Synthesis, metabolism and in vitro cytotoxicity studies on novel lavendamycin antitumor agents

https://doi.org/10.1016/j.bmc.2010.01.037Get rights and content

Abstract

A series of lavendamycin analogues with two, three or four substituents at the C-6, C-7 N, C-2′, C-3′ and C-11′ positions were synthesized via short and efficient methods and evaluated as potential NAD(P)H:quinone oxidoreductase (NQO1)-directed antitumor agents. The compounds were prepared through Pictet–Spengler condensation of the desired 2-formylquinoline-5,8-diones with the required tryptophans followed by further needed transformations. Metabolism and toxicity studies demonstrated that the best substrates for NQO1 were also the most selectively toxic to NQO1-rich tumor cells compared to NQO1-deficient tumor cells.

Introduction

Lavendamycin (1), a naturally occurring 7-aminoquinoline-5,8-dione antitumor antibiotic, was first isolated from the fermentation broth of Streptomyces lavendulae by Balitz et al.1 and its structure was determined by Doyle et al.2 The latter study determined that lavendamycin is a pentacyclic structure with two moieties including quinoline-5,8-dione and indolopyridine (β-carboline) (Chart).2 Lavendamycin is similar to another potent antibiotic antitumor agent streptonigrin (2).1, 2, 3, 4, 5 The clinical use of both of these antibiotics has been precluded because of their toxicity toward human cells.2, 6, 7, 8 Initial structure–activity relationship (SAR) studies have demonstrated that the essential moiety for the cytotoxic activity of lavendamycin and streptonigrin is the 7-aminoquinoline-5,8-dione moiety.7

As it was thought that some analogues of the naturally occurring antibiotic 1 might show activity toward human tumors, efficient methods were needed to prepare these derivatives. There were two previous reports on the synthesis of lavendamycin methyl ester (3) (above and Table 1) by Kende9 and Boger10 but both were impractical, since they produced compound 3 in an overall yield of less than 2% and in 9- and 20-steps, respectively. Our group was able to develop two synthetic methods11, 12 that produced 3 in an overall yield of about 40% in only 5-steps. These efficient methods, particularly those reported in 1996,12 made it possible to prepare a large number of substituted lavendamycins needed for various in vitro and in vivo screening tests.

We previously reported that a significant number of these lavendamycin analogues have potent biological activity including anti-HIV reverse transcriptase13 and antitumor activity.14, 15, 16 More importantly, several screening tests on a number of lavendamycin analogues have shown that a significant number of these compounds possess low animal toxicity.13, 17 The NCI in vivo studies have shown that the maximum tolerated dose of three derivatives 3, 4 and 5 (Table 1) in mice is 400 mg/kg which is 31 and 1000 times higher than that for lavendamycin and streptonigrin, respectively.13, 17 The lavendamycin analogue 6 reduced tumor volume (up to 80%) in mice bearing tumor xenografts at a daily dose of 300 mg/kg for 10 days without exhibiting drug-related weight loss or lethality.17 A recent study also demonstrated that the normal rat kidney epithelial cell line (NRK-52E) exhibited much less sensitivity to several lavendamycin analogues compared to the tumor cells with the same origin.17 When lavendamycin analogues 7 and 6 at a daily dose of 300 mg/kg and 8 at a daily dose of 100 mg/kg were administered to nude mice for eight and seven days, respectively, no drug-related deaths or toxicity were observed.17 Analogues 8, 7 and 6 (Table 1) inhibited tumor growth in nude mice by 69%, 88% and 78%, respectively, when administered at 100, 150 and 300 mg/kg for 7, 8 and 8 days, respectively.17

NAD(P)H:quinone oxidoreductase 1 (NQO1, DT-diaphorase) is a two-electron reductase, characterized by its capacity for utilizing either NADH or NADPH as reducing cofactors and by its inhibition by dicoumarol.18 It is primarily located in the cytosol (>80%), but a recent study has reported a substantial pool of NQO1 in the nuclei of cancer cells.19 NQO1 is generally categorized as a detoxification enzyme, and it can protect the cell from a broad range of chemically reactive metabolites including electrophiles and reactive oxygen species.20 NQO1 can also function as an activating enzyme, specifically for the reductive activation of antitumor quinones and other bioreductive anticancer agents.21

NQO1 can be induced by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and polycyclic aromatic hydrocarbons.21 Induction by procarcinogens and the capacity of NQO1 for detoxifying reactive metabolites suggest that changes in expression of NQO1 may occur during carcinogenesis.22 Marked elevations in NQO1 activity and mRNA content have been documented in both preneoplastic tissues and established tumors. Tumors or cancer cell lines where increased NQO1 expression has been observed include those from the lung, liver, colon, and breast.23 In addition, advanced, metastatic tumors appear to express higher levels of NQO1 than non-metastatic tumors.24

Correlations between NQO1 activity in cancer cells and cytotoxicity of antitumor quinones to those cells have been reported.25, 26, 27 In our previous studies with several series of indole- and quinolinequinone antitumor agents, including analogues of lavendamycin, those compounds that were the best substrates for NQO1 were also found to be selectively toxic to cell lines with high levels of NQO1 compared to cell lines that were deficient in NQO1.15, 16, 28, 29, 30 In this report, we describe the synthesis, metabolism by recombinant human NQO1 and anticancer activity of novel analogues of lavendamycin with functional group substitutions on both the quinoline-5,8-dione and indolopyridine moieties.

Section snippets

Synthetic chemistry

Table 4 presents the structures of a total of 25 lavendamycins that were the subject of our biological studies in this report. As shown in Scheme 1, Pictet–Spengler (P–S) condensation of the quinolinedione aldehydes (Table 2) with tryptophans (Table 3) yielded the target lavendamycins 4058 listed in Table 4. The amino lavendamycins 5964 were prepared respectively by the acid hydrolysis of the corresponding 7-acylamino lavendamycins 40–45.

High resolution mass spectroscopy of 67 (Scheme 2) did

Conclusions

Twenty-five novel lavendamycin analogues were synthesized, characterized and evaluated as NQO1-directed antitumor agents. In line with our earlier computational and structure–activity studies,15, 16 analogues with small substituents at the R1 position and groups capable of hydrogen bonding/van der Waals interactions at the R3 position generally made the best substrates for NQO1. Consequently, many of these compounds were also selectively toxic to the cancer cells with elevated NQO1 activity.

General methods

For general methods, see Ref. 16. In nearly all of the experimental work-ups, solvents were evaporated under reduced pressure and heat using a rotaevaporator. In each experiment, the progress of the reaction was monitored by TLC and the reaction stopped when TLC showed its completion. In some instances, no NMR signals were observed for active H’s, especially when DMSO-d6 was the solvent of use.

7-N-Formamido-2-formylquinoline-5,8-dione (20)

In a 25 mL two-necked round-bottomed flask, equipped with a stirring bar, a condenser and an argon

Acknowledgments

The financial support by the National Institutes of Health is greatly appreciated (NIH Grants: R15CA74245, M.B.; R15CA78232 and P20RR017670, H.D.B.). We also thank professor David Williams and the staff at the mass spectroscopy laboratory at Indiana University for their help in obtaining mass spectral data.

References and notes (34)

  • T.W. Doyle et al.

    Tetrahedron Lett.

    (1981)
  • A.S. Kende et al.

    Tetrahedron Lett.

    (1984)
  • M. Behforouz et al.

    Bioorg. Med. Chem.

    (2007)
  • L. Ernster

    Methods Enzymol.

    (1967)
  • P. Talalay et al.

    Methods Enzymol.

    (2004)
  • D. Ross et al.

    Chem. Biol. Interact.

    (2000)
  • S. Danson et al.

    Cancer Treat. Rev.

    (2004)
  • N. Robertson et al.

    Eur. J. Cancer

    (1994)
  • T. Fryatt et al.

    Bioorg. Med. Chem.

    (2004)
  • D.M. Balitz et al.

    J. Antibiot.

    (1982)
  • W.R. Erickson et al.

    J. Am. Chem. Soc.

    (1985)
  • W.R. Erickson et al.

    J. Am. Chem. Soc.

    (1987)
  • K.V. Rao et al.

    J. Am. Chem. Soc.

    (1963)
  • C.A. Hackethal et al.

    Antibiot. Chemother.

    (1961)
  • D.L. Boger et al.

    J. Med. Chem.

    (1987)
  • W.L. Wilson et al.

    Antibiot. Chemother.

    (1961)
  • D.L. Boger et al.

    J. Org. Chem.

    (1985)
  • Cited by (41)

    • Biological activities of polypyridyl-type ligands: implications for bioinorganic chemistry and light-activated metal complexes

      2021, Current Opinion in Chemical Biology
      Citation Excerpt :

      These compounds exhibit antineoplastic activity against several cancer types and possess other biological effects such as neuroprotection, which may be related to their metal-binding capabilities. Additional naturally occurring chelators that have been extensively studied are eilatin [60], streptonigrin [61–65], and lavendamycin [66–68]. As these ligands contain substituents at the ortho-positions, they may exhibit a mechanistic connection to synthetic ligands with similar structures.

    View all citing articles on Scopus
    View full text