PABOCO-20-O-SN-38 (‘CL2A-SN-38’), and two other derivatives with cross-linker attachment at the 10-position of SN-38 instead of the 20-position: maleimido-[x]-Phe-Lys-N(Me)-(CH2)2-N(Me)-CO-10-O-SN-38 (‘CL2D-SN-38’) and maleimido-[x]-Phe-Lys-PABOCO-N(Me)-(CH2)2-N(Me)-CO-10-O-SN-38 (‘CL2E-SN-38’). In these, PAB is p-aminobenzyl moiety and ‘x’ contains a short PEG group to increase solubility. Conjugates of the humanized anti-CEACAM5 MAb, hMN-14 (labetuzumab), were prepared by reacting the maleimide-appended SN-38 derivatives with mildly reduced hMN-14. The SN-38/MAb molar substitutions were in the range of 5.9 to 6.3. In vitro studies showed a similarity in the stabilities, in mouse serum, of CL2 and CL2A-based linkers, but an order of magnitude greater stability for conjugates with linker at the 10-position; however, CL2D-SN-38 linker was inert to dipeptide cleavage by cathepsin B at the lysosomal pH of ∼ 5. The latter finding was also reflected in the loss of cytotoxicity due to hMN-14-CL2D-SN-38 conjugate in the LS174T colon cancer cell line in vitro. All conjugates were also evaluated in an exploratory preclinical therapy experiment in nude mice with aggressive s.c. LS174T human colon cancer xenografts. Conjugates were administered i.p. in groups of tumor-bearing animals (n = 10) at a schedule of 0.39 mg/kg of SN-38 equivalent twice weekly × 2 weeks. Untreated animals received saline. Mean tumor volumes (cm3) of animals in various treatment groups, on day 18, were: 0.073 ± 0.035 for the hMN-14-CL2-SN-38 positive control group; 0.055 ± 0.032 for the hMN-14-CL2A-SN-38 group; 0.518 ± 0.239 for the hMN-14-CL2D-SN-38 group; 0.490 ± 0.224 for the hMN-14-CL2E-SN-38 group; and 1.09 ± 0.88 for the untreated group. Median survival times (days) were 54.5, 55, 30, 25, and 26.5 for these groups, respectively. These results showed that a cleavable peptide was not necessary in the SN-38 conjugate design, as hMN-14-CL2A-SN-38 was equivalent to hMN-14-CL2-SN-38 in efficacy, and that hMN-14-CL2E-SN-38 was surprisingly much less efficacious than the CL2 and CL2A-based conjugates despite its higher relative stability.

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 2439.
Abstract 2440: CD47 is a prognostic factor and an antibody target that synergizes with rituximab to eradicate non-Hodgkin lymphoma
Authors: Chao Mark P. , Alizadeh Ash A. , Tang Chad Z. , Myklebust June H. , Varghese Bindu , Jan Max , Cha Adriel C. , Tan Brent T. , Park Christopher Y. , Levy Ronald , Weissman Irving L. , Majeti Ravindra
Published: February 2011

Monoclonal antibodies are standard therapeutics for several cancers including the anti-CD20 antibody rituximab for B cell non-Hodgkin lymphoma (NHL). Rituximab and other antibodies are not curative, and must be combined with cytotoxic chemotherapy for clinical benefit. Here we identify CD47 as an attractive monoclonal antibody target in NHL. A major function of CD47 is to inhibit phagocytosis, through binding its receptor, SIRP-alpha, on phagocytes. We hypothesize that NHL cells over-express CD47 to evade immune phagocytosis and that blockade of CD47 signaling by a monoclonal antibody can eliminate NHL cells by enabling phagocytic engulfment. Here we investigate the therapeutic potential of an anti-CD47 antibody alone and in combination with rituximab for the treatment of NHL. We predict that the combination of anti-CD47 antibody and rituximab through utilization of two independent antibody mechanisms will result in synergistic phagocytosis and elimination of NHL cells.

We found that CD47 protein is highly expressed on primary human NHL cells compared to normal B cell counterparts. Higher CD47 gene expression predicted an inferior outcome in several cohorts of NHL patients, providing independent prognostic value. Blocking anti-CD47 antibodies preferentially enabled phagocytosis of NHL cells but not normal cell counterparts in vitro while the combination of anti-CD47 antibody and rituximab resulted in synergistic phagocytosis of NHL cells. In vivo, anti-CD47 antibody treatment of human NHL-engrafted mice reduced lymphoma burden and prolonged survival compared to IgG control treatment. Furthermore, anti-CD47 antibody treatment in combination with rituximab led to elimination of lymphoma and cure of NHL-engrafted mice compared to either antibody treatment as a single agent. The above in vivo results were observed in both a disseminated and localized human NHL cell line mouse model as well as in primary human lymphoma xenotransplants. These antibodies synergized through a novel mechanism combining Fc receptor (FcR)-dependent and FcR-independent stimulation of phagocytosis that might be applicable to many other cancers. Together, these data provide the rationale for utilizing an anti-CD47 antibody either alone or in combination with rituximab in treating human NHL.

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 2440.
Abstract 2443: Tetraspanin CD151 as a target for antibody-based cancer therapy
Authors: HAEUW Jean-François F.
Published: February 2011

The tetraspanin CD151 is known to be functionally linked to cancer metastasis. In human, increased expression of CD151 is indicative of a poor prognosis in different cancer types. Whereas its precise mechanism of action remains obscure, CD151 was shown to regulate cell motility and adhesion through association with laminin-binding integrins such as α3β1 or α6β4, and more recently the communication between endothelial and tumor cells. Overexpression of CD151 in tumor cells enhances cell migration, cell invasion and experimental metastasis. Silencing of CD151 with specific siRNA was shown to delay tumour formation in vivo in a mouse model of breast cancer. Several anti-CD151 monoclonal antibodies (Mabs) were shown to display anti-metastatic activity in vivo. Inhibition of metastasis was not attributed to any effect of these Mabs on tumor cell growth but was essentially linked to inhibition of cell motility. However, a more direct effect could be expected. For these reasons, we generated different anti-CD151 Mabs via immunisation with transfectant cells and evaluated their anti-tumor activity in xenograft and orthotopic mouse models to further investigate the role of CD151 in cancer progression in vivo. Interestingly, anti-CD151 Mabs inhibited potently primary tumor growth in different xenograft models including prostate, lung and breast cancer models. These results demonstrate that CD151 could be of importance for controlling tumor cell proliferation. In addition, as anticipated, we showed that anti-CD151 Mabs were also able to inhibit metastasis in vivo in an orthotopic A549 lung cancer model with well characterized lung metastases. The mechanism of action of these Mabs on tumor cells is under investigation and preliminary in vitro data will be presented. Altogether, our results strongly suggest that CD151 could function at multiple cancer stages, including not only metastasis cascade steps but also earlier steps of primary tumor growth, reinforcing thus the interest of this innovative target in oncology. Mabs targeting CD151 may be of significant interest for cancer biotherapy.

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 2443.
Abstract 2445: Involvement of neutrophils and natural killer cells in the anti-tumor activity of alemtuzumab in xenograft tumor models
Authors: Siders William M. , Shields Jacqueline , Garron Carrie , Hu Yanping , Boutin Paula , Shankara Srinivas , Weber William , Roberts Bruce , Kaplan Johanne M.
Published: February 2011

Alemtuzumab (Campath) is a recombinant humanized IgG1 monoclonal antibody directed against CD52, an antigen expressed at high levels on the surface of normal and malignant B and T lymphocytes. Alemtuzumab is approved for the treatment of B-cell chronic lymphocytic leukemia (B-CLL), but the exact mechanism by which the antibody depletes malignant lymphocytes in vivo is not clearly defined. To address this issue, the anti-tumor activity of alemtuzumab was studied in disseminated and subcutaneous xenograft tumor models. The density of CD52 target antigen on the surface of tumor cells appeared to be an important factor as an overall correlation was observed between the anti-tumor activity of alemtuzumab and levels of CD52 expression. Deglycosylation of alemtuzumab resulted in a loss of complement-dependent cytotoxicity (CDC) and antibody-dependent cell-mediated cytotoxicity (ADCC) in vitro and was found to abolish anti-tumor activity in vivo. Individual inactivation of these effector mechanisms in tumor-bearing mice indicated that the protective activity of alemtuzumab in vivo was primarily dependent on ADCC mediated by neutrophils and to a lesser extent NK cells, as evidenced by the loss of tumor growth inhibition caused by removal of these populations with antibodies to Gr-1 or asialo-GM-1, respectively. In contrast, inactivation of complement by treatment with cobra venom factor or depletion of macrophages using clodronate liposomes had no significant impact on the anti-tumor activity of alemtuzumab in vivo. Increasing the number of circulating neutrophils by treatment with G-CSF enhanced the anti-tumor activity of the antibody thus providing further evidence for the involvement of neutrophils as effector cells in the activity of alemtuzumab.

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 2445.
Abstract 2446: Biodistribution of antibody drug conjugates by LC-MS
Authors: Srinivasan Mohan , Thevanayagam Lourdes , Chakraborty Indrani , Bell Alasdair , Deshpande Shrikant , Pan Chin , Cardarelli Pina M. , Gangwar Sanjeev , Chong Colin , Cortez Orville , Klitzing David , Jiang Jerry , Greenbaum Meghan , Phillips James
Published: February 2011

Introduction: A method to study biodistribution of adduct forming antibody drug conjugates (ADCs) by LC-MS

Description: ADCs are increasingly used to deliver cytotoxic payloads to tumor cells. While the targeted approach has vastly improved the toxicity profiles of these drugs, it is still important to understand the bio-distribution and the enhanced tumor localization this approach offers. Radio-labeled methods are still the most reliable approach to study the bio-distribution, but may not be able to differentiate between the drug that is present in the tissue from those that have formed covalent adducts upon non-specific internalization. Here, we describe a mass-spectrometric methods to identify these covalent adducts these cytotoxic agents form and how such methods could be utilized to map the bio-distribution in xenograft models.

Data: The antibody drug conjugate is highly localized in the tumor with a %ID/gm value of almost 40 within 24 hours of dosing and no non-specific accumulation in other organs observed.

Conclusions: A simple and robust LC-MS based method has been developed to understand the bio-distribution of antibody-drug conjugates by detecting and quantifying the actual and final chemical state of the toxin in the targeted cell. This method could be used to evaluate the (1) PK of antibody-drug conjugates (2) make lead selections at advanced preclinical stages of antibody-drug development (3) understand the ADME properties of the chosen antibody-drug conjugate

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 2446.
Abstract 2447: Transient lymphocyte decrease due to adhesion & migration following treatment with catumaxomab (anti-EpCAM x anti-CD3) in vivo
Authors: Eberhardt Andrea , Dettmar Kirsten , Atz Judith , Lindemann Carsten , Schroeder Petra , Kluge Michael
Published: February 2011

Catumaxomab is a trifunctional, bispecific antibody targeting EpCAM and CD3 which redirects T-cells to EpCAM expressing tumor cells and is able to evoke T-cell cytotoxic responses. The hybrid Fc region of catumaxomab provides a third functional binding site, which is able to bind and activate Fcγ receptor (FcγR)-positive accessory cells.

In clinical trials a transient decrease in peripheral blood lymphocyte counts was observed following i.p. administration of catumaxomab. Lymphocyte decrease may be a result of either decreased production or increased destruction of lymphocytes, redistribution of lymphocytes or various unknown / multifactorial pathogeneses.

We determined whether the observed transient decrease in patients might be due to antibody induced adhesion of lymphocytes to endothelial cells or to migration of lymphocytes into the tissue. Therefore, we investigated the influence of catumaxomab on adhesion of human T cells to endothelial cells in vitro. Catumaxomab physiologically activated T-cells by increasing expression of the activation marker CD69 and induced the release of cytokines, including TNFα and IFNγ. TNFα increased expression of adhesion molecules CD54 and CD62E on endothelial cells.

Furthermore, catumaxomab dose-dependently increased adhesion of T cells to endothelial cells, and the adhesion was further increased when the endothelial cells were preactivated with TNFα. Thus, catumaxomab increases adhesion of T-cells to endothelial cells, which is due to antibody mediated upregulation of adhesion molecules on T-cells and production of cytokines that upregulate endothelial cell adhesion molecules.

Furthermore, we showed that treatment of BALB/c mice with the related trifunctional antibody BiLu (anti-human EpCAM x anti-mouse CD3), that binds to mouse CD3, also resulted in a dose-dependent transient decrease in CD3+ T-cells (both CD4+ and CD8+) that returned to the normal range within 48 h.

These results provide a rationale for the transient and reversible decrease in peripheral lymphocyte counts observed following catumaxomab administration in clinical studies, which is likely to be due to redistribution of lymphocytes and not due to depletion of circulating lymphocytes or stem cells.

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 2447.
Abstract 2453: P16, a novel monoclonal antibody specifically recognizing human placental growth factor (PLGF), blocks PLGF-VEGFR1 interaction in vitro and inhibits angiogenesis and tumor growth in vivo
Authors: Xu Yiqing , Hu Hong-Qun , Luo Shi-Ping , Sun Ya-Nan , Zhou Qing , Chen Zui , Song Xiao-Qi , Zhou Qunmin
Published: February 2011

Placental growth factor play (PLGF) is a homolog of vascular endothelial growth factor (VEGF) and binds to VEGF receptor-1. Previous studies have indicated its synergistic role with VEGF in angiogenesis, while a recent study suggested its independent role in promoting growth and metastasis of VEGF-inhibitor-resistant tumors. In order to further investigate the role of PLGF in tumor angiogenesis in relationship to VEGF, we have generated a novel PLGF-specific monoclonal antibody (mAb). Hybridomas secreting PLGF-specific mAbs were obtained by fusing splenocytes from mice immunized with yeast-derived recombinant human PLGF protein with Sp/20 myeloma cells; the mAb clone that showed highest binding affinity, designed as P16, was selected for further study. This P16 mAb recognizes human PLGF, in both native and denatured forms, but not VEGF-A, VEGF-B or VEGF-D. P16 mAb is of IgG1-kappa type. Sequence analysis of cDNA encoding the heavy and light chains of P16 mAb showed novel variable regions. Using P16 mAb in immunohistochemical staining assay, increased levels of PLGF protein were found in a number of human tumors from lung, breast and colon. PLGF proteins were also detected in a wide variety of normal human tissues including lung, stomach, colon and kidney, contrary to previous reports. Importantly, P16 mAb blocked the binding of PLGF to VEGFR-1 in a dose-response manner, and inhibited the PLGF-mediated endothelial cell line growth in vitro. Treatment of human tumor xenografts (A673 rhabdomyosarcoma) in nude mice with P16 mAb resulted in marked inhibition of tumor growth, which is similar in degree to the tumor inhibition by VEGF-specific antibody Bevacizumab. A humanized form of P16 mAb was constructed and showed similar inhibition of tumor growth in xenograft nude mice model. In conclusion, P16 mAb is a novel mAb specifically recognizing human PLGF but not VEGF and blocks the binding of PLGF to its receptor. It inhibits angiogenesis and tumor growth, suggesting an important and independent role of PLGF in tumor growth. Further studies on its mechanisms of action and its clinical applications are under way.

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 2453.
Abstract 2457: The MUC1 membrane-bound mucin is an actor in renal clear-cell carcinoma
Authors: Bouillez Audrey , Gnemmi Viviane , Hémon Brigitte , Seuningen Isabelle Van , Zini Laurent , Leroy Xavier , Aubert Sébastien , Perrais Michael
Published: February 2011

MUC1, an O-glycoprotein membrane-bound mucin, is overexpressed in renal clear-cell carcinomas (CRCC) with correlation to two major prognostic factors, Tumor-Node-Metastasis stage and nuclear Fürhman grade. Previously (Aubert et al., Cancer Res 2009), we have shown that (i) MUC1 was significantly overexpressed in metastatic CRCC vs non-metastatic CRCC and (ii) MUC1 is a target gene of HIF-1 transcription factor which is a part of the hypoxia pathway, the main renal carcinogenetic pathway. To better understand the roles of MUC1 in CRCC, we used two renal cell lines expressing MUC1 (Caki-2 cells) or not (ACHN cells). Caki-2 cells were stably transfected with shRNA targeting MUC1 and ACHN cells with an expressing vector containing MUC1 cDNA. Proliferation, migration and invasion properties were studied in vitro in the different cellular clones using Boyden chambers coated with Matrigel, wound healing assay and MTS cell proliferation assay, respectively. Our results showed that (i) MUC1 expression was associated with an increase of invasion and migration properties of renal carcinoma cells whereas no effect on proliferation and apoptotic rate was observed and (ii) levels of phospho-JNK were higher in MUC1 expressing cells. Results of (i) transcriptomic analysis on 44K Agilent microarrays, (ii) signaling pathway screening and (iii) subcutaneous xenografts will be presented. Our results show that MUC1 plays a role in biological properties of renal cancer cells suggesting important function for this mucin in tumour progression and confirms its potential as a therapeutic target in this type of cancer.

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 2457.
Abstract 2462: Overexpression of prostaglandin E2 EP4 receptor enhances mouse skin tumorigenesis
Authors: Simper Melissa S. , Bowen Rebecca A. , Surh Inok , Fischer Susan M.
Published: February 2011

Chronic inflammation has recently been suggested to be involved in cancer development, and cyclooxygenase-2 (COX-2), a key mediator of the inflammatory response, is known to be upregulated in several cancers. COX-2 is responsible for the production of prostaglandins, and prostaglandin E2 (PGE2) is the major prostaglandin involved in inflammation. PGE2 binds to four receptors (EP1-EP4), and the EP4 receptor has recently been shown to be important in carcinogenesis depending on the tissue in which it is expressed. To determine the role of EP4 in skin carcinogenesis, transgenic mice were generated that overexpress EP4 in the basal layer of the skin under the control of the bovine keratin 5 promoter (BK5.EP4). Using a two-stage chemical carcinogenesis protocol, the BK5.EP4 mice were shown to develop both papillomas and squamous cell carcinomas earlier than wild type (WT) mice. However, a tumor observed before 12-O-tetradecanoylphorbol-13-acetate (TPA) treatment on one BK5.EP4 mouse suggested that TPA was not necessary for the promotion of the tumors. Therefore a 7,12-dimethylbenz[a]anthracene (DMBA)-only carcinogenesis protocol was performed, and by 30 weeks, the BK5.EP4 mice again developed more carcinomas compared to none on the WT mice. Immunohistochemistry performed on DMBA-treated mice demonstrated that the BK5.EP4 epidermis began to detach 48 hours after treatment with total detachment occurring 5 days after treatment, while WT mice had only increased epidermal thickening. By 7 to 10 days after DMBA treatment, patches were observed on the BK5.EP4 skin where it appeared the hair and epidermis were detaching, which were not seen on the WT mice. MMP-9, MMP-7, and E-cadherin expression also were upregulated in the BK5.EP4 mice at the same time detachment of the epidermis was occurring. However, no differences in proliferation, stem cell numbers, or CYP1A1 and CYP1B1 expression have been observed. These data suggest that overexpression of EP4 in the mouse skin can increase tumorigenesis, possibly through the wounding of the skin and the subsequent inflammation and epidermal regeneration acting as a tumor promoter. Supported by CA100140.

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 2462.
Abstract 2471: Immunosuppressive effect of elevated epidermal ornithine decarboxylase activity
Authors: Keough Martin P. , DeFeo Karen , Hayes Candace S. , Gilmour Susan K.
Published: February 2011

Tumor promoting agents, such as phorbol esters, promote tumorigenesis via induction of inflammation and hyperplasia. However, the immunosuppressive/immunomodulating properties of tumor promoting agents may play a relevant role in tumor promotion as well. Because elevated polyamine biosynthesis is sufficient to promote skin tumorigenesis, we hypothesized that increased activity of epidermal ornithine decarboxylase (ODC), a key regulatory enzyme in polyamine biosynthesis, may suppress the cutaneous immune response in addition to stimulating proliferation. To investigate the role of ODC in the control of cutaneous inflammation, a well-known model of T cell mediated inflammation was used. A contact hypersensitivity (CHS) response was elicited in the ears of both ODC transgenic mice, in which ODC is targeted to the epidermis, and their normal littermates by topical sensitization and challenge with oxazolone. Unlike in normal littermate mice, ear swelling was dramatically reduced in ODC transgenic mice after elicitation of CHS. Flow cytometric and immunohistochemical analyses of the infiltrating immune cells revealed fewer infiltrating immune cells, especially neutrophils, in the elicited ears of ODC transgenic mice. The specific ODC inhibitor, α-difluoromethylornithine, abrogated the suppressive effects of ODC in CHS reactions. Adoptive transfer of sensitized lymphocytes from either ODC transgenic or normal littermate mice to naïve ODC transgenic or wild-type mice indicated that elevated epidermal ODC activity suppress both the sensitization and elicitation phases of CHS. Cytokines are central mediators of immune functions associated with inflammatory reactions. The cytokine profile of ex vivo stimulated lymphocytes isolated from draining lymph nodes showed reduced levels of IL-6 and IL-10 in sensitized and challenged ODC transgenic mice. In vivo labeling of dendritic cells by topical application of fluorescein isothiocyanate (FITC) showed fewer FITC-labeled dendritic cells in the draining lymph nodes of ODC transgenic mice compared to normal littermates. Vascular permeability was assessed using Evans blue extravasation assays during the elicitation phase of CHS. At 24 hours after induction of CHS, there was 45% less plasma extravasation of Evans blue dye into the ear skin of ODC transgenic mice compared to normal littermates. Collectively these data indicate elevated epidermal ODC activity can suppress a number of functions associated with cutaneous inflammatory reactions. In addition to promoting cellular proliferation, the immune suppression mediated by increased epidermal ODC activity may provide an environment conducive to tumor growth. This work was supported by NIH grant CA070739.

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 2471.
Abstract 2477: The effect of dissociated glucocorticoids on 12-O-tetradecanoylphorbol-13-acetate (TPA) induced skin inflammation in SENCAR mice
Authors: Kowalczyk Piotr , Sosnowska Renata , Kowalczyk Magdalena , Tolstykh Olga , Junco Jacob , Walaszek Zbigniew , Slaga Thomas J. , Hanausek Margaret
Published: February 2011

Glucocorticoids are very effective in preventing carcinogen- and tumor promoter-induced skin inflammation, hyperplasia, and mouse skin tumor formation when applied to skin together with a carcinogen. Their control of cellular functions is mediated by a well-known transcription factor, glucocorticoid receptor (GR). GR acts via two different mechanisms: transcriptional regulation that requires DNA-binding and protein-protein interaction between GR and other transcription factors, such as nuclear factor kappa B (NF-κB) or activator protein 1 (AP-1), which is independent of DNA binding. We hypothesize that only the transrepression activities of the GR are sufficient to suppress skin tumor development. To test our hypothesis, we have synthesized two glucocorticoids (RU24858 and RU24782) that induce only transrepression activities of the GR. These two compounds are able to bind to the GR with high affinity and repress the AP-1 and NF-κB activity while showing reduced transactivation activities in vitro. Different doses (0.001, 0.01 and 0.1 mg) of RU24858, RU24782, desoxymethasone (DES) and fluocinolone acetonide (FA) or acetone were applied to the dorsal skin of female SENCAR mice 20 min. prior to TPA application (2 μg), two times per week for two weeks. Keratinocytes proliferation rates were estimated by measuring epidermis thickness and BrdU incorporation index. Immunohistochemistry and RT-PCR techniques were also used to evaluate expression level of genes involved in inflammation and cell cycle regulation. All studied compounds did not influence keratinocytes proliferation when applied alone. Application of DES, FA and RU24858 reversed TPA induced hyperplasia, in dose-depended manner, while RU24782 treatment had no effect on epidermis thickness or the BrdU incorporation index in TPA treated skins. All tested compounds were able to decrease c-jun mRNA level both in acetone and TPA-treated skins. DES, FA and RU24858 were also able to decrease elevated by TPA mRNA level of IL-6, COX-2 and iNOS. RU24782 did not influence mRNAs levels of these genes. These findings suggest that RU24782 had no effect on TPA-induced inflammation and hyperplasia in SENCAR mice, while RU24858 was able to reduce all tested inflammation markers, having only slight effect on epidermal hyperplasia. Supported by NIH grants CA 0796065-06 and P30 CA 54174-16S1.

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 2477.