A B S T R A C T Previous studies have shown that plasma transcobalamin II (TCII) facilitates the cellular uptake of [57Co]vitamin B13 (Be) by a variety of tissues, but the lack of an intrinsic label on the protein moiety of the TCII-Bn complex has made it impossible to determine the role and fate of TCII during this process. We have labeled homogeneous rabbit and human TCII with 'I-labeled N-succinimidyl-3-(4-hydroxyphenyl) propionate and have performed in vivo experiments in rabbits.When 'M-labeled rabbit TCII-['"Co] B1s and 'I-labeled bovine albumin were simultaneously injected intravenously, we observed that 'I and 'Co were cleared from plasma at a faster rate (ti = 11 h) than 'I and that 12J and 57Co were present in excess of 'I in the kidney, liver, spleen, heart, lung, and small intestine j h after injection. Later, "7Co remained in excess of PI, but the ratio of 'I to 'I decreased progressively in all of these tissues; 'I fragments (mol wt less than 1,000) appeared in the plasma and were rapidly excreted in the urine. After 1 h following injection, 'Co was present in excess of 'I in the plasma. Additional experiments revealed that both isotopic moieties of human TCII-Bn were cleared from rabbit plasma 30% faster than their rabbit TCII-Bn3 counterparts and that apo-rabbit TCII and apo-human TCII were cleared from rabbit plasma 30% faster than holo-rabbit TCII and holo-human TCII, respectively.These studies, and appropriate control experiments, indicate that TCII and the TCII-Bu2 complex are cleared

Leptin protein may not be produced but rather stored in human gastric cells. Leptin-induced increases in the proliferation of gastric mucosa cells suggests that leptin might contribute to mucosal integrity and gastroprotection.

No abstract

293 Background: RAD001 is an oral mTOR inhibitor that has exhibited activity in breast cancer. Triple negative breast cancer cells are unable to repair double stranded DNA breaks and hence have sensitivity to platinum agents that cause interstrand cross-links. Rapamycin acts synergistically with platinum agents to induce apoptosis and inhibit proliferation in at least two different breast cancer cell lines (including ER/PR negative cell lines). We propose that combination RAD001 and carboplatin may have activity in triple-negative breast cancer. Methods: The primary objective of the study is to determine clinical benefit (complete remission; CR, partial remission; PR and stable disease; SD) and the toxicity of this combination in women with triple negative metastatic breast cancer who had received 0-3 prior chemotherapy regimens for metastatic disease. Prior carboplatin was allowed. Women with treated brain metastasis were eligible. Secondary objectives were to determine progression free survival. According to the original study plan, carboplatin AUC 6, was to be given intravenously every three weeks. 5 mg of RAD001 was to be given daily with a 3 patient run-in and then 10 mg daily if there were no dose-limiting toxicities. Due to a surprising amount of thrombocytopenia with this combination the dose of carboplatin was first amended to AUC 5 and most recently to AUC 4 with 5 mg of RAD001 (and no plan to escalate to 10 mg). Results: Eleven patients of a planned 25 have been recruited thus far. Median age is 62. Median number of prior regimens is 1. Of the 6 patients assessable for response at this time, four have SD and two have had a PR. 1 SD was achieved in a patient progressing on prior carboplatin at study entry. Five of 7 patients assessable for toxicity had grade 3 or 4 thrombocytopenia and 2 patients had grade 3 neutropenia. All patients have had treatment held and/or dose reductions secondary to hematological toxicity. There have been no non-hematological grade 3 or 4 toxicities. Conclusions: Clinical benefit was observed in all 6 evaluable patients. Dose limiting thrombocytopenia was an unexpected side effect requiring protocol amendment. We continue to accrue study subjects at the amended dosing.

Purpose/Objective(s): Inflammatory breast cancer (IBC) is a highly aggressive and radiation resistant malignancy with a dismal prognosis despite multimodality therapy, including ionizing radiation. We have previously shown that the unique pathogenic properties of IBC result in part from over-expression of translation initiation factor eIF4G1, which is part of the eIF4F translation initiation complex, along with eIF4E and eIF4A. eIF4F is regulated by mTOR, providing a promising target for anti-cancer therapeutics. We demonstrated that protein synthesis is highly regulated during IR by the DNA-damage response (DDR) pathway through mTOR signaling. Many key proteins required for the DDR pathway are encoded by mRNAs that require high levels of the eIF4F complex and mTOR activity for their efficient translation. We hypothesized that upregulation of eIF4F in IBC plays a crucial role in the radio-resistance of disease. Materials/Methods: Experiments were conducted in IBC SUM149 cells. eIF4G1, eIF4E and eIF4A were silenced through the generation of stable cell lines that express tetracycline-inducible shRNAs. eIF4A was also inhibited using the pharmacologic investigational inhibitor DAMD-PatA. Radiation sensitivity in vitro was determined by cell survival assay. Tumor xenografts were generated by the injection of stable shRNA inducible cell lines into nude mice. IBC SUM149 cancer stem cells (CSC) from both in vitro and in vivo experiments were analyzed by a combination of cell surface marker analysis, mammosphere formation and Aldefluor assays. Results: We show that moderate inhibition by silencing of individual components (or by pharmacologic inhibition of eIF4A) of the eIF4F complex prevents IBC xenograft tumor growth and strongly enhances radiosensitivity. In contrast to results obtained for non-transformed breast epithelial cells, reducing the high levels of eIF4G1 in epithelial IBC cells in 2D cultures provides no enhancement in radiation sensitivity. Rather, SUM149 IBC cells harbor a substantial population of CSCs, which are the cells that are strongly dependent on high levels of eIF4G1, and which are selectively radio-sensitized as a result of eIF4G1-silencing. CSCs also require eIF4E and eIF4A activity in order to survive radiation treatment. We also demonstrate that silencing of eIF4G1 radio-sensitizes the stem cell population within IBC tumor xenografts. Radio-resistance of IBC cells is likely mediated by differential responses to the DDR in the stem-cell populations and by selective mRNA translation of proteins involved in the DDR pathway. Conclusions: Our results demonstrate that regulation of mRNA translation plays an important role in conferring radio-resistance to advanced breast cancers, particularly by allowing the survival of the CSC compartment. While inhibition of eIF4F enhances radiation sensitivity in non-transformed cells, this process is abrogated in IBC due to enrichment of a radiation resistant CSC population, demonstrating translational control of the breast cancer stem cell population, and providing a novel understanding of the role of the regulation of mRNA translation in radiation resistance of breast cancer. Citation Information: Cancer Res 2012;72(24 Suppl):Abstract nr P5-03-02.