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Lizards of the clade Iguanidae (sensu lato) are primarily a New World group. Thus, the remarkable presence of an endemic lineage of iguanas (family Opluridae) on the isolated Indian Ocean island of Madagascar has long been considered a biogeographic anomaly. Previous work attributed this disjunct extant distribution to: (1) vicariance at about 140-165 Ma, caused by the breakup of Gondwana and the separation of South America, Africa, and Madagascar (with subsequent extinction of iguanas on Africa, and potentially other Gondwanan landmasses), (2) vicariance at about 80-90 Ma, caused by the sundering of hypothesized land-bridge connections between South America, Antarctica, India, and Madagascar, or (3) long-distance overwater dispersal from South America to Madagascar. Each hypothesis has been supported with molecular divergence dating analyses, and thus the biogeographic origin of the Opluridae is not yet well resolved. Here we utilize genetic sequences of ultraconserved elements for all Iguania families and the majority of Iguanidae (s.l.) genera, and morphological data for extant and fossil taxa (used for divergence dating analyses), to produce the most comprehensive dataset applied to date to test these origin hypotheses. We find strong support for a sister relationship between the Opluridae (Madagascar) and Leiosauridae (South America). Divergence of the Opluridae from Leiosauridae is dated to between the late Cretaceous and mid-Paleogene, at time when Madagascar was already an island and was isolated from all other Gondwanan landmasses. Consequently, our results support a hypothesis of long-distance overwater dispersal of the Opluridae lineage, either directly from South America to Madagascar or potentially via Antarctica or Africa, leading to this radiation of iguanas in the Indian Ocean.

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A33, a monoclonal antibody that targets colon carcinomas, was labeled with (125)I or (131)I and the relative therapeutic efficacy of the 2 radiolabeled species was compared in a human colon cancer xenograft system.Nude mice bearing human SW1222 colon carcinoma xenografts were administered escalating activities of (125)I-A33 (9.25-148 MBq) or (131)I-A33 (0.925-18.5 MBq), (125)I- and (131)I-labeled control antibodies, unlabeled antibody, or no antibody. The effects of treatment were assessed using the endpoints of tumor growth delay and cure.Tumor growth delay increased with administered activity for all radiolabeled antibodies. Approximately 4.5 times more activity was required for (125)I-A33 to produce therapeutic effects that were equivalent to those of (131)I-A33. This ratio was approximately 7 for a nonspecific, noninternalizing isotype-matched, radiolabeled control antibody. Unlabeled A33 antibody had no effect on tumor growth. Approximately 10 times more activity of (125)I-A33 produced toxicity similar to that of (131)I-A33, and this ratio fell to approximately 6 for radiolabeled control antibody.Treatment with (125)I-A33 resulted in a relative therapeutic gain of approximately 2 compared with (131)I-A33 in this experimental system.

Radiolabeling monoclonal antibodies (mAbs) allows the evaluation of biodistribution of constructs in vivo through gamma camera imaging and also permits quantitation of mAb uptake in tumors through biopsy-based counting techniques. The quantitation of radiolabeled mAb uptake in cancer patients is complicated by the attenuation of gamma emissions of routinely used isotopes (e.g., 131I and 111In) and the spatial resolution and sensitivity of gamma cameras.We used the positron-emitting isotope 124I (half-life [T1/2] = 4.2 d) to label the recombinant humanized anti-colorectal cancer A33 antibody (huA33) and evaluated its biodistribution properties and PET imaging characteristics in BALB/c nude mice bearing SW1222 colorectal xenografts and control colon tumors.The immunoreactivity of radioconjugate was 78% as determined using the cell-binding Lindmo assay. The apparent association constant was found to be 2.2 x 10(9) M(-1), and the number of antibody binding sites per cell was 371,000. The radioconjugate was found to be stable in serum obtained from mice at various times after injection. Assuming a two-compartment model with a four-parameter fit of mean blood levels, the T1/2alpha was 1.5 h and the T1/2beta was 38.2 h. Excellent tumor uptake was obtained, with maximal uptake reaching 50.0 +/- 7.0 percentage injected dose per gram of tumor by 4 d after injection. Specificity of localization was shown by lack of uptake in control tumor. PET imaging detected antigen-positive tumor by 4 h after injection, and high-resolution images were obtained by 24 h after injection.In clinical trials using PET, huA33 labeled with 124I has potential for imaging and staging colon tumors and quantifying antibody uptake in colon tumors in vivo.

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Most available pancreas cancer cell lines have been in culture for long periods of time, have not been extensively characterized from the cell biology standpoint, or lack differentiated properties.We have established four new cell lines from ductal pancreatic cancers (IMIM-PC-1, IMIM-PC-2, SK-PC-1, and SK-PC-3). The phenotype and functional properties of the cell lines were analyzed using ultrastructural methods, antibodies detecting cytokeratin polypeptides and mucin epitopes, and cDNA probes of epithelial differentiation markers.IMIM-PC-2 and SK-PC-1 cells grow as a polarized monolayer, form domes, and express all CK polypeptides typical of simple epithelia and the MUC1 mucin. IMIM-PC-1 and SK-PC-3 are morphologically less differentiated and express low or undetectable levels of CK7 and MUC1. By Northern blotting, we found that SK-PC-1 and SK-PC-3 cells express carbonic anhydrase II and that the cystic fibrosis transmembrane regulator was undetectable in the four lines. Secretin induces a marked stimulation of cAMP levels in all cell lines except for SK-PC-3. Cytogenetic analysis demonstrates their human origin.Levels of CK7 and MUC1 are associated with less differentiated cultures. The new cell lines should be useful tools to study the cell biology of exocrine pancreas cancer.

The chimeric monoclonal antibody KM871, directed against the G(D3) antigen, is under evaluation for its potential to target melanoma. To facilitate the in vivo evaluation of biodistribution properties and measurement of pharmacokinetics, KM871 was radiolabeled with (125)I via tyrosine residues and with (111)In via the bifunctional metal ion chelator C-functionalized trans-cyclohexyl diethylenetriaminepentaacetic acid (CHX-A"-DTPA) to lysine residues. Using antigen-positive SK-MEL-28 melanoma cells, immunoreactivities of 42 and 40% cell binding were obtained, respectively, for the two radioconjugates. Binding was enhanced in the presence of added unlabeled antibody. A humanized A33 antibody was similarly labeled with the two isotopes and used as a control. To determine and compare in vivo biodistribution characteristics of KM871 radiolabeled with (111)In or (125)I, mixtures of the radioconjugates were injected i.v. into BALB/c nude mice bearing G(D3)-positive-SK-MEL-28 melanoma xenografts. Gamma camera images were acquired; groups of five mice were sacrificed at various time intervals, and tumors, blood, and tissues were analyzed. (111)In-labeled CHX-A"-DTPA-KM871 showed a maximum tumor uptake of 41.9 +/- 7.0% injected dose/g at 72 h with prolonged retention over a 15-day period. The tumor:blood ratio was 3:1 by 72 h, and higher ratios were observed at later time points. No abnormal accumulation of (111)In-labeled conjugate was found in normal tissues. In contrast, there was little accumulation of (125)I-labeled KM871 in the same tumors. The specificity of antibody localization was confirmed by the low tumor uptake values for radiolabeled control antibody. Gamma camera imaging demonstrated excellent uptake of (111)In-labeled CHX-A"-DTPA-KM871 in the xenografts. Chromatographic analyses of xenograft cytosolic extracts demonstrated tumor internalization and catabolism of radiolabeled KM871 with the formation of small molecular weight metabolites. Laser scanning confocal microscopy demonstrated that the majority of intracellular KM871 is localized to lysosomes. Despite the catabolism of the radioconjugate, a dose-dependent increase in KM871 tumor localization was shown through immunohistochemical examination of xenograft biopsies. This study demonstrates for the first time the in vivo localization of a radiolabeled anti-G(D3) monoclonal antibody to G(D3)-expressing xenografts using gamma camera scanning techniques and tumor cell internalization of KM871 tagged with a green fluorescent dye, Alexa Fluor 488, through confocal microscopy. KM871 has potential for targeting tumors in patients with melanoma.

Mouse mAb M111 identifies a cell surface glycoprotein of 115,000 to 135,000 Da. M111 was expressed constitutively in subsets of cells of multiple lineages at discrete stages of cell maturation, suggesting that M111 is a differentiation Ag of the three germ layers. Ag expression could be induced by IFN-gamma but not by IFN-alpha, IFN-beta, or TNF. Induction of M111 expression was maximal at 48 h of culture in 200 U/ml of IFN-gamma and was independent of induction of class II MHC Ag. Induction was dependent on the cell type used. Nine colon cancer cell lines of undifferentiated phenotype were constitutively M111-; IFN-gamma induced M111 expression in seven of them. In contrast, IFN-gamma failed to induce M111 expression in six of six M111- ovarian cancer cell lines. Eight normal fibroblast cultures tested were M111-; they could not be induced to express M111. Three of five sarcoma cell lines were M111+; culture in IFN-gamma induced an increase in M111 expression in all of them. Constitutive and IFN-gamma-induced expression of M111 was independent of constitutive and induced expression of HLA class I and II molecules. IFN-gamma-mediated induction of M111 expression was not accompanied by coordinate changes in the expression of other differentiation traits. These results suggest that expression of the M111 gene is controlled by two mechanisms, one related to differentiation and the other activated by IFN-gamma.