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Purpose There is an unmet need for biomarkers for identifying patients likely to benefit from anticancer treatments, selecting dose, and understanding mechanisms of resistance. Plasma vascular endothelial growth factor (VEGF) and soluble VEGF receptor 2 (sVEGFR-2) are known to be modulated by VEGF pathway inhibitors. It is unknown whether chemotherapy or VEGFR inhibitor/chemotherapy combinations induce changes in these or other cytokines and angiogenic factors (CAFs) and whether such changes could be markers of benefit. Methods Thirty-five plasma CAFs were analyzed using multiplexed bead arrays and enzyme-linked immunosorbent assays from 123 patients with non–small-cell lung cancer in a randomized phase II study who received vandetanib, a VEGFR and epidermal growth factor receptor inhibitor, monotherapy carboplatin and paclitaxel (CP), or the combination (VCP). Changes in CAFs at days 8, 22, and 43 from baseline were correlated with progression risk. Results VEGF increased and sVEGFR-2 decreased by day 43 in the vandetanib arm, whereas a distinct pattern was observed in the CP and VCP arms, with significant decreases in interleukin (IL) -12, IL-1 receptor antagonist, and matrix metalloproteinase 9 (MMP-9) and increased macrophage chemoattractant protein 1. In each treatment arm, changes in different markers were associated with progression risk. For example, increases in IL-8 with VCP, MMP-9 with CP, and VEGF with vandetanib monotherapy were associated with increased progression risk, and increase in intercellular adhesion molecule 1 with vandetanib was associated with decreased risk. Conclusion Vandetanib and chemotherapy treatment led to distinct patterns of CAF changes; the combination resembled chemotherapy alone. Changes in specific CAFs correlated with clinical outcome, but markers differed for each treatment arm. CAF profiling may provide insights into the biologic effects of treatment and identify drug-specific markers of activity and clinical benefit.

CD26 is an antigen with key role in T-cell biology and is expressed on selected subsets of aggressive T-cell malignancies. To elucidate the role of CD26 in tumor behavior, we examine the effect of CD26 depletion by small interfering RNA transfection of T-anaplastic large cell lymphoma Karpas 299. We show that the resultant CD26-depleted clones lose the ability to adhere to fibronectin and collagen I. Because anti–integrin β1 blocking antibodies also prevent binding of Karpas 299 to fibronectin and collagen I, we then evaluate the CD26-integrin β1 association. CD26 depletion does not decrease integrin β1 expression but leads to dephosphorylation of both integrin β1 and p38 mitogen-activated protein kinase (MAPK). Moreover, our data showing that the p38MAPK inhibitor SB203580 dephosphorylates integrin β1 and that binding of the anti-CD26 antibody 202.36 dephosphorylates both p38MAPK and integrin β1 on Karpas 299, leading to loss of cell adhesion to the extracellular matrix, indicate that CD26 mediates cell adhesion through p38MAPK-dependent phosphorylation of integrin β1. Finally, in vivo experiments show that depletion of CD26 is associated with loss of tumorigenicity and greater survival. Our findings hence suggest that CD26 plays an important role in tumor development and may be a novel therapeutic target for selected neoplasms.

CD26 is a Mr 110,000 surface-bound glycoprotein with diverse functional properties, including having a key role in normal T-cell physiology and the development of certain cancers. In this article, we show that surface expression of CD26, especially its intrinsic dipeptidyl peptidase IV (DPPIV) enzyme activity, results in enhanced topoisomerase IIα level in the B-cell line Jiyoye and subsequent in vitro sensitivity to doxorubicin-induced apoptosis. In addition, we show that expression of CD26/DPPIV is associated with increased phosphorylation of p38 and its upstream regulators mitogen-activated protein kinase kinase 3/6 and apoptosis signal-regulating kinase 1 and that p38 signaling pathway plays a role in the regulation of topoisomerase IIα expression. Besides demonstrating that CD26 effect on topoisomerase IIα and doxorubicin sensitivity is applicable to cell lines of both B-cell and T-cell lineages, the potential clinical implication of our work lies with the fact that we now show for the first time that our in vitro results can be extended to a severe combined immunodeficient mouse model. Our findings that CD26 expression can be an in vivo marker of tumor sensitivity to doxorubicin treatment may lead to future treatment strategies targeting CD26/DPPIV for selected human cancers in the clinical setting. Our article thus characterizes the biochemical linkage among CD26, p38, and topoisomerase IIα while providing evidence that CD26-associated topoisomerase IIα expression results in greater in vitro and in vivo tumor sensitivity to the antineoplastic agent doxorubicin.

Lactoferrin has been identified as a factor in human colostrum that accounts for increased incorporation of thymidine into the DNA in an in vitro rat crypt enterocyte bioassay. We have examined lacatoferrin-stimulated thymidine incorporation by comparing the effects of ironfree lactoferrin (apolactoferrin) with those of iron-saturated lactoferrin (diferric lactoferrin) under conditions that inhibit the transfer of iron between these iron-binding proteins in the bioassay system. In addition, we have compared the dose-response relationships of diferric lactoferrin and apolactoferrin. The results demonstrated that lactoferrin, independent of iron-binding states, promoted the incorporation of thymidine into the DNA of rat crypt enterocytes. These observations suggest a previously unreported nutritional role for lactoferrin that is independent of its iron-binding capacity.

362 Background: Cediranib is a highly potent and selective signaling inhibitor of vascular endothelial growth factor (VEGF) receptors−1, −2, −3 and c-Kit. In a phase II trial, patients (pts) with mRCC were randomized 3:1 to treatment with cediranib or placebo to primarily assess the difference in change in tumor size at 12 weeks. Cediranib resulted in a significant reduction in tumor size vs. placebo (Mulders et al. ESMO. 2009). Here we investigated soluble biomarkers in serum and concentration changes during treatment. Methods: Pts received cediranib 45 mg/day PO or placebo for the first 12 weeks on treatment. At that time (or upon progression if earlier), treatment was unblinded and pts on placebo were given the option of receiving cediranib. Serum was collected from 61 pts at baseline (BL; cediranib 46, placebo 15), from 50 pts on day (D) 28, and from 45 pts on D84. Multiplex bead suspension arrays and ELISA were used to measure CAF concentrations including VEGF, placental growth factor (PlGF), sVEGFR-2, PDGFbb, HGF, MMP-9, multiple chemokines and interleukins (IL). The objectives of this exploratory analysis were to assess whether baseline CAFs were associated with changes in tumor size after 12 weeks of treatment and whether CAF levels changed while on treatment. Results: Of 39 CAFs available at BL, lower than median (‘low') concentrations of IL-10, VEGF, PlGF, stem cell factor (SCF), and monokine-induced by interferon-gamma (MIG) were associated with larger decreases in tumor size than high concentrations (independent of treatment arm), whereas the opposite trend was observed for IL-5 and TRAIL. Consistent with inhibition of VEGF signaling, cediranib treatment resulted in changes over time in VEGF (increase), sVEGFR-2 (decrease) and PlGF (increase) concentrations. In addition, SCF, a c-Kit ligand and M-CSF decreased, while VCAM-1 and TRAIL increased during cediranib treatment. Conclusions: Candidate CAFs associated with response to cediranib in mRCC such as IL-10 and VEGF were identified. Patients treated with cediranib showed distinct CAF changes compared with placebo. These results require independent validation in a larger trial. [Table: see text]

14687 Background: Recent studies suggest that specific subpopulations of monocytes and myeloid progenitors, particularly those expressing VEGFR-1, play a key role in promoting tumor angiogenesis an...

In a search for dietary factors that might stimulate enterocyte proliferation, we developed an assay for thymidine incorporation into DNA using harvested crypt cells from mature rat small intestine. Human colostrum stimulated a significant increase in thymidine incorporation into rat crypt cell DNA during a 60-min period of incubation. When the protein with biological activity was purified to a single peak by sequential ion exchange and gel filtration chromatography, it was found to have the characteristics of lactoferrin. The protein was identical to lactoferrin standards by sodium dodecyl sulfate polyacrylamide gel electrophoresis, isoelectric focusing, and doublediffusion immunologic precipitation. All available human lactoferrins stimulated thymidine uptake and all reacted with a lactoferrin polyclonal antibody. Human lactoferrin appears to be a potent activator of thymidine incorporation into DNA in incubated rat crypt cells, a nutritional function not previously reported.

Lactoferrin has been identified as the factor in human colostrum that accounts for the increased incorporation of thymidine into DNA in a rat crypt enterocyte bioassay. Commercially available infant formulas used in the refeeding of infants who have severe dietary intolerance associated with mucosal atrophy were tested in this bioassay. In contrast to human milk, no stimulation was observed with these formulas in the assay. All formulas inhibited basal thymidine incorporation. The degree of inhibition of the basal assay progressed from 14% with cow's-milk formula to 30% with soy-based formulas to 45% with hydrolyzed casein formulas. When the formulas were supplemented with lactoferrin to match the level of that in human milk, the result was a 35% relative increase in thymidine incorporation into DNA. The quantitative level of response was primarily influenced by the baseline inhibitions associated with the specific formulas. The most profound effect was observed with the hydrolyzed casein formula, when lactoferrin supplementation reversed the baseline inhibition but did not result in a response greater than that seen in control basal bioassays. The supplementation of infant formulas with lactoferrin improved the thymidine incorporation in rat crypt enterocytes; a response equivalent to that of human milk, however, would require a redesign of other formula components to reduce basal inhibition.