Potentiation of antitumor activity of irinotecan by chemically modified oligonucleotides.

Co-administration of synthetic chemically modified oligonucleotides with irinotecan, a selective topoisomerase I inhibitor, provided a significant enhancement in the antitumor activity of irinotecan. The enhancement of antitumor activity of irinotecan with co-administration of chemically modified oligonucleotides was observed in several tumor models--pancreatic cancer (Panc-1), colon cancer (HCT-116) and melanoma (A375). Inhibition of tumor growth in all three models required the co-administration of irinotecan and chemically modified oligonucleotides, but was independent of the nucleotide sequence of the oligonucleotides. The potentiation of antitumor activity was dependent on the dose of irinotecan and chemically modified oligonucleotides administered. The enhancement of antitumor activity of irinotecan was also observed by co-administration of a phosphorothioate oligonucleotide, however, to a lesser extent than did chemically modified oligonucleotides, suggesting that metabolic stability of the oligonucleotide contributes to the enhancement of antitumor activity seen with irinotecan. The co-administration of dextran sulfate sodium with irinotecan showed insignificant potentiation of antitumor activity of irinotecan, suggesting that the enhancement of antitumor activity of irinotecan observed was not a result of polyanionic characteristic of oligonucleotides. Co-administration of irinotecan and chemically modified oligonucleotides did not result in increased toxicity in the tumor models studied. Potentiation of antitumor activity of irinotecan observed with co-administration of oligonucleotides suggests that the oligonucleotides affect the pharmacokinetics and/or metabolism of irinotecan. The use of chemically modified oligonucleotides together with irinotecan may increase the therapeutic index of irinotecan in cancer patients and continued development of such agents should be considered.

Antitumor activity and toxicity of novel nitroheterocyclic phosphoramidates.

A series of novel nitroheterocyclic phosphoramidates has been evaluated for antitumor activity in murine and xenograft tumor models and for toxicity in mice. Significant increases in lifespan and long-term survivors were noted in L1210 leukemia and B16 melanoma models, and both complete and partial tumor regressions were observed in the MX-1 breast cancer xenograft model. All compounds exhibited some degree of toxicity to granulocyte/macrophage progenitors in the bone marrow of mice. Two drugs were selected for further toxicologic, histopathologic, and pharmacokinetic evaluations. Toxicity of potential clinical significance was observed only in the bone marrow at the highest drug dose; otherwise no significant abnormalities in blood chemistries or organ histopathology were noted. The bone marrow lesions consisted of reduced numbers of progenitor cells in the myeloid and erythroid series; platelets were not affected. The compounds were eliminated rapidly by first-order kinetics, with half-lives in the 4-12 min range. The best of these compounds exhibits excellent antitumor activity and minimal toxicity at therapeutically effective doses in mice.

Preclinical antitumor activity of 6-hydroxymethylacylfulvene, a semisynthetic derivative of the mushroom toxin illudin S.

6-Hydroxymethylacylfulvene (HMAF; MGI 114) is a novel semisynthetic antitumor agent derived from the sesquiterpene mushroom toxin illudin S. In vitro cytotoxicity determinations produced IC50 concentrations (concentrations required for 50% inhibition of growth) ranging from 160 nM in sensitive MCF-7 human mammary carcinoma cells to 17 microM in relatively insensitive murine B16 melanoma cells. In vivo antitumor activity was consistent with in vitro sensitivity. HMAF was very effective in human tumor xenograft models, including MX-1 breast carcinoma, MV522 lung adenocarcinoma, and HT-29 colon carcinoma, but not murine B16 melanoma or P388 leukemia. Excellent responses were observed in animals bearing MX-1 tumors administered i.v. or i.p. doses of 3-7.5 mg/kg daily for 5 days, with complete regression recorded in 29 of 30 animals administered i.v. HMAF. Extensive tumor shrinkage was also observed with MV522, and significant tumor growth inhibition was obtained with HT-29 when animals received 5 daily i.p. doses ranging from 3.75 to 7.5 mg/kg. Complete regressions were also observed in individual animals with MV522 and HT-29. The excellent activity of HMAF in several human solid tumor xenografts, including the more refractory MV522 and HT-29 models, warrants the further investigation of this novel agent in clinical trials.

Evaluation of a novel bis-naphthalimide anticancer agent, DMP 840, against human xenografts derived from adult, juvenile, and pediatric cancers.

The new bis-naphthalimide antitumor agent (R,R)2,2'-[1,2-ethanediylbis[imino(1-methyl-2.1-ethanediyl)]-bis(5 -nitro 1H-benz[de]-isoquinoline-1,3-2H) dione] dimethanesulfonate (DMP 840) was evaluated against parental and multidrug-resistant human KB cell lines in vitro and against these lines growing as xenografts in immune-deprived mice. In vitro, KB8-5 cells were 50-fold resistant to vincristine but only 16-fold resistant to DMP 840 as measured by clonogenic survival. For in vivo evaluation, DMP 840 was given by i.v. injection daily for 9 days or for 5 days/week for 2 consecutive weeks [(dx5)2]. In contrast to the cross-resistance of KB cell lines in vitro, both KB3-1 and KB8-5 tumors were highly and equally sensitive to DMP 840; only KB3-1 xenografts demonstrated sensitivity to vincristine, which was consistent with the in vitro results. DMP 840 was also evaluated against a panel of human tumors comprising colon adenocarcinoma and rhabdomyosarcoma xenografts. Against eight lines of colon adenocarcinoma, DMP 840 caused a high frequency of partial and complete regressions in two lines and significant inhibition of growth in two lines. DMP 840 caused complete regressions in five of six lines of advanced rhabdomyosarcomas, demonstrating a broad range of effective dose levels. The pattern of activity against this tumor panel was similar but not identical to that of two inhibitors of topoisomerase I. There was no cross-resistance to DMP 840 in xenografts selected for resistance to vincristine or in a rhabdomyosarcoma selected for resistance to the topoisomerase I inhibitor topotecan. In contrast, a colon tumor selected for topotecan resistance was completely resistant to DMP 840. Slight cross-resistance to DMP 840 was demonstrated in a rhabdomyosarcoma xenograft that was selected for primary resistance to melphalan and was cross-resistant to topoisomerase I inhibitors. The pattern of activity and cross-resistance in these tumors was compared with that shown by two agents that inhibit topoisomerase I: topotecan and CPT-11.

Efficacy of DMP 840: a novel bis-naphthalimide cytotoxic agent with human solid tumor xenograft selectivity.

DMP 840, a novel bis-naphthalimide, was evaluated for antitumor efficacy in several tumor models in mice. As measured by a tumor growth inhibition assay, i.v. administration of DMP 840 to athymic nude mice at doses at or below the maximum tolerated dose resulted in curative activity against four human solid tumor xenografts, MX-1 mammary carcinoma, CX-1 and DLD-2 colon adenocarcinomas, and LX-1 lung carcinoma, producing full or incomplete regressions and/or percent tumor growth inhibition of > or = 96%. The efficacy of DMP 840 in the models was dose dependent. The activity of DMP 840 against the human tumors surpassed that demonstrated by several clinically used and investigational anticancer agents. In long-term growth delay studies, DMP 840 induced full regressions in 20 of 20 mice bearing MX-1 tumors, and tumors in one-half of these mice remained regressed for over 5 months. In addition, DMP 840 was curative against exponentially growing DLD-2 tumors staged at 500 mg and MX-1 tumors staged at 1000 mg. The bis-naphthalimide was equally efficacious when administered i.v. or i.p. but was slightly less active after oral dosing. Against both the MX-1 mammary carcinoma and the DLD-2 colon adenocarcinoma, some measure of schedule dependence was observed; the optimum schedule was daily for 9 days. Against L1210 and P388 murine leukemias, DMP 840 demonstrated little or no activity and was inactive against B16 murine melanoma. Overall, these results suggest that DMP 840 may be a human solid tumor selective cytotoxic agent.

XB596, a promising bis-naphthalimide anti-cancer agent.

We have synthesized a promising class of bis-naphthalimide anti-tumor agents. A representative compound in this series, XB596, exhibits potent in vitro growth inhibitory activity against several human and murine leukemic and solid tumor lines in culture, with IC50 values ranging from 7.2 to 147.5 nM. XB596 was almost as equally growth inhibitory against three doxorubicin-resistant cell lines compared with their parental lines. Using a human tumor colony-forming assay, XB596 demonstrated cytocidal activity against fresh human tumors taken directly from patients, with 23 of 25 evaluable tumors responding to a continuous exposure of 1 microgram/ml of XB596. When L1210 cells were incubated with XB596 for 1 h, the incorporation of uridine and thymidine into RNA and DNA, respectively, was inhibited with IC50 values of 0.14 microM. DNA single-strand breaks, but not double-strand breaks, were detected in XB596-treated L1210 cells. XB596 bound to DNA with guanine-cytosine sequence selectivity as shown by an indirect ethidium bromide displacement assay. XB596 was shown to interact with DNA by a spectrophotometric titration assay, with an estimated binding constant of 4.7 +/- 2.2 +/- 10(6) M-1. XB596 unwound supercoiled DNA as measured by agarose gel electrophoresis. These data are consistent with XB596 being a DNA intercalator. In vivo, XB596 demonstrated good anti-tumor activity against two human solid tumors (DLD-2 colon adenocarcinoma and MX-1 mammary carcinoma) xenografted in nude mice, but has not demonstrated anti-leukemic activity. In summary, XB596 is a pre-clinical anti-cancer agent which interacts with DNA and demonstrates good in vivo anti-tumor activity against human solid tumor xenografts.

Chemotherapy of mammary carcinomas arising in ras transgenic mice.

Transgenic female mice carrying the V-Ha-ras transgene linked to the MMTV promoter, which developed mammary carcinomas, were treated with selected cancer chemotherapy drugs. Agents were administered i.p. on a daily x 9 schedule when mice developed tumors that were 50-100 mg in size. Drugs which are clinically effective against breast cancer were quite efficacious in the transgenic model at their maximum tolerated dose. Doxorubicin produced excellent responses in tumor-bearing transgenic mice, with several mammary carcinomas undergoing tumor shrinkage. Two anthrapyrazoles, DuP 937 and DuP 941, novel anticancer drugs with phase 2 activity against breast cancer, were as effective as doxorubicin in the oncomice. Mitoxantrone, a synthetic agent with some properties similar to the anthracyclines, also had antitumor activity, but not as pronounced as obtained with doxorubicin or the anthrapyrazoles. Cisplatin, a drug with limited use in human breast cancer, only caused modest antitumor responses. A computerized data analysis method based on the area under the tumor growth curve was developed to better quantitate the data and provide statistical information. This quantitative analysis confirmed the high statistical significance of the activity of doxorubicin or the anthrapyrazoles in the ras transgenic model, and defined an excellent dose response relationship for each drug tested. Our results suggest that the ras transgenic model may be useful for identifying drugs that have efficacy for breast cancer in women.

Effects of modulation of basic fibroblast growth factor on tumor growth in vivo.

BACKGROUND: Recombinant human basic fibroblast growth factor (rHu-bFGF) is known to stimulate proliferation in some tumor cells and to modulate tumor vascularization. PURPOSE: The purpose of this study was to examine the possible role of this agent in the development of tumors. The study was designed to determine the effects of modulating bFGF activity in vivo in tumor models from cell lines with different responses to bFGF and with different content and receptor levels of bFGF. METHODS: Two tumor cell lines (human DLD-2 colon carcinoma and rat C6 glioma) were characterized for bFGF content and bFGF receptor levels by Western blot analysis in cultured cells and by studies of [125I]rHu-bFGF binding to sections from xenografts grown in nude mice. Tumor cell proliferation was monitored after treatment with rHu-bFGF or the DG2 or DE6 IgG monoclonal antibody to rHu-bFGF in culture and in vivo. RESULTS: C6 cells exhibited 7800 high-affinity receptors for rHu-bFGF per cell (dissociation constant [Kd] = 46 pM), while DLD-2 cells lacked high-affinity receptors. rHu-bFGF stimulated [3H]thymidine uptake by C6 cells, but the addition of DG2 IgG prevented this stimulation; rHu-bFGF had no effect on [3H]thymidine incorporation by DLD-2 cells. C6 cells had higher levels of immunoreactive bFGF than did DLD-2 cells. The xenografts from both cell lines exhibited high-affinity [125I]rHu-bFGF binding that was concentrated on vascular-like structures. rHu-bFGF at a dosage of 0.25 mg/kg given intraperitoneally daily for 18 days caused a twofold increase in DLD-2 tumor weight but had little effect on the growth of C6 xenografts. In contrast, daily intravenous injections of DG2 IgG given to mice had no effect on DLD-2 tumor growth but reduced growth of C6 tumors by approximately 30%--a statistically significant difference. CONCLUSIONS: The addition of exogenous rHu-bFGF or of a neutralizing antibody resulted in significant alterations in tumor growth in vivo, which were specific for tumor type and bFGF characteristics. While some of these effects may be mediated by the bFGF-responsive endothelial cells of the tumor vasculature (DLD-2 colon carcinoma), others may result from inhibition of bFGF-dependent tumor cell proliferation (C6 glioma). IMPLICATIONS: Studies that measure tumor blood flow are necessary to confirm that these effects are mediated by changes in tumor vasculature.

Basic fibroblast growth factor: a potential autocrine regulator of human glioma cell growth.

Basic fibroblast growth factor (bFGF) is a potent mitogen and angiogenic factor. bFGF is expressed by a variety of solid human tumors and has been implicated as an autocrine regulator of tumor growth. Different solid tumor lines including glioma, colon carcinoma and melanoma were examined for intracellular immunoreactive bFGF, high- and low-affinity bFGF receptors and mitogenic response to bFGF when grown in chemically defined medium. All tumor lines contained significant levels of bFGF. In addition, all tumor lines contained subsets of five forms of immunoreactive bFGF, as well as 0.68-20 x 10(6) low affinity bFGF binding sites (Kd = 15-300 nM). Most, but not all lines exhibited high affinity bFGF receptors (Kd = 25-40 pM). Glioma cell lines were distinguished by expressing the highest levels of bFGF protein as well as the most high-affinity receptors for bFGF. Furthermore, glioma cell lines were the only tumor type mitogenically responsive to bFGF. These results indicate that glioma cells express high levels of this potent mitogen and angiogenic factor relative to human colon carcinoma and melanoma cells. The expression of bFGF and bFGF receptors by glioma cells may be related to abnormal growth and neoplastic progression in these tumors.

Structure-activity relationship of quinoline carboxylic acids. A new class of inhibitors of dihydroorotate dehydrogenase.

The novel anticancer drug candidate brequinar sodium [DuP 785, NSC 368390, 6-fluoro-2-(2'-fluoro-1,1'-biphenyl-4-yl)-3-methyl-4-quinoline carboxylic acid sodium salt] inhibits dihydroorotate dehydrogenase, the fourth enzyme in the de novo pyrimidine biosynthetic pathway leading to the formation of UMP. Sixty-nine quinoline 4-carboxylic acid analogs were analyzed as inhibitors of L1210 dihydroorotate dehydrogenase. This structure-activity relationship study identified three critical regions of brequinar sodium and its analogs, where specific substitutions are required for the inhibition of the activity of dihydroorotate dehydrogenase. The three principal regions are: (i) the C(2) position where bulky hydrophobic substituents are necessary, (ii) the C(4) position which has a strict requirement for the carboxylic acid and its corresponding salts, and (iii) the benzo portion of the quinoline ring with appropriate substitutions. These results will be useful in the elucidation of the precise nature of the interaction between brequinar sodium and dihydroorotate dehydrogenase.

Synthesis and biological evaluation of 2-styrylquinazolin-4(3H)-ones, a new class of antimitotic anticancer agents which inhibit tubulin polymerization.

A novel series of 2-styrylquinazolin-4(3H-ones which inhibited tubulin polymerization and the growth of L1210 murine leukemia cells was discovered. Extensive structure-activity relationship studies suggest that the entire quinazolinone structure was required, but activity was further enhanced by halide or small hydrophobic substituents at position 6. These analogues did not substantially interfere with the binding of radiolabeled colchicine, vinblastine, or GTP to tubulin and weakly stimulated GTP hydrolysis uncoupled from polymerization. Several analogues have shown in vivo tumor growth inhibitory activity in the L1210 leukemia model, with the lead compound 5o exhibiting good antitumor activity against murine solid tumors as well as human tumor xenografts.

Basic fibroblast growth factor-like activity and receptors are expressed in a human glioma cell line.

Basic fibroblast growth factor (bFGF), a potent mitogen and angiogenic peptide, has been examined as an autocrine regulator of glioma cell growth. The addition of purified bovine pituitary bFGF to an established human glioma cell line, SNB-19, doubled the density of these cells in chemically defined medium. Half-maximal stimulation occurred at 8.2 ng/ml (480 pM). Also, human recombinant bFGF (hr-bFGF) significantly enhanced the growth of SNB-19 cells in soft agar. SNB-19 cells expressed both high and low affinity binding sites for hr-bFGF. These cells expressed approximately 13,000 high affinity sites/cell (Kd = 16.6 +/- 1.7 pM) and 9.5 x 10(6) low affinity sites/cell (Kd = 61.2 +/- 4.1 nM). The results of cross-linking experiments with iodinated hr-bFGF demonstrated the presence of two bands with molecular masses of 145 and 130 kDa. High affinity receptors were also demonstrated in SNB-19 tumors grown in nude mice. SNB-19 cell extracts contained mitogenic activity that eluted from heparin-agarose with high salt (1.2-2 M NaCl) and exhibited many properties normally associated with authentic bFGF. This material cross-reacted with a monoclonal antibody to hr-bFGF, comigrated with hr-bFGF by Western blot analysis, competed with 125I-hr-bFGF in a radioreceptor assay, and stimulated SNB-19 cell growth. These results indicate that a human glioma cell line both expresses and utilizes a bFGF-like growth factor. Such a factor may be an important autocrine regulator of glioma cell growth and may also facilitate its neoplastic progression.

Distribution of the novel anticancer drug candidate Brequinar sodium (DuP 785, NSC 368390) into normal and tumor tissues of nude mice bearing human colon carcinoma xenografts.

The distribution of the novel anticancer drug candidate Brequinar Sodium (DuP 785, NSC 368390) was studied in control mice and mice implanted subcutaneously with human colon carcinoma xenografts. Mice were given radiolabeled 14C-Brequinar Sodium intravenously. Brequinar concentrations in blood and various tissues were determined at 1, 6, and 24 h after drug administration. Within 1 h Brequinar distributed to the tumor and all other tissues studied. The tumor-to-blood drug concentration ratios ranged from 0.19 to 0.41. Radioactivity in the liver and small intestine at 1 h accounted for 17% and 13%, respectively, of the dose given. Elimination rates of Brequinar from all tissues were approximately equal to that from blood. Comparison of blood concentrations determined by both radioactivity and HPLC methods suggests that the intact drug is probably the only form in the blood.

Plasminogen activator and inhibitor activity in human glioma cells and modulation by sodium butyrate.

The activity of the serine protease plasminogen activator (PA), which correlates with tumorigenicity and metastatic capacity, was examined using the 125I-labeled fibrin plate assay in cell extracts from four human glioma lines as a function of growth in vitro. Cell-associated inhibitory activity to plasmin and urokinase-type PA was also measured concurrently. The relative PA activities differed markedly among the lines, whereas inhibitory activities did not. Two lines, SNB-19 and SNB-75, exhibited maximal PA activities (1-6 m Plough units/micrograms protein) as cultures approached confluence, whereas two other lines, SNB-56 and SNB-78, expressed low PA activity at all times (less than 0.2 m Plough units/micrograms protein). The PA of SNB-19 cell extracts was predominantly urokinase-type PA. In addition to having the highest PA levels, SNB-19 and SNB-75 were the most clonogenic in soft agar and tumorigenic in nude mice. In contrast, SNB-56 and SNB-78 were poorly clonogenic in soft agar and were not tumorigenic in nude mice. Measured directly, inhibitory activities to plasmin, urokinase-type PA, and tissue-type PA were detected in SNB-19 (high PA) and SNB-56 (low PA) cell extracts. However, there were no qualitative or quantitative differences in inhibitor effects between SNB-19 and SNB-56 suggesting that the differences in PA activity between these lines resulted from changes in PA activity and were not due to differential plasminogen activator inhibitor effects. The ability of the differentiating agent sodium butyrate (NaB) to modulate total PA activity was also examined. Peak SNB-19 cell PA activity was decreased in a concentration (Ki, 0.75 mM) and time-dependent manner by the addition of nontoxic amounts of NaB. The dose-dependent decrease in PA activity induced by NaB was most likely due to an effect on PA itself, since the action of inhibitor on urokinase was unchanged in response to NaB. These results suggest that net cellular PA activity in glioma cells is a balance between relative PA activity and inhibitor(s) effects and that this balance can be modulated by sodium butyrate.

Artificial capillary culture studies of human tumor cell growth, differentiation, and marker production.

The morphological characteristics and the production of biochemical markers were determined for 8 human tumor cell lines grown in artificial capillary culture. Comparisons were made with nude mouse xenografts and conventional monolayer or suspension cultures. Capillary histologies reproduced the features of neoplastic differentiation and glandular formation exhibited by the original human tumors and xenografts. The concentrations of specific biochemical markers, such as carcinoembryonic antigen, aspartate aminotransferase, and immunoglobulin, were higher in the pericellular culture medium in capillary culture. The capillary environment influenced the expression of biochemical heterogeneity by the DLD-1 colon carcinoma cell line and its derivative clones. Spontaneous differentiation of K562 leukemia cells was increased in the capillary system. These results indicate that the artificial capillary is a useful and relevant system for the study of cultured human tumor cells.

Mechanism of action of the novel anticancer agent 6-fluoro-2-(2'-fluoro-1,1'-biphenyl-4-yl)-3-methyl-4-quinolinecarbo xylic acid sodium salt (NSC 368390): inhibition of de novo pyrimidine nucleotide biosynthesis.

Exposure of cultured clone A human colon tumor cells to 25 to 75 microM of NSC 368390 [6-fluoro-2-(2'-fluoro-1,1'-biphenyl-4-yl)-3-methyl-4-quinolinecarbox yli c acid sodium salt, DuP 785] for 48 to 72 h resulted in a 99.9% cell kill as determined by clonogenic assay. Cells exposed to NSC 368390 became depleted in intracellular pools of uridine 5'-triphosphate and cytidine 5'-triphosphate. Both uridine 5'-triphosphate and cytidine 5'-triphosphate were decreased to 50% of levels in control cells at 3 h and were undetectable at 15 h after addition of 25 microM of NSC 368390 to the cultures. Similar effects were observed in L1210 leukemia cells. Addition of 0.1 mM of uridine or cytidine restored intracellular pools of uridine 5'-triphosphate and cytidine 5'-triphosphate to control levels and rescued clone A cells from NSC 368390 cytotoxicity. Addition of uridine circumvented NSC 368390 cytotoxicity in L1210 cells, but addition of cytidine did not. This result is consistent with the fact that L1210 cells lack cytidine deaminase and thus cannot form uridine or its anabolites from cytidine. These results indicated that NSC 368390 inhibits a step in the de novo biosynthetic pathway leading to uridine 5'-monophosphate. Therefore, the effects of NSC 368390 on the six enzymes that comprise the de novo pathway leading to the formation of uridine 5'-monophosphate were examined. The results showed that NSC 368390 was a potent inhibitor of dihydroorotate dehydrogenase, the fourth enzyme in the pathway; thus, this study demonstrates that NSC 368390 exerts its tumoricidal effect by inhibiting a step in de novo pyrimidine biosynthesis resulting in the depletion of critical precursors for RNA and DNA synthesis.

Tumor heterogeneity and drug resistance.

Drug resistance has long been identified as a major reason for therapy failure in cancer patients. Concurrently, work from many laboratories in the past 10 years has established tumor heterogeneity as a phenomenon of critical importance in the natural history of individual neoplasms. The two most sinister aspects of intraneoplastic diversity in human solid tumors are the genesis of clones with metastatic potential, and the existence of drug-resistant variants in primary cancers and their metastases. Thus, recent investigations on drug resistance and on tumor heterogeneity have converged to focus attention on the clonal organization of primary tumors and their metastases as the underlying basis for anticancer drug resistance. This review examines the degree of heterogeneity observed within tumors and the relationship of this diversity to resistance that might be anticipated for any given agent. A question critical to our discussion is "How many subpopulations are there?" The impact of multiple tumor clones on therapy is next discussed in relationship to normal tissue tolerance, the barrier clinicians face regardless of the specific agent used in treatment. Finally, laboratory and clinical approaches are presented for addressing a drug resistance problem that is seemingly overwhelming because of its complex biological roots.

Enhancement of the responses of human colon adenocarcinoma cells to X-irradiation and cis-platinum by N-methylformamide (NMF).

The ability of the maturational agent N-methylformamide (NMF) to modify the response of exponentially growing clone A human colon adenocarcinoma cells to x-irradiation, cis-platinum (cis-DDP), or x-irradiation combined with cis-platinum was studied using an in vitro clonogenic assay. When clone A tumor cells were adaptively grown in medium containing 1% NMF (V/V) for 3 passages prior to experiments, a significantly increased sensitivity to x-irradiation as compared to non-NMF treated cells was found. This increased sensitivity was most marked in the low dose region of the survival curve (as indicated by a large increase in the alpha constant in the linear-quadratic equation), and is similar to the increased radiosensitivity observed after treatment of these tumor cells with N,N-dimethylformamide (DMF). Growth in NMF medium also sensitized these cells to the cytotoxic effects of a 1 hr treatment with cis-DDP at 37 degrees C. A dose enhancement factor of about 1.8 was found at the 10% level of survival for the NMF adapted and cis-DDP treated cells as compared to control cells. Clone A cells were treated either immediately prior to or immediately after x-irradiation with a single low dose of cis-DDP (1.5 microgram/ml, 1 hour at 37 degrees C) after adaptation to growth in NMF containing medium, and the modification of the X ray survival curve was compared to cells not exposed to NMF and to NMF-treated cells also treated with cis-DDP. For the non-NMF treated cells, the low dose cis-DDP treatment produced no change in the survival parameters of the X ray survival curve. However, the NMF adapted cells exhibited an additional decrement in cell survival, indicating that the effect of NMF on radiation on cis-DDP cell killing was additive in nature when all 3 agents were combined in this protocol. Also, there was no difference between the sequences of cis-DDP (1 hr, 37 degrees C) + X rays versus X rays + cis-DDP (1 hr, 37 degrees C). These data indicate that combinations of differentiation inducing agents, together with chemotherapeutic agents and X rays, may be a promising avenue of investigation in developing strategies for cancer treatment.

Activity of a novel 4-quinolinecarboxylic acid, NSC 368390 [6-fluoro-2-(2'-fluoro-1,1'-biphenyl-4-yl)-3-methyl-4-quinolinecarb oxylic acid sodium salt], against experimental tumors.

A novel, substituted 4-quinolinecarboxylic acid (NSC 339768) demonstrated antitumor activity against L1210 leukemia and B16 melanoma in the National Cancer Institute's Developmental Therapeutics Program. An extensive analogue synthesis program was initiated; over 200 derivatives were synthesized and tested for anticancer activity. One of these compounds, 6-fluoro-2-(2'-fluoro-1,1'-biphenyl-4-yl)-3-methyl-4-quinolinecarboxylic acid sodium salt, NSC 368390 (DuP-785), was selected for further investigation because of its efficacy against a spectrum of human solid tumors and its water solubility. In initial studies with L1210 leukemia, the compound caused an increase in life span of greater than 80%. The activity was schedule dependent, and the compound was equally efficacious when administered i.p., i.v., s.c., or p.o. In tests against human tumors xenografted under the renal capsule of nude mice, NSC 368390 when injected i.p. in doses of 20-40 mg/kg daily for 9 days inhibited the growth of the MX-1 breast, LX-1 lung, BL/STX-1 stomach, and CX-1 colon carcinomas by greater than 90%. NSC 368390 also inhibited the growth of three distinct human colon carcinomas, the HCT-15, clone A, and DLD-2 tumors, growing s.c. in nude mice. An i.p. dose of 25 mg/kg given daily for 9 days inhibited the growth of the DLD-2 colon cancer by 98%. 1-beta-D-Arabinofuranosylcytosine and Adriamycin were ineffective, and fluorouracil was only moderately effective against these colon tumors. Because of its good activity against human colon tumors and other human carcinomas and its water solubility, NSC 368390 (DuP-785) is being developed as a Phase 1 anticancer agent.

Ultrastructural evidence of dimethylformamide-induced differentiation of cultured human colon carcinoma cells. Increased expression of desmosomes.

N,N-dimethylformamide (DMF) induces differentiation of human colon carcinoma (DLD-1) cells in culture and reduces their tumorigenicity in nude mice. The current investigation analyzed DLD-1 (clone D) cells for ultrastructural evidence of differentiation. Examination of treated and untreated confluent monolayers by transmission electron microscopy revealed an occasional intracytoplasmic lumen indicative of adenocarcinoma. DMF-treated cells showed no signs of a toxic reaction. Cytoplasmic organelles were essentially unchanged except for an increase in tonofilaments and associated desmosomes. The number of desmosomes per unit length of contiguous cell border increased almost sixfold in treated monolayers. No other type of cell junction was seen. The increased frequency of desmosomes in DMF-treated cultures is significant because of the direct correlation known to exist between the number of desmosomes and degree of differentiation of some human carcinomas. Desmosomes serve as foci of cell adhesion and are reduced in number in some invasive tumors. Whether the supernumerary desmosomes in DMF-treated cells contribute to the reduction in malignant behavior of these cells in vivo remains to be determined.