Design, synthesis, and biological evaluation of analogues of the antitumor agent, 2-(4-[(7-chloro-2-quinoxalinyl)oxy]phenoxy)propionic acid (XK469).

2-(4-[(7-Chloro-2-quinoxalinyl)oxy]phenoxy)propionic acid (XK469) is among the most highly and broadly active antitumor agents to have been evaluated in our laboratories and is currently scheduled to enter clinical trials in 2001. The mechanism or mechanisms of action of XK469 remain to be elaborated. Accordingly, an effort was initiated to establish a pharmacophore hypothesis to delineate the requirements of the active site, via a comprehensive program of synthesis of analogues of XK469 and evaluation of the effects of structural modification(s) on solid tumor activity. The strategy formulated chose to dissect the two-dimensional parent structure into three regions-I, ring A of quinoxaline; II, the hydroquinone connector linkage; and III, the lactic acid moiety-to determine the resultant in vitro and in vivo effects of chemical alterations in each region. Neither the A-ring unsubstituted nor the B-ring 3-chloro-regioisomer of XK469 showed antitumor activity. The modulating antitumor effect(s) of substituents of differing electronegativities, located at the several sites comprising the A-ring of region I, were next ascertained. Thus, a halogen substituent, located at the 7-position of a 2-(4-[(2-quinoxalinyl)oxy]phenoxy)propionic acid, generated the most highly and broadly active antitumor agents. A methyl, methoxy, or an azido substituent at this site generated a much less active structure, whereas 5-, 6-, 8-chloro-, 6-, 7-nitro, and 7-amino derivatives all proved to be essentially inactive. When the connector linkage (region II) of 1 was changed from that of a hydroquinone to either a resorcinol or a catechol derivative, all antitumor activity was lost. Of the carboxylic acid derivatives of XK469 (region III), i.e., CONH2, CONHCH3, CON(CH3)2, CONHOH, CONHNH2, CN, or CN4H (tetrazole), only the monomethyl- and N,N-dimethylamides proved to be active.

Preclinical efficacy of thioxanthone SR271425 against transplanted solid tumors of mouse and human origin.

A highly active and broadly active thioxanthone has been identified: N-[[1-[[2-(Diethylamino)ethyl]amino]-7-methoxy-9-oxo-9H-thioxanthen++ +-4-yl] methylformamide (SR271425, BCN326862, WIN71425). In preclinical testing against a variety of subcutaneously growing solid tumors, the following %T/C and Log10 tumor cell kill (LK) values were obtained: Panc-03 T/C = 0, 5/5 cures; Colon-38 (adv. stage) T/C = 0, 3/5 cures, 4.9 LK; Mam-16/C T/C = 0, 3.5 LK; Mam-17/0 T/C = 0, 2.8 LK; Colon-26 T/C = 0, 1/5 cures, 3.2 LK; Colon-51 T/C = 0, 2.7 LK; Panc-02 T/C = 0, 3.1 LK; B16 Melanoma T/C = 13%, 4.0 LK; Squamous Lung-LC12 (adv. stage) T/C = 14%, 4.9 LK; BG-1 human ovarian T/C = 16%, 1.3 LK; WSU-Brl human breast T/C = 25%, 0.8 LK. The agent was modestly active against doxorubicin (Adr)-resistant solid tumors: Mam-17/AdrT/C =23%, 0.8 LK; and Mam-16/C/Adr T/C = 25%, 1.0 LK, but retained substantial activity against a taxol-resistant tumor: Mam-16/C/taxol T/C = 3%, 2.4 LK. SR271425 was highly active against IV implanted leukemias, L1210 6.3 LK and AML1498 5.3 LK. The agent was equally active both by the IV and oral routes of administration, although requiring approximately 30% higher dose by the oral route. Based on its preclinical antitumor profile, it may be appropriate to evaluate SR271425 in clinical trials.

Preclinical antitumor efficacy of analogs of XK469: sodium-(2-[4-(7-chloro-2-quinoxalinyloxy)phenoxy]propionate.

A series of quinoxaline analogs of the herbicide Assure was found to have selective cytotoxicity for solid tumors of mice in a disk-diffusion-soft-agar-colony-formation-assay compared to L1210 leukemia. Four agents without selective cytotoxicity and 14 agents with selective cytotoxicity were evaluated in vivo for activity against a solid tumor. The four agents without selective cytotoxicity in the disk-assay were inactive in vivo (T/C > 42%). Thirteen of the fourteen agents with selectivity in the disk-assay were active in vivo (T/C < 42%). Five of the agents had curative activity. These five agents had a halogen (F, Cl, Br) in the 7-position (whereas Assure had a CI in the 6 position). All agents with curative activity were either a carboxylic acid, or a derivative thereof, whereas Assure is the ethyl ester of the carboxylic acid. All other structural features were identical between Assure and the curative agents. Assure had no selective cytotoxicity for solid tumors in the disk-assay, and was devoid of antitumor activity. The analog XK469 is in clinical development.

Preclinical antitumor activity of XK469 (NSC 656889).

XK469 (NSC 656889) is a water-soluble member of the novel quinoxaline family of antitumor agents. In vitro, XK469 demonstrated selective cytotoxicity for several murine solid tumors including colorectal and mammary adenocarcinoma cell lines, when compared to both leukemia and normal epithelial cells. In vivo, XK469 was active against 7/7 murine tumors tested, including pancreatic ductal carcinomas #02 and #03, colon adenocarcinomas #38 and #51/A, mammary adenocarcinoma #16/C and the Adriamycin resistant mammary adenocarcinomas #16/C/ADR and #17/ADR. XK469 was efficacious both intravenously and orally. Regardless of dosing schedule, conventional mice tolerated higher total doses than SCID or nu/nu mice did. Despite these reduced doses, XK469 was active against xenografts of 4/6 human tumor lines including mammary adenocarcinoma MX-1, the small cell lung cancer DMS 273, the prostate model LNCaP and the CNS tumor SF295. The lower doses in the xenograft studies were below curative levels. The dose-limiting toxicity appeared to be myelosuppression with rapid host recovery (5-8 days), and in vitro assays of XK469 toxicity to murine bone marrow neutrophil progenitors CFU-GM (colony forming unit-granulocyte/macrophage) demonstrated concentration-dependent toxicity from 0.5-30 microg/mL. The difference in drug tolerance between BDF1 and SCID mice was detected in vitro as a 3-fold difference in the IC90 for CFU-GM, despite similar IC50 values. Comparative in vitro hematotoxicology studies revealed that human bone marrow CFU-GM tolerated XK469 as well as their SCID counterparts (IC90 values 5.7 vs. 7.4 microg/mL). Based on comparison with previously tested anti-cancer agents, these data suggest that humans will be able to tolerate XK469 doses that are efficacious against human tumor xenografts.

Induction of the estrogen specific mitogenic response of MCF-7 cells by selected analogues of estradiol-17 beta: a 3D QSAR study.

Analogues of estradiol-17 beta (E2) have been evaluated for estrogen receptor (ER) binding affinity and mitogenic potential in the human breast cancer cell line MCF-7. These 42 compounds represent subtle modifications of the natural estrogen structure through the placement of hydroxyl, amino, nitro, or iodo groups around the ring system in addition to, or as replacement of, the 3- and 17 beta-hydroxyls of E2. The mitogenic activity of the analogues was found to be related to ER binding only to a limited extent. In order to elucidate structural features that are uniquely responsible for receptor binding affinity or mitogen potential of estrogens, the three-dimensional quantitative structure-activity (QSAR) method Comparative Molecular Field Analysis (CoMFA) was employed. Separate CoMFA models for receptor binding and cell growth stimulation were optimized through the use of various alignment rules and region step size. Whereas the CoMFA contour plots did outline the shared structural requirements for the two measured biological properties, specific topological features in this set of estrogens were delineated that distinguish mitogenic potential from ER binding ability. In particular, steric interference zones which affected growth extend in a band from above the A-ring to position 4 and below, whereas the ER binding steric interference zones are limited to isolated polyhedra in the 1, 2 and 4 positions and the alpha face of the B-ring. In addition, electronegative features located around the A-, B-, or C-rings contribute to receptor affinity. However, growth is dependent only on electronegative and electropositive properties near the 3-position. In a final QSAR model for the mitogenic response, the value of ER binding was included along with structural features as a descriptor in CoMFA. The resulting 3D-QSAR has the most predictive potential of the models in this study and can be considered a prototype model for the general evaluation of a steroidal estrogen's growth stimulating ability in MCF-7 cells. For example, the location of D-ring contours illustrate the model's preference for 17 beta-hydroxy steroids over the less mitogenic 17 alpha- and 16 alpha-hydroxy compounds. In addition, the enhanced mitogenic effect of steric bulk in the 11 alpha-position is also evident. The QSAR studies in this report illustrate the fact that while ER binding may be a required factor of the estrogen dependent growth response in MCF-7 cells, particular structural characteristics, in addition to those responsible for tight receptor binding, must be present to induce an optimal mitogenic response. Therefore, this report demonstrates that the CoMFA QSAR method can be utilized to characterize structural features of test compounds that account for different types of estrogenic responses.

Treatment of human prostate tumors PC-3 and TSU-PR1 with standard and investigational agents in SCID mice.

Both the PC-3 and the TSU-PR1 prostate tumor models were found to be satisfactory for chemotherapeutic investigations in ICR-SCID mice. The 30 to 60 mg fragments implanted took in all mice (as judged by 100% takes in the controls of all experiments as well as the passage mice). The tumor volume doubling time was 4.0 days for PC-3 and 2.5 days for TSU-PR1. Nine agents were evaluated IV against early stage subcutaneous PC-3 tumors, with Nano-piposulfan being the only agent highly active (4.9 log kill). Three other agents were moderately active: Taxol (1.5 log kill), Cryptophycin-8 (1.6 log kill), Vinblastine (1.0 log kill). Five agents were inactive: VP-16, Adriamycin, CisDDPt, 5-FUra, and Cyclophosphamide. Ten agents were evaluated IV against early stage subcutaneous TSU-PR1 tumors. Three agents were highly active, producing > 6 log kill and cures: Taxol (5/5 cures), Cryptophycin-8 (5/5 cures), Vinblastine (2/4 cures). Two other agents were moderately active: Nano-piposulfan (1.2 log kill), and Cyclophosphamide (1.1 log kill). Five agents were inactive: VP-16, Adriamycin, CisDDPt, 5-FUra, and BCNU. In part, activity was determined by the ability of the SCID mice to tolerate meaningful dosages of the agents. Agents producing granulocyte toxicity (e.g., Adriamycin) were poorly tolerated and appeared less active than expected. Vinblastine, producing little or no granulocyte toxicity was very well tolerated and appeared to be more active than expected.

Discovery of cryptophycin-1 and BCN-183577: examples of strategies and problems in the detection of antitumor activity in mice.

Historically, many new anticancer agents were first detected in a prescreen; usually consisting of a molecular/biochemical target or a cellular cytotoxicity assay. The agent then progressed to in vivo evaluation against transplanted human or mouse tumors. If the investigator had a large drug supply and ample resources, multiple tests were possible, with variations in tumor models, tumor and drug routes, dose-decrements, dose-schedules, number of groups, etc. However, in most large programs involving several hundred in vivo tests yearly, resource limitations and drug supply limitations have usually dictated a single trial. Under such restrictive conditions, we have implemented a flexible in vivo testing protocol. With this strategy, the tumor model is dictated by in vitro cellular sensitivity; drug route by water solubility (with water soluble agents injected intravenously); dosage decrement by drug supply, dose-schedule by toxicities encountered, etc. In this flexible design, many treatment parameters can be changed during the course of treatment (e.g., dose and schedule). The discovery of two active agents are presented (Cryptophycin-1, and Thioxanthone BCN 183577). Both were discovered by the intravenous route of administration. Both would have been missed if they were tested intraperitoneally, the usual drug route used in discovery protocols. It is also likely that they would have been missed with an easy to execute fixed protocol design, even if injected i.v.

Crystal structure, receptor binding, and gene regulation of 2- and 4-nitroestradiols.

Crystal structures of 2-nitroestradiol and 4-nitroestradiol showed two different molecular conformations for each compound. The crystal structure of 4-nitroestradiol, as well as that of 4-nitroestrone-3-methyl ether, displayed a nitro group in which the oxygens were perpendicular to the aromatic ring and were this nonconjugating. On the other hand, the nitro-oxygens in 2-nitroestradiol were periplanar, with the aromatic ring permitting conjugating. This latter structure bound to estrogen receptor with 1/1000th the affinity of estradiol and was inefficient in gene stimulation. 4-Nitroestradiol possessed a relative binding affinity 40-fold greater than that of the 2-nitro derivative and actively induced responsive genes at a concentration of 10(-8) M. Whereas binding affinity can be explained primarily by polar groups and skeletal structure, gene induction may be linked to electronic induction in ring A that causes a requisite electronegative isopotential around the molecule. This electronegative characteristic also produces conformational changes in the alicyclic backbone of the estrogen, specially ring B, which could interfere with the molecular fit of the nitroestradiols with estrogen receptor.

Preclinical anticancer activity of cryptophycin-8.

Cryptophycin-8 was prepared by the conversion of the epoxide group on cryptophycin-1 to a chlorohydrin. In the studies reported here, cryptophycin-8 was evaluated for preclinical activity against subcutaneous tumors of both mouse and human origin. At the highest non-toxic single course treatment, the following results were obtained (Table A). Cryptophycin-8 was less potent than cryptophycin-1 by approximately 4-fold; however, it was both more water soluble and had greater therapeutic efficacy, as demonstrated by % T/C, tumor cell log kill values, range of dose effectiveness and host cures.

Formulation and antitumor activity evaluation of nanocrystalline suspensions of poorly soluble anticancer drugs.

PURPOSE: Determine if wet milling technology could be used to formulate water insoluble antitumor agents as stabilized nanocrystalline drug suspensions that retain biological effectiveness following intravenous injection. METHODS: The versatility of the approach is demonstrated by evaluation of four poorly water soluble chemotherapeutic agents that exhibit diverse chemistries and mechanisms of action. The compounds selected were: piposulfan (alkylating agent), etoposide (topoisomerase II inhibitor), camptothecin (topoisomerase I inhibitor) and paclitaxel (antimitotic agent). The agents were wet milled as a 2% w/v solids suspension containing 1% w/v surfactant stabilizer using a low energy ball mill. The size, physical stability and efficacy of the nanocrystalline suspensions were evaluated. RESULTS: The data show the feasibility of formulating poorly water soluble anticancer agents as physically stable aqueous nanocrystalline suspensions. The suspensions are physically stable and efficacious following intravenous injection. CONCLUSIONS: Wet milling technology is a feasible approach for formulating poorly water soluble chemotherapeutic agents that may offer a number of advantages over a more classical approach.

Preclinical antitumor activity of CI-994.

CI-994 [aka: acetyldinaline; PD 123654; 4-acetylamino-N-(2'aminophenyl)-benzamide] (Figure 1) is a novel antitumor agent with a unique mechanism of action. It is the acetylated metabolite of dinaline, a compound previously identified as having cytotoxic and cytostatic activity against several murine and human xenograft tumor models. CI-994 had activity against 8/8 solid tumors tested (log cell kills at the highest non-toxic dose): pancreatic ductal adenocarcinoma #02 (4.7); pancreatic adenocarcinoma #03 (3.0; 1/6 cures); colon adenocarcinoma #38 (1.6); colon adenocarcinoma #51/A (1.1); mammary adenocarcinoma #25 (1.7); mammary adenocarcinoma #17/ADR (0.5); Dunning osteogenic sarcoma (4.0); and the human prostate carcinoma LNCaP (1.2). CI-994 had the same spectrum of activity in vivo as dinaline. It also behaved similarly in schedule comparison/toxicity trials. Prolonged administration with lower drug doses was more effective than short-term therapy at higher individual doses. If doses were kept between 40 and 60 mg/kg/injection, prolonged administration (> 50 days) was tolerated with no gross toxicity. Doses > or = 90 mg/kg/injection caused lethality after 4-5 days of administration. The maximum tolerated total dose was also increased with smaller individual doses administered for prolonged intervals. Clinical Phase I trials are ongoing with this agent.

Relationship between estrogen structure and conformational changes in estrogen receptor/DNA complexes.

The effect of estrogen structure on the conformation of the complex formed with estrogen receptor (ER) and the consensus estrogen response element (EREc) has been examined with gel mobility shift assay. Proteins in MCF-7 cell extracts formed three distinct complexes with ERE. Only the slowest moving complex contained ER as indicated by binding with anti-ER antibodies H222 and D547. This ER-ERE complex displayed increased electrophoretic mobility when formed in the presence of estradiol (E2) and bound radiolabeled 16 alpha-iodoestradiol. The antiestrogen ICI 164,384 decreased the mobility of the ER-ERE complex and blocked the effect of E2. The results reported here indicate that the position and location of hydroxyl groups on the estratriene nucleus is an important factor in determining the mobility of ER-EREc (or a variant ERE) in gel shift assays. The ability of E2 analogs to cause conformational changes detectable as altered mobility was not directly related either to their binding affinity for ER or to their ability to activate E2 responsive genes. Although several dihydroxyestrogens (estradiol-16 alpha, 1- and 2-hydroxyestratrien-17 beta-ol) caused an increase in the mobility of the ER-EREc, other ligands (estradiol-17 alpha, 4-hydroxyestratriene-17 beta-ol, 3-hydroxy estratriene, estratrien-17 beta-ol and 5-androsten-3 beta, 17 beta-diol) with a capacity for activating at least some E2 responsive genes in MCF-7 cells had little or no effect. On the basis of these and previously published results, it can be concluded that specific structure features of estrogens are responsible for conformational changes of ER-ERE complexes detectable in gel-shift assays. Furthermore, the identified structural characteristics of the ligand which are required for gel-shift are not the same as those previously reported to be essential for stimulation of transcriptional activity of ER.

Comparative efficacy of DMP 840 against mouse and human solid tumor models.

BACKGROUND: DMP 840 is a compound from a class of bis-naphthalimide antitumor agents that recently completed Phase I clinical trials at three North American centers and is currently undergoing Phase II testing. Preclinically, it was shown to have curative activity against a variety of human tumor xenograft models. PURPOSE: To test DMP 840 both in vitro and in vivo for antiproliferative activity against predominantly mouse tumor models. METHODS: A disk diffusion soft agar colony formation assay was used to determine the in vitro growth inhibitory activity against a selection of mouse and human tumor cell lines, and the comparable selective mouse solid tumors were used for in vivo testing. RESULT: In vitro DMP 840 exhibited equal cytotoxicity for human tumors (including MX-1 directly cultured from nude mice), mouse tumors and normal cells. In vivo DMP 840 was only modestly active or inactive against the following mouse tumors: Mam 16/C, T/C = 30% (T/C = Percent Tumor Growth Inhibition); Mam 16/C/ADR, T/C = 33%; Colon 38, T/C = 9%; Panc 03, T/C = 53%; Colon 51/A, T/C = 28%; Panc 02, T/C = 52%; P388/0, 36% ILS (Percent Increased Life Span) and P388/ADR, 14% ILS. Furthermore, the antitumor activity was only observed at the highest non-toxic dose and was associated with a large body weight loss. In contrast, the agent was highly active against the human breast tumor MX-1 implanted subcutaneously in either athymic nude or SCID mice (Nudes: T/C = 0%; 1/5 cures; SCIDS: T/C = 0%; 5/5 cures). CONCLUSIONS: Although there was no selective cytotoxicity in our clonogenic assay for human versus mouse tumor cell lines, selective activity in vivo for human xenograft tumors was noted. Overall, this compound is rather unique in its differential degree of in vivo activity for human versus mouse tumors. IMPLICATIONS: Phase II trials, which are ongoing, will help determine if the preclinical in vivo selective activity of DMP 840 translates to clinical activity in man.

Skeletal conformations and receptor binding of some 9,11-modified estradiols.

The effect of the modification of the 9-11 positions on the skeletal conformation of estradiol (E2) has been analyzed by X-ray crystallography and MM2 molecular mechanics. The 11 beta-hydroxyl and 11-keto analogs of E2 maintained ring conformations which were similar to the natural hormone (E2). Introduction of a double bond at position 9-11 induced a flattening of the entire steroid molecule. An 11 alpha-hydroxyl group brought about significant changes in the alicyclic rings of E2. 9 beta-Estradiol and 11-keto-9 beta-estradiol formed ring conformations which were significantly bent from E2 (below the plane of the A-ring). Examination of the affinity of these C-ring analogs of E2 for the human estrogen receptor has shown extreme variations. A hydroxyl group placed either alpha or beta at the 11-position yielded ligands with vastly different and reduced affinities for the receptor. The low affinity of 11 alpha-hydroxyestradiol (1/300th of E2) may be due to the drastic structural change induced in the alicyclic portion of the molecule, as well as, to the steric or electrostatic effects of the alpha-hydroxyl group upon the receptor protein. An 11 beta-hydroxyl group diminished the receptor binding to 1/60th that of E2 without alicyclic ring distortions, whereas a 9-11 unsaturation reduced the binding to 1/5th although this steroid displayed a flattening of rings B, C, and D. The 11-keto function, which had little effect on the conformation of the estrogen nucleus, reduced the affinity of this ligand to 1/1000th that of E2. The negative bend at the C-ring of 11-keto-9 beta-estradiol and 9 beta-estradiol prevented these ligands from binding receptor. Some of the observed receptor interactions were related to structural alterations in the estrogen ring system induced by modifications on the 9-11 region.

Antitumor efficacy of interleukin-2 alone and in combination with adriamycin and dacarbazine in murine solid tumor systems.

Recombinant interleukin-2 (IL-2)/chemotherapy combinations have recently entered clinical trial. The rationale for sequencing has primarily been empiric or based on in vitro data. To establish in vivo models for chemoimmunotherapy trials, we investigated IL-2 alone and in combination with dacarbazine (DTIC) and adriamycin. IL-2 (as a single agent given i.v. at 1-3 x 10(5) Cetus units once daily for 5 days, repeated 7-10 days later), was highly active against an immunogenic line of colon adenocarcinoma no. 11/A [tumor growth inhibition (T/C) = 0% with cures]. It was modestly active against colon adenocarcinoma no. 38 (T/C = 39%), mammary adenocarcinoma no. 16/C (T/C = 18%), and B16 melanoma (T/C = 21%). IL-2 was inactive against colon adenocarcinoma no. 7/A (T/C = 83%). Combination trials were done using DTIC and IL-2 against colon no. 7/A and upstaged colon no. 11/A. The combination of adriamycin and IL-2 was tested against mammary adenocarcinoma no. 16/C. In the DTIC/IL-2 combination trials, the combination was superior over either agent used alone. In the IL-2/adriamycin trials, the combination was no better than adriamycin alone at optimum dosages.

Activity of datelliptium acetate (NSC 311152; SR 95156A) against solid tumors of mice.

Datelliptium acetate (NSC 311152) is a water soluble analogue of ellipticine. It is a solid tumor selective compound. In vitro, in a disk diffusion, soft agar colony formation assay (25 micrograms/disk), the compound demonstrated solid tumor selectivity (compared to leukemia L1210) against colon adenocarcinoma 38 and pancreas ductal carcinoma 03. Upon intravenous administration, NSC 311152 was effective in vivo against a variety of murine solid tumors. Responses at maximum tolerated doses were: colon #07/A (T/C = 33%); 0.60 log cell kill), #38 [T/C = 0%; 4.2 log cell kill), colon #51/A (T/C = 2%; 1.2 log cell kill), undifferentiated colon #26/A (T/C = 38%; 0.4 log kill), mammary #16/C (T/C = 10%; 1.7 log cell kill), and pancreatic ductal carcinoma #03 (T/C = 0%; 80% cures through day 38). It was ineffective against pancreas #02 (T/C = 45%), mammary 17/A (T/C = 53%), and 17/A/ADR (T/C = 52%). At efficacious doses acute neurotoxicity (i.e. stupor and lethargy) and weight loss were noted (with rapid recovery from both toxicities). There were no delayed toxicities. The agent was slightly necrotizing and produced pain on SC injections. In lieu of its preclinical efficacy and toxicity profiles, we recommend further clinical investigation of this agent.

Antitumor efficacy of PD115934 (NSC 366140) against solid tumors of mice.

PD115934 (NSC 366140) is a soluble pyrazoloacridine derivative presently undergoing preclinical toxicology evaluation with the anticipation of Phase I human investigation. The agent displayed both human and murine solid tumor selectivity in vitro in a soft agar disk diffusion assay, relative to its activity against murine L1210 leukemia. In vivo it was highly active against solid tumors colon adenocarcinoma 38 and pancreas ductal carcinoma 03, which was consistent with the cellular cytotoxicity seen in the disk diffusion assay. A log cell kill of greater than 4.0 was demonstrated in vivo against both models. PD115934 was administered by both bolus and infusional therapy. After completion of these trials, it was determined that this compound was a schedule category III agent, i.e., a schedule-independent agent with peak plasma level toxicity. The main toxicity encountered with infusional therapy was myelosuppression. With bolus therapy, central nervous system toxicities were dose limiting. On the basis of our preclinical infusion studies, we recommend a 2-h infusion twice weekly in humans in order to obtain a total dose of 360 mg/m2 over 8 weeks.

Activity of batracylin (NSC-320846) against solid tumors of mice.

Batracylin (NSC 320846) is a water insoluble, solid tumor active compound discovered by the Development Therapeutics Program of the National Cancer Institute (NCI). In vivo, the NCI found this compound to be highly active [median treated tumor mass/median control tumor mass (T/C) = 0 to 20%] both orally and intraperitoneally against colon 38. In a disk diffusion, soft agar colony formation assay (500 ug/disk), we found solid tumor selectively (compared to leukemia L1210) against colon adenocarcinoma 38 (0-170 zu:L1210 leukemia; greater than 950 zu:C8), colon adenocarcinoma 9 (0-170 zu:L1210; greater than 950 zu:C9), colon adenocarcinoma 7/A (0-170 zu:L1210; 250-400 zu:C7), and pancreas ductal carcinoma 03 (0-170 zu:L1210; greater than 950 zu:Panc 03 (200 zone units [zu] = 6.5 mm zone of inhibition of cultured tumor colonies from drug disk). In vivo we have tested batracylin against mammary adenocarcinoma 16/C, colon 9, colon 38, colon 51, Panc 03, and hepatoma 129. Upon oral administration, batracylin was effective against colon 9 (T/C = 2.4%) and marginally active against colon 38 (T/C = 39%). Batracylin was orally ineffective against Panc 03 (T/C greater than 100%), colon #51 (T/C = 77%) and hepatoma 129 (T/C greater than 100%). Upon subcutaneous administration, batracylin was effective against colon #9 (T/C = 0%), and Panc 03 (T/C = 15%) but ineffective against mammary 16/C (T/C greater than 100%). At efficacious doses, delayed neurotoxicity, hepatic toxicity and a significant host weight loss was noted (with slow recovery). Both our in vitro data and the NCI in vivo data confirm its scant activity against L1210 (%ILS = 8 to 16%). Although showing activity against selected murine solid tumors, it lacked curative potential with early stage disease [C38, C9, Panc 03] and has shown relative inactivity in vitro against human solid tumor cell lines (H-125, CX-1, HCT-8, HCT-116). Batracylin has entered large animal toxicology trials at the NCI, anticipating phase I clinical evaluation.

5-Fluorouracil containing combinations in murine tumor systems.

5-FUra is a familiar member of a variety of clinically useful regimens. Examples include: Cytoxan-Adriamycin-5-FUra (CAF); Methotrexate-Prednisone-Vincristine-Cytoxan-5-FUra (the Cooper regimen); and CisDDPt-5-FUra. In addition to its clinical utility, over 20 different 5-FUra-combinations have been reported to be therapeutically synergistic in experimental systems. In this report, we will attempt to evaluate the reasons behind the widespread utility of this agent in combination therapy and suggest future directions for research efforts.

Chemotherapy of the squamous cell lung cancer LC-12 with 5-fluorouracil, cisplatin, carboplatin or iproplatin combinations.

The combination of Cis-dichlorodiammineplatinum (II) (CisDDPt) + 5-Fluorouracil (5-FU) was compared with two CisDDPt analogues + 5-FU [Iproplatin (CHIP) + 5-FU and Carboplatin (CBDCA) + 5-FU] for relative efficacy against advanced stage squamous cell lung tumors (LC-12) in Balb/c mice. At equitoxic dosages, the numbers of regressions and cures were similar for the three combinations (5-FU/CISDDPt 2/10 PR's, 2/10 CR's, 2/10 cures; 5-FU/CBDCA 1/10 PR's, 5/10 CR's, 3/10 cures; 5-FU/CHIP 1/10 PR's, 3/10 CR's, 3/10 cures). The tumor growth delay among the mice not cured was slightly superior in the 5-FU/CisDDPt regimen. All the agents were active singly against this tumor model. Based on these results, the substitution of CBDCA or CHIP for CisDDPt in a FU regimen did not offer a cytotoxic advantage. Because of different dose limiting toxicities for the platinum compounds the possibility exists that these analogues could be used in drug combinations in substitution for CisDDPt.