New targets are required for the control of advanced metastatic disease. We investigated the use of cadherin RGD motifs, which activate the α2β1integrin pathway, as targets for the development of therapeutic monoclonal antibodies (mAb). Cadherin 17 (CDH17) fragments and peptides were prepared and used for immunization and antibody development. Antibodies were tested for inhibition of β1 integrin and cell adhesion, proliferation, and invasion assays using cell lines from different cancer types (colorectal, pancreatic, melanoma, and breast cancer). Effects of the mAbs on cell signaling were determined by Western blot analysis. Nude mice were used for survival analysis after treatment with RGD-specific mAbs and metastasis development. Antibodies against full-length CDH17 failed to block the binding to α2β1 integrin. However, CDH17 RGD peptides generated highly selective RGD mAbs that blocked CDH17 and vascular-endothelial (VE)-cadherin-mediated β1 integrin activation in melanoma and breast, pancreatic, and colorectal cancer cells. Antibodies provoked a significant reduction in cell adhesion and proliferation of metastatic cancer cells. Treatment with mAbs impaired the integrin signaling pathway activation of FAK in colorectal cancer, of JNK and ERK kinases in colorectal and pancreatic cancers, and of JNK, ERK, Src, and AKT in melanoma and breast cancer. , RGD-specific mAbs increased mouse survival after inoculation of melanoma and colorectal cancer cell lines to cause lung and liver metastasis, respectively. Blocking the interaction between RGD cadherins and α2β1 integrin with highly selective mAbs constitutes a promising therapy against advanced metastatic disease in colon cancer, melanoma, and, potentially, other cancers. .

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The leishmanioses, vector-borne diseases caused by the trypanosomatid protozoan Leishmania, are transmitted to susceptible mammals by infected phlebotomine sand flies that inoculate promastigotes into hemorrhagic pools created in host skin. We assumed that promastigotes are delivered to a blood pool, and analyzed early promastigote interactions (0–5 min) with host components, which lead to parasite endocytosis by blood leukocytes, and to host infection. Promastigotes were incubated with NHS or with heparinized blood in near-physiological conditions, and we used cell radioimmunoassay and flow cytometry to measure the on-rate constants (k+1) of promastigote interactions with natural opsonins and erythrocytes. We obtained quantitative data for parasitized cells to determine the time-course of promastigote binding and internalization by blood leukocytes. In these reactions, promastigotes bind natural opsonins, immune adhere to erythrocytes and activate complement cytolysis, which kills ∼95% of promastigotes by 2 min post-infection. C3-promastigote binding is a key step in opsonization; nascent C3-promastigotes are the substrate for two simultaneous reactions, C3-promastigote immune adherence (IA) to erythrocytes and complement-mediated promastigote killing. The k+1 for IA was 75-fold greater than that for promastigote killing, showing that IA facilitates promastigote endocytosis and circumvents lysis. At 5 min post-infection, when reaction velocity is still linear and promastigote concentration is not limiting, 17.4% of granulocytes and 10.7% of monocytes had bound promastigotes, of which ∼50% and ∼25%, respectively, carried surface-bound (live) or internalized (live and dead) leishmanias. Of other leukocyte types, 8.5% of B cells bound but did not internalize promastigotes, and T cells, NK cells and CD209+ dendritic cells did not bind parasites. These data show that, once in contact with blood, promastigote invasion of human leukocytes is an extremely rapid and efficient reaction, and suggest that the IA reaction constitutes a central strategy for this parasite in subverting host innate immune defenses.