Biomedical subjects

T Tsuruo

Publications and source records attributed to T Tsuruo.

At least 217 records · Page 12Linked to original sources

PubMed1995

Cysteine produced from lymph node stromal cells suppresses apoptosis of mouse malignant T-lymphoma cells.

Mouse malignant T-lymphoma CS-21 cells can grow when cocultured with CA-12 lymph node stromal cells, but they undergo apoptotic cell death with DNA fragmentation when separated from CA-12 stromal cells. In the course of examining the effects of the soluble factor(s) secreted by CA-12 stromal cells on CS-21 cell growth, we found that cysteine produced from CA-12 stromal cells promoted CS-21 cell growth and suppressed apoptosis in the isolated culture of CS-21 cells. The expression of anti-apoptotic protein Bcl-2, however, was not induced by cysteine. These results indicate that cysteine produced from CA-12 stromal cells participates not only in the cell growth but also in the inhibition of CS-21 apoptosis without inducing the expression of Bcl-2 protein.

Animals↗

PubMed1995

Novel dithiane analogues of tiapamil with high activity to overcome multidrug resistance in vitro.

Dithiane analogues of tiapamil are highly active as modifiers of P-glycoprotein mediated multidrug resistance (MDR) in vitro. In an assay using the P-glycoprotein over-expressing cell line KB-8-5, the most active analogues for decreasing vincristine resistance were the racemate Ro 11-5160 and its two enantiomers, Ro 44-5911 (R) and Ro 44-5912 (S). In the KB-8-5 assay, the resistance modification index (RMI) of Ro 11-5160 was approximately 12-fold higher than those of the most active reference compounds tested, dipyridamole, cepharanthine, reserpine and cyclosporin A, when compared at concentrations equal to one-tenth of the IC50 of each compound (RMI0.1). The enantiomers have similar resistance modifying activities, but the (S) enantiomer Ro 44-5912 is somewhat more active, fully reverting the vincristine sensitivity of KB-8-5 cells to the level of the parental KB-3-1 cells at a concentration of 2 microM. The (R) enantiomer attained this level of modification at a concentration of 3.5 microM. These concentrations are both well below their IC50 values for KB-8-5 cells (150 microM). The enantiomers appear to interact with P-glycoprotein because they inhibited [3H]azidopine and [3H]-vinblastine binding to plasma membrane fractions prepared from resistant K562/ADR cells. However, in addition to their resistance modifying activities with KB-8-5 cells, these compounds also decreased the IC50 values of vincristine and doxorubicin with KB-3-1 cells that do not express detectable levels of P-glycoprotein. Ro 44-5911 overcame doxorubicin and vincristine resistance in three colorectal cancer cell lines (DLD-1, WiDr and COLO 201) that express P-glycoprotein. No effect was seen with the 3 colorectal cell lines on the IC50 values of three drugs not related to the MDR phenotype, 5-fluorouracil, 5'-deoxy-5-fluorouridine and cis-diaminodichloroplatinum (II). The in vitro vasodilatory activity of these dithianes, measured with strips of rat aorta contracted with KCl, was about 5% of that of verapamil. These results suggest that diathianes could be useful agents for MDR modification in vivo.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

PubMed1995

Binding diversity of antibodies against external and internal epitopes of the multidrug resistance gene product P-glycoprotein.

P-glycoprotein (Pgp) is a trans-membraneous protein that is associated with multidrug resistance (MDR) in human cancer, including hepatocellular carcinomas and leukemias. There is no consensus regarding methods of choice for analysis of Pgp expression, and development of reliable analytical methods is now essential. We have studied the the Pgp expression in human hepatoma and leukemia cell lines using flow cytometry. The aim of the study was to compare binding properties of anti-Pgp antibodies reacting with surface (MRK16, UIC2) and cytoplasmic (C219, JSB-1) epitopes to assess which antibody performed best with respect to fluorescence discrimination. By histogram subtraction the fractions of resistant human hepatoma cells positive for Pgp were 99% (MRK16), 97% (UIC2), 77% (JSB-1), and 51% (C219), demonstrating variations in antibody reactivity. The resolution in detecting decreasing levels of Pgp in hepatoma cells was superior for the externally binding antibodies, showing that there is a correlation between antibody reactivity and fluorescence discrimination. Similar results were obtained for parental and resistant KG1a human leukemia cell lines. The Pgp epitopes remained reactive to the anti-Pgp MAbs after methanol fixation and cryopreservation. By dual parameter flow cytometry it was shown that Pgp expression in viable cells may be assessed together with uptake of epirubicin, which was low in cells expressing high levels of Pgp and vice versa. In conclusion, all tested antibodies proved useful for flow cytometric detection of high levels of Pgp, but the externally binding ones were superior in detection of low and variable levels of Pgp.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

PubMed1995

Involvement of CD45 in adhesion and suppression of apoptosis of mouse malignant T-lymphoma cells.

Mouse malignant T-lymphoma CS-21 cells undergo apoptotic cell death in vitro in the absence of lymph node stromal cells but escape apoptosis and proliferate when they are attached to CA-12 stromal cells. A monoclonal antibody raised against CS-21 cell surface molecules (MCS-5) recognized a M(r) 168,000 protein, inhibited binding of CS-21 cells to CA-12 stromal cells, and suppressed apoptosis in CS-21 cells. To identify the M(r) 168,000 protein, we purified it with MCS-5 affinity chromatography and ion exchange chromatography. Partial amino acid sequences of the purified M(r) 168,000 protein were identical to those of CD45, a transmembrane tyrosine phosphatase. The purified protein possessed tyrosine phosphatase activity and was recognized by an anti-CD45 monoclonal antibody. The M(r) 168,000 protein was identified as CD45. To determine the CD45 isoform, we cloned the CD45 gene from the cDNA library of CS-21. Sixteen of 18 clones encoded CD45RO (CD45 lacking exons 4, 5, and 6), and the remainder lacked exons 4, 5, 6, and 7. Like MCS-5, an anti-CD45 monoclonal antibody, also inhibited binding of CS-21 cells to CA-12 cells and suppressed apoptosis in CS-21 cells. Our present results indicate that CD45RO expressed on CS-21 cells mediates adhesion to CA-12 cells and suppression of apoptosis.

Amino Acid Sequence↗

PubMed1995

Cloning, expression and chromosomal localization of a novel gene for protein tyrosine phosphatase (PTP-U2) induced by various differentiation-inducing agents.

Previously, we cloned two gene fragments encoding novel protein tyrosine phosphatases, termed PTP-U1 and PTP-U2. Here, we report the full-length sequence, expression, and chromosomal localization of the PTP-U2 gene. The cDNA for PTP-U2, which was obtained from a human normal kidney library, predicts a protein of 1216 amino acids, -140 kDa, that contains a single transmembrane domain and a single intracellular catalytic domain. The extracellular domain of PTP-U2 contains 14 putative N-glycosylation sites and eight repeats of fibronectin type III-like motif. These data suggest that PTP-U2 is structurally similar to HPTP beta and DPTP10D, which have been reported previously. Northern blot analysis revealed that there were two different transcripts for PTP-U2. In kidney and brain, gene expression of PTP-U2 was detected as a 5.4 kb mRNA and in lung and placenta as 3.5 kb. The 3.5 kb transcript was also detected in human leukemia cell lines (eg., U937). Interestingly, its gene expression was enhanced by various differentiation-inducing agents, such as phorbol ester, dihydroxy vitamin D3, retinoic acid, and dimethyl sulfoxide. The bacterially expressed PTP-U2 fusion protein exhibited intrinsic tyrosine phosphatase activity. The PTP-U2 gene was assigned to chromosome 12p13.2-p13.3.

Amino Acid Sequence↗

PubMed1995

Aspartate-based inhibitor of interleukin-1 beta-converting enzyme prevents antitumor agent-induced apoptosis in human myeloid leukemia U937 cells.

We found that a novel protease inhibitor, benzyloxycarbonyl-Asp-CH2OC(O)-2,6-dichlorobenzene (Z-Asp-CH2-DCB), which can preferentially inhibit interleukin-1 beta-converting enzyme (ICE), completely blocked the apoptotic cell death of human myeloid leukemia U937 cells caused by etoposide, camptothecin, 1-beta-D-arabinofuranosyl-cytosine and Adriamycin, as well as TNF-alpha, anti-Fas antibody and staurosporine. However, Z-Asp-CH2-DCB did not block non-apoptotic cell death of U937 cells caused by etoposide during prolonged incubation periods. These results indicate that ICE or ICE-like proteases inhibited by Z-Asp-CH2-DCB are involved in a common pathway of apoptotic cell death in U937 cells.

Antineoplastic Agents↗

PubMed1995

Purification and characterization of the platelet-aggregating sialoglycoprotein gp44 expressed by highly metastatic variant cells of mouse colon adenocarcinoma 26.

A platelet-aggregating sialoglycoprotein with a molecular weight of 44,000 (gp44) was immunochemically purified from highly metastatic mouse adenocarcinoma cells. The rat monoclonal antibody (mAb) 8F11 used in the purification procedure has been generated previously against NL-17 cells derived from the mouse colon 26 cell line, mAb 8F11 inhibits NL-17 cells from inducing platelet aggregation and suppresses their experimental metastasis to the lung. The purified gp44 induced mouse platelet aggregation in a dose-dependent manner without any plasma component. This aggregation was completely inhibited by mAb 8F11. The gp44 was partially characterized by sequential enzymatic hydrolysis of carbohydrates and was found to be O-glycans enriched. When gp44 was sequentially treated with N-glycanase and neuraminidase, it lost platelet aggregation activity. Further treatment with O-glycanase resulted in a loss of the reactivity to mAb 8F11. These results suggest that sialylated carbohydrate chains of gp44 are involved in the induction of platelet aggregation and may play an important role in the colonization of NL-17 cells in the lung.

Adenocarcinoma↗

PubMed1995

Induction of resistance to 1-beta-D-arabinofuranosylcytosine in human H9 cell line by simian immunodeficiency virus.

We examined drug sensitivity of human T cell acute lymphoblastic leukemia H9 cells chronically infected with simian immunodeficiency virus (SIVmac) and found that the retrovirus-infected H9 cells showed 8.2-fold resistance to 1-beta-D-arabinofuranosylcytosine (Ara-C). In the infected cells, Ara-CTP levels decreased to 20% of that found in uninfected H9 cells after 3 h incubation at Ara-C concentration of 1 microM, and 8.1-fold increase of cytidine deaminase activity was observed in the infected H9 cells. A competitive inhibitor of cytidine deaminase, 3, 4, 5, 6-tetrahydrouridine (THU), at 100 microM reversed Ara-C resistance in the infected cells. These results indicate that inducing increased cytidine deaminase activity by SIVmac infection conferred Ara-C resistance to H9 cells. An understanding of these cellular differences in drug sensitivity may aid in the development of therapeutic strategies against retrovirus-infected cells.

Cell Division↗

PubMed1995

Human melanoma invasion and metastasis enhancement by high expression of aminopeptidase N/CD13.

Aminopeptidase N/CD13 is a Zn(2+)-dependent exoprotease present on the cell surface as a transmembrane protein. Our previous studies using aminopeptidase inhibitors and antibodies demonstrated that aminopeptidase N is involved in the degradation and invasion of the extracellular matrix (ECM) by metastatic tumor cells. In the present study we transfected human A375M melanoma cells with eukaryotic plasmid expression vectors that contained full length cDNA of aminopeptidase N/CD13 and examined their characteristics. The transfectants that expressed extremely high levels of aminopeptidase N/CD13 degraded type IV collagen and invaded ECM more actively than the parental and control vector-transfected cells. Furthermore, the aminopeptidase N/CD13-transfected A375M cells had significantly augmented lung colonizing potential in nude mice. The results show that the aminopeptidase N/CD13 plays an active role in degradation and invasion of ECM and may be involved in the molecular mechanisms of blood-borne metastasis.

Animals↗

PubMed1995

H-31 human breast cancer cells stimulate type I collagenase production in osteoblast-like cells and induce bone resorption.

Bone is one of the most common sites of metastasis in breast cancer. For metastasis to occur in bone, tumor cells must induce osteolysis by osteoclasts. Degradation of the osteoid layer by type I collagenase is a necessary process before osteolysis can occur because the osteoid layer hinders osteoclasts from adhering to bone. In this study, we investigated the function of H-31 human breast cancer cells in inducing type I collagenase production and in enhancing bone resorption. H-31 cells did not themselves produce type I collagenase whereas MG-63 human osteoblast-like cells and MC3T3-E1 mouse osteoblast cells constantly produced type I collagenase. When these osteoblast-like cells were cocultured with H-31 cells, type I collagenase production was enhanced. The same enhancement occurred when the conditioned medium of H-31 cells was added to the osteoblast-like cells. The activity of this type I collagenase was inhibited by EDTA and minocyclin, an inhibitor of matrix metalloproteinases, hence it was identified as matrix metalloproteinase-1 (MMP-1). H-31 cells exhibited chemotactic migration towards collagen; therefore, collagen degraded by MMP-1 may play an important role in the localisation of breast cancer cells like H-31 to bone. In an organ culture system using newborn mouse calvaria, the conditioned medium of H-31 cells increased the concentration of calcium in the medium, and this effect was inhibited by minocyclin, indicating that bone resorption occurred in this system. Based on these observations, we speculate that type I collagenase produced by osteoblast cells in response to breast cancer cells (exemplified by H-31) may facilitate degradation of the osteoid layer and the homing of breast cancer cells to bone. This can lead to osteolysis by osteoclasts, a crucial event for bone metastasis.

Bone Resorption↗

PubMed1995

Delivery and cytotoxicity of RS-1541 in St-4 human gastric cancer cells in vitro by the low-density-lipoprotein pathway.

RS-1541 is a 13-O-palmitoyl derivative of rhizoxin, an inhibitor of tubulin polymerization. RS-1541 has been shown to bind preferentially to plasma lipoproteins and to exhibit selective and sustained uptake by tumors in mice. To elucidate a mechanism of RS-1541 cytotoxicity, the cellular uptake and the cytotoxicity of a complex of RS-1541 with human low-density lipoprotein (RS-1541/LDL complex) were investigated in cultured St-4 human gastric cancer cells. Both the cellular uptake and the cytotoxicity of the RS-1541/LDL complex were greater in cells with higher LDL-receptor activities than in control cells. Excess amounts of LDL or 1 microM of monensin, a proton ionophore, significantly inhibited both the uptake and the cytotoxicity of the complex. Chloroquine, an inhibitor of lysosomal enzymes, decreased the intracellular level of rhizoxin liberated from RS-1541 and suppressed the cytotoxicity of the RS-1541/LDL complex. However, a detergent-aided solution of RS-1541 showed very low cellular uptake and cytotoxicity, irrespective of the LDL-receptor activities of these cells. These results demonstrate that the RS-1541/LDL complex is incorporated into the cells via the LDL receptor and that it manifests its cytotoxic activity after forming rhizoxin, the original antitumor agent, in the lysosomes.

Antibiotics, Antineoplastic↗

PubMed1995

Relationship between multidrug resistant gene expression and multidrug resistant-reversing effect of MS-209 in various tumor cells.

MS-209 is a novel quinoline compound which can overcome multidrug resistance (MDR) both in vitro and in vivo, while having a low level of side effects, and is now being evaluated in a clinical phase II study. Reverse transcription-polymerase chain reaction (RT-PCR) was used to quantitate the expression levels of MDR genes in various mouse and human tumor cell lines. The MDR gene and the beta actin gene, as the internal reference standard, were coamplified separately, and the relative expression of the MDR gene was represented by the MDR/beta actin ratio. The in vitro MDR-reversing effect of MS-209 was then compared with the MDR gene expression (MDR/beta actin ratio). We found a significant correlation between these two parameters. Moreover, a significant correlation was also observed between the level of expression of the MDR1 gene and that of P-glycoprotein in human cell lines. Therefore, the efficacy of MS-209 seems to specifically depend on the level of MDR gene expression (P-glycoprotein). From these observations, it is suggested that RT-PCR assays of MDR1 gene in tumor biopsy specimens might be an effective means to predict the response of tumor cells to combination therapy with MS-209.

Animals↗

PubMed1995

Reversal of multidrug resistance by a novel quinoline derivative, MS-209.

MS-209, a novel quinoline derivative, was examined for its reversing effect on multidrug-resistant tumor cells. MS-209 at 1-10 microM completely reversed resistance against vincristine (VCR) in vitro in multidrug-resistant variants of mouse leukemia P388 cells (VCR-resistant P388/VCR and Adriamycin (ADM)-resistant P388/ADM) and human leukemia K562 cells (VCR-resistant K562/VCR and ADM-resistant K562/ADM). MS-209 at 1-10 microM also completely reversed resistance against ADM in vitro in P388/VCR cells, K562/VCR cells, and K562/ADM cells. In ADM-resistant P388 (P388/ADM) cells, however, ADM resistance was only partially reversed at the MS-209 concentrations tested. MS-209 enhanced the chemotherapeutic effect of VCR in P388/VCR-bearing mice. When MS-209 was given p.o. at 80 mg/kg twice a day (total dose, 160 mg/kg per day) with 100 micrograms/kg VCR, a treated/control (T/C) value of 155% was obtained. MS-209 also enhanced the chemotherapeutic effect of ADM in P388/ADM-bearing mice. The most prominent effects were obtained when MS-209 was given with 2 mg/kg ADM, yielding T/C values of 150%-194% for the combined treatment at an MS-209 dose of 200-450 mg/kg. MS-209 inhibited [3H]-azidopine photolabeling of P-glycoprotein efficiently. Furthermore, the accumulation of ADM in K562/ADM cells was increased more efficiently by MS-209 than by verapamil. These results indicate that MS-209, like verapamil, directly interacts with P-glycoprotein and inhibits the active efflux of antitumor agents, thus overcoming multidrug resistance in vitro and in vivo.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

PubMed1995

Intracellular activation and cytotoxic action of RS-1541 against cultured human tumor cells.

RS-1541, an acyl-derivative of rhizoxin (Fig. 1), is a potent antitumor compound. This agent showed cytotoxicity in vitro on some cultured human tumor cells, although it was less potent than rhizoxin. Rhizoxin exhibited antitumor effects by inhibiting the polymerization of tubulin, whereas RS-1541 did not inhibit tubulin polymerization in vitro. However, cell cycle analysis in vivo showed that the two agents had the same mode of action. The cytotoxicity of RS-1541 was enhanced when the initial cell density of the cells was increased. The cytotoxicity was also enhanced when the membrane fraction of St-4 cells, which were the most sensitive to RS-1541 among the cell lines tested, was added to the target cells. When St-4 cells were incubated with [14C]-RS-1541, significant amounts of [14C]-rhizoxin were produced within the cells. Further fractionation of the crude membrane showed that the activity that enhanced the cytotoxicity of RS-1541 (RS-1541-enhancing activity) belonged to the mitochondrial-lysosomal fraction, not to the microsomal fraction. Both the enhancing activity and the activity that converting [14C]-RS-1541 to [14C]-rhizoxin (RS-1541-converting activity) were inhibited by treatment with chloroquine, an inhibitor of lysosomal function. Cholesterol esterase derived from Candida cylindracea had RS-1541-enhancing and -converting activities. These data suggest that RS-1541 exerts its cytotoxic action after being converted to rhizoxin within the cells by a lysosomal enzyme such as cholesterol esterase.

Antibiotics, Antineoplastic↗

PubMed1995

Reconstitution of purified P-glycoprotein into liposomes.

To study the molecular function of the multidrug-resistance gene product P-glycoprotein, we purified and reconstituted it into liposomes. Twelve detergents were examined in an attempt to solubilize and reconstitute the transport activity of K562/ADM membrane proteins containing P-glycoprotein. We found that transport activity was effective reconstituted after solubilization with cholate, glycocholate and taurocholate. Other detergents, such as CHAPS, Triton X-100 and deoxycholate, diminished the transport activity. The K562/ADM membrane was solubilized by 1% glycocholate, and P-glycoprotein was purified by MRK-16 immunoaffinity column chromatography to a homogeneous single band on sodium dodecyl sulfate/polyacrylamide gel electrophoresis. The purified P-glycoprotein was reconstituted by detergent dialysis into liposomes composed of phosphatidylcholine, phosphatidylethanolamine and phosphatidylserine. The reconstituted P-glycoprotein specifically bound [3H]azidopine and had an ATPase activity that was slightly stimulated when vincristine was added. Furthermore, though its activity was reduced, the reconstituted P-glycoprotein was shown to be an ATP-dependent transporter of vincristine.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

PubMed1995

Expression of the 85-kd membrane protein in primary human breast cancer: relationship to hormone receptor levels, DNA ploidy, and tumor grade.

Using the monoclonal antibody (MAb) MRK20 the immunohistochemical expression of the 85-kd membrane protein (MP)/cell adhesion molecule (CAM) CD36 was investigated in tumors from 41 patients with untreated breast adenocarcinoma. DNA ploidy as measured by flow cytometry and estrogen (ER) and progesterone receptor (PgR) tumor contents also were determined. Immunohistochemistry on frozen tissue showed expression of 85-kd MP localized to the carcinoma cells in tumors from 16 patients (39%). These samples also displayed variable staining of the capillary endothelium. Twenty-one samples (51%) showed 85-kd MP expression in small blood vessels. Thus, a total of 37 of 41 tumors (90%) were MAb MRK20 positive in carcinoma and/or mesenchymal cells. Expression of 85-kd MP in carcinoma cells was negatively associated with expression of ER (P < .05) and PgR (P < .01) receptors and positively associated with nondiploid tumors (P < 0.05) and tumor grade (P < .05). Axillary lymph node status, tumor size, and (patient) age were not associated with expression of 85-kd MP. In cytological smears obtained by fine-needle aspiration (FNA) cytology from 19 of the samples, expression of 85-kd MP in carcinoma cells showed a good correlation with tumor cell positivity in the corresponding histological section. Expression of 85-kd MP in breast carcinoma cells thus was associated with some recognized adverse prognostic factors. Future studies of its pathophysiological and prognostic roles in breast cancer seem warranted.

Adenocarcinoma↗

PubMed1995

Induction of hepatic P-glycoprotein enhances biliary excretion of vincristine in rats.

To clarify the contribution of P-glycoprotein to the biliary excretion of vincristine in rats, the effects of induction of hepatic P-glycoprotein by a phenothiazine treatment on the biliary excretion of [3H]vincristine were investigated. Immunoblot analysis using C219, a monoclonal antibody to P-glycoprotein, demonstrated that the phenothiazine treatment increased the P-glycoprotein level in isolated bile canalicular membrane vesicles approximately 6.5-fold. Transport of [3H]vincristine to canalicular membrane vesicles from the phenothiazine-treated and control rats revealed ATP-dependency, with an overshoot that results from the consumption of medium ATP. The maximum ATP-dependent uptake was increased in canalicular membrane vesicles from the phenothiazine-treated rats approximately 2-fold compared to the control. The biliary excretion of [3H]vincristine was further studied using an indicator dilution method in a single-pass perfused liver. The ratios of the cumulative amount of [3H]vincristine excreted into the bile ot the amount of [3H]vincristine taken up by the liver at 15, 30 and 90 min were significantly increased in the phenothiazine-treated rats by 60, 45 and 25%, respectively, compared to the control rats. Furthermore, the corrected mean residence time of [3H]vincristine in hepatocytes in the phenothiazine-treated rats was reduced to 21 min from that in the control rats (30 min), supporting the contention that the induction of hepatic P-glycoprotein on the bile canalicular membrane function sas a transporter not only in the isolated membrane but also in the more physiological perfused liver system. One must be cautious in the interpretation of the data, however, since phenothiazine can induce other proteins which might affect the behavior of [3H]vincristine.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

PubMed1995

In vivo imaging and specific targeting of P-glycoprotein expression in multidrug resistant nude mice xenografts with [125I]MRK-16 monoclonal antibody.

Multidrug resistance (MDR) in tumors is associated with P-glycoprotein (Pgp) expression. In vivo quantitation of Pgp may allow MDR to be evaluated noninvasively prior to treatment planning. The purpose of this study was to radiolabel MRK-16, a monoclonal antibody that targets an external epitope of P-glycoprotein, and perform in vivo quantitation of P-glycoprotein in a MDR xenograft nude mouse model. MRK-16 was labeled with 125I by the iodogen method, with subsequent purification by size exclusion chromatography. Groups of 10 Balb c mice were each xenografted with colchicine-resistant or sensitive neuroblastoma cell lines, respectively. Whole body clearance and tumor uptake over time was quantitated by gamma camera imaging, and biodistribution studies were performed with [125I]MRK-16 and an isotype matched control antibody, A33. Quantitative autoradiography and immunohistochemistry analysis of tumors was also evaluated to confirm specific targetting of [125I]MRK-16. Peak tumor uptake was at 2-3 days post-injection, and was significantly greater in resistance compared to sensitive tumors (mean % injected dose/g +/- SD) (18.76 +/- 2.94 vs 10.93 +/- 0.96; p < 0.05). Quantitative autoradiography verified these findings (19.13 +/- 0.622 vs 12.08 +/- 0.38, p < 0.05). Specific binding of [125I]MRK-16 was confirmed by comparison to [131I]A33 in biodistribution studies, and localized to cellular components of tissue stroma by comparison of histologic and autoradiographic sections of sensitive and resistant tumors. Immunoblot analysis demonstrated a 4.5-fold difference in P-glycoprotein expression between sensitive and resistant cell lines without colchicine selective pressure. We conclude that in vivo quantitation of P-glycoprotein in MDR tumors can be performed with [125I]MRK-16.(ABSTRACT TRUNCATED AT 250 WORDS)

ATP Binding Cassette Transporter, Subfamily B, Mem↗