Novel Lavendamycin Analogues as Potent HIV-Reverse Transcriptase Inhibitors: Synthesis and Evaluation of Anti-Reverse Transcriptase Activity of Amide and Ester Analogues of Lavendamycin

Article abstract of DOI:10.1021/jm0304414

Novel lavendamycins including two water soluble derivatives were synthesized via short and efficient methods. Pictet-Spengler condensation of 7-N-acylamino-2-formylquinoline-5,8-diones with tryptophans produced lavendamycin esters or amides 11-17. Lavenda

Full text of DOI:10.1021/jm0304414

J . Med. Chem. 2003, 46, 5773-5780
5773
Novel La ven d a m ycin An a logu es a s P oten t HIV-Rever se Tr a n scr ip ta se
In h ibitor s: Syn th esis a n d Eva lu a tion of An ti-Rever se Tr a n scr ip ta se Activity of
Am id e a n d Ester An a logu es of La ven d a m ycin
Mohammad Behforouz,* Wen Cai, Mark G. Stocksdale, J ennifer S. Lucas, J oo Yong J ung, Daniel Briere,
Aiqin Wang, Kevin S. Katen, and Nancy C. Behforouz*
Chemistry and Biology Departments, Ball State University, Muncie, Indiana 47306-0440
Received April 30, 2003
Novel lavendamycins including two water soluble derivatives were synthesized via short and
efficient methods. Pictet-Spengler condensation of 7-N-acylamino-2-formylquinoline-5,8-diones
with tryptophans produced lavendamycin esters or amides 11-17. Lavendamycins 18-21 were
obtained, respectively, by further transformations of 13-15 and 17. Several lavendamycins
were found to be potent HIV reverse transcriptase inhibitors with very low toxicity in vitro
and in vivo. Several compounds also acted either additively or synergistically to inhibit enzyme
activity together with AZT-triphosphate.
In tr od u ction
suppression, and limit the emergence of drug-resistant
HIV mutants.
Human immunodeficiency virus (HIV) infection, with
its clinical progression to AIDS, is one of the leading
causes of morbidity and mortality in the world.1 Al-
though highly active anti-retroviral therapy (HAART)
using a combination of anti-retrovirals has been highly
effective in suppressing HIV load and decreasing mor-
tality of these infections, the emergence of drug resis-
tance among HIV and inherent toxicity of most of the
anti-retroviral therapies makes the continued search for
novel anti-HIV drugs imperative.2 Indeed, it is now
estimated that over 30% of patients are failing combina-
tion therapy. Such failure is partly due to the develop-
ment of drug resistance and drug toxicity but also to
Earlier work has shown that two structurally related
antibiotics produced by Streptomyces, streptonigrin (1)
and lavendamycin (2a ), have significant biological activ-
the lack of patient compliance due to demanding drug
ity.7 Streptonigrin and several of its derivatives, par-
ticularly its alkyl esters, were shown to have potent
anti-reverse transcriptase activity against HIV-RT.8
Unfortunately both the streptonigrins and the parent
lavendamycin were found to be highly toxic. Until
recently, evaluation of the therapeutic potential of
synthetic lavendamycin analogues were precluded due
to the lack of efficient synthetic methods. Indeed, initial
reported total syntheses of lavendamycin methyl ester
involved a large number of steps with low overall yields
of 0.5-2%.9 In 1993 and 1996, we reported short and
efficient methods for the preparation of lavendamycin
methyl ester in excellent overall yields of 33-40%.10 We
now describe the total synthesis of a number of novel
demethyllavendamycin esters, amides as well as two
water-soluble derivatives via short and practical meth-
ods. This is the first report to describe the synthesis of
these lavendamycins possessing the full pentacyclic
structure of the system and its C-7 amino and C-2′ acid
functions and their derivatives. In addition, efficient
methods are introduced to synthesize water-soluble
lavendamycins essential for meaningful in vivo studies
and potential chemotherapeutic application. This is also
the first report showing the high degree of activity that
these novel nonnucleoside lavendamycin agents have in
inhibiting HIV-RT. Fortunately these inhibitors also
2a,3
dosing.
Currently approved HIV drugs include six
nucleoside HIV reverse transcriptase (HIV-RT) inhibi-
tors, zidovudine (AZT), didanosine(ddI), stuvudine (d4T),
lamivudine, (3TC), zalcitabine (ddC), and abacavir
(ABC); three nonnucleoside RT inhibitors, nevirapinne,
delavirdine, and efavirenz; and five protease inhibitors,
squinavir, indinavir, ritonavir, nelfinavir, amprenovir,
and lopiavir.3 Currently HAART using different com-
binations of these drugs is the preferred treatment for
AIDS patients.2a,4 Nonnucleoside HIV-1 reverse tran-
scriptase inhibitors (NNRTI) are becoming increasingly
important additions to this combination retroviral
therapy, and these compounds tend to have good
antiviral potency, high specificity, and low toxicity.5 It
has been suggested that NNRTI’s used in combination
regimens with protease inhibitors (PI) may be used in
the future either as first-line or second-line or salvage
therapy in patients who need to change anti-retroviral
treatments.6 Furthermore, anti-HIV combination strat-
egies that demonstrate favorable drug interactions (e.g.,
synergy) may allow the use of individual agents below
their toxic concentrations, provide more complete viral
* To whom correspondence should be addressed. M. Behforouz,
Chemistry Department, Ball State University, Phone: 765-2858070.
Fax: 765-2858804. E-mail: mbehforo@bsu.edu.
10.1021/jm0304414 CCC: $25.00 © 2003 American Chemical Society
Published on Web 11/21/2003

5774 J ournal of Medicinal Chemistry, 2003, Vol. 46, No. 26
Behforouz et al.
Sch em e 1
Sch em e 2
Ta ble 1. Structures of Lavendamycins, Reaction Conditions,
and Yields
Ta ble 2. Lavendamycins Derived from 13-15, 17
no.
R1
R2
R3
no.
R1
R2
OCH3
R3 % yield
CH3 79
solvent
xylene
anisole
anisole
h (°C )
18
19
20
21
CH3CO
CH3CO
H
OH
H
H
H
H
11 CH3CO
12 n-C3H7CO OC4H9-n
13 CH3CO
14 CH3CO
15 CH3CO
16 CH3CO
19 (reflux)
3.5 (reflux)
18 (152 °C)
OCH2CH2OPO3H2
OC8H17-n
H
H
54
72
OCH2Ph
OCH2CH2OH H
OC8H17-n
NH2
H
NH2
87-94 DMF/anisole 2 (reflux)
H
H
H
48
62
63
xylene
anisole
anisole
3 (reflux)
14 (reflux)
13 (reflux)
by the hydrogenolysis of 13 in the presence of Pd/black
in CH2Cl2-MeOH-THF mixture. The free amino lav-
endamycins 20 and 21 were obtained by the acid
hydrolysis of the corresponding 15 and 17, respectively,
in 64 and 96.5% yields, according to our reported method
for the conversion of 11 to lavendamycin methyl ester
(2b).10a Table 2 shows the structures of compounds 18-
21.
Tryptophan 5 was prepared according to our own
procedure. 11 Tryptophans 6, 7, 9, and 10 were obtained
by the neutralization of the commercially available salts
with ammonium hydroxide. The procedure for the prep-
aration of 1,2-ethanediol ester 8 is shown in Scheme 2.
Aldehyde 3 was prepared according to our previously
reported method10a,12 as shown in Scheme 3. A similar
procedure was used for the preparation of 7-butyramido-
2-formylquinoline-5,8-dione (4).
17 n-C3H7CO NH2
have low toxicity both in vitro and in vivo. In addition
to being inhibitory alone, several of the analogues also
display additive or synergistic activity against HIV RT
together with AZT-triphosphate. The synthesis of these
lavendamycin analogues is briefly described, and the
biological evaluation of these compounds presented.
Resu lts a n d Discu ssion
Syn th etic Ch em istr y. As shown in Scheme 1, Pic-
tet-Spengler condensation of 7-N-acylamino-2-formyl-
quinoline-5,8-diones 3 or 4 with â-methyltryptophan
methyl ester (5) or trytophan butyl, benzyl, 1,2-
ethanediol, octyl, esters (6-9), or tryptophan amide (10)
yielded the corresponding lavendamycin derivatives
11-17, respectively, in yields of 79, 54, 72, 94, 48, 62,
and 63% . In a typical procedure, aldehydes 3 or 4 (0.1
mmol) were mixed with the corresponding tryptophan
derivatives in 60 mL of dry anisole or xylene under
argon, and while being magnetically stirred, the mixture
was gradually heated to reflux over a period of 3 h. The
resulting clear solution was refluxed until TLC showed
the absence of the starting materials. The mixture was
concentrated or evaporated to dryness. The products
were either precipitated from the concentrated solu-
tions10a or purified by washing with acetone followed
by ether. The structures of the resulting lavendamycins,
the reaction conditions, and the yields are shown in
Table 1. Carboxylic acid 18 was prepared in 62% yield
Water soluble dihydrogen phosphate 19 was prepared
by the Mitsunobu method (Scheme 4).13
An ti-HIV Rever se Tr a n scr ip ta se a n d In Vitr o
a n d In Vivo Toxicit y St u d ies. The lavendamycin
analogues, 11, 12, 16, 18-21, were tested for their
ability to inhibit the HIV reverse transcriptase using a
modified assay of Okada et al.14 Briefly, the analogues
were initially dissolved in either DMSO or HEPES
buffer (pH 7.2) and then diluted to the appropriate
concentration before addition to the assay mixtures.
Varying concentrations of either AZT-TP or the ana-
logues or both were added to wells of a 24-well microtiter
3
plate containing 1 unit /mL HIV-RT, HTTP, the tem-
plate primer [poly(rA)-oligo (dT)12-18], and appropriate

Novel Lavendamycin Analogues as HIV-RT Inhibitors
J ournal of Medicinal Chemistry, 2003, Vol. 46, No. 26 5775
Sch em e 3
Sch em e 4
Ta ble 3. Inhibitory Activity of Lavendamycin Analogues on
HIV-RT and Cellular Cytotoxicity
water soluble analogues, namely the dihydogen phos-
phate 19, had the greatest activity with 15 µM inhibit-
ing the HIV-RT >92% (data not shown) with an IC50 of
3 µM. With the exception of compound 21, the concen-
trations at which 50% cytotoxicity (CC50’s) in tissue
culture were observed for the analogues were consider-
ably higher than the IC50’s toward both normal mouse
splenocytes and human lymphocytic cell lines. Further-
more, the levels of cytotoxicity for both human and
murine cells was comparable in most cases, which
allows for further testing in both murine and human
systems. At 10 and 20 µM concentrations, all the
analogues had a reversible inhibitory effect on normal
Con A-stimulated murine spleen cells, i.e., when the
drugs were removed from the cultures after 12 h, the
growth resumed at the same level as unexposed control
cells. This suggests that at the levels tested, the drugs
are cytostatic rather than cytocidal (data not shown).
As the human cells studied are lines permissive for HIV
replication and commonly used to examine the effective-
ness of potential anti-HIV therapeutic agents, these
compounds may be readily evaluated for anti-HIV
activity in culture.15 Furthermore, we found that BALB/c
mice both survived and tolerated the highly active anti-
HIV RT analogue 19 with little weight loss when given
either 50 mg/kg as one ip injection or 75 mg/kg given ip
in two injections within a 24 h period. In other studies,
we have found that several of the lavendamycin ana-
logues have remarkably low animal toxicity. For ex-
ample, the National Cancer Institute found that in mice
the maximum tolerated dose of 21 was 400 mg/kg and
15 was tolerated in SCID mice at 300 mg/kg/ day for 10
days with no animal death or loss of weight (data not
shown). Interestingly, lavendamycin and its analogues
appear to be much less toxic in experimental animals
than the related streptonigrin and its analogues.7b,16
Although the maximum tolerated dose of the naturally
occurring parent lavendamycin in mice has been re-
mean IC50
(µM)a ( SEM (normal mouse spleen)
compd (no. of expts)
mean CC50 (µM)b
mean CC50 (µM)c
(human cell lines)
( SEM (no. of expts) ( SEM (no. of expts)
11
12
16
18
19
20
21
15.1 ( 5.9 (3)
5.8 ( 1.5 (4)
12.7 ( 5.2 (6)
13.3 ( 4.4 (3)
3.0 ( 0.5 (3)
7.1 ( 2.7 (5)
20.5 ( 5 (3)
62 ( 5 (4)
81 ( 17 (4)
50 ( 7 (4)
47 ( 8.5 (4)
29 ( 5 (4)
48 ( 10 (4)
15 ( 2 (4)
31 ( 2 (4)
50 ( 15 (4)
95 ( 45 (4)
81 ( 35 (4)
27 ( 3 (4)
47 ( 14 (4)
17 ( 3 (4)
a
Mean concentration of analogue at which 50% inhibition of
b
HIV RT activity toward
1 unit of enzyme occurred. Mean
concentration of analogue at which normal mouse spleen stimu-
lated with concanavalin A was inhibited by 50%. c Mean concen-
tration of analogue at which human lymphocytic cell lines H-9 and
CEM was inhibited by 50%.
salts and buffers. Several control wells without any
inhibitors were also included on each microtiter plate.
The reactions were allowed to proceed for 1 h at 37 °C.
The amount of radioactivity incorporated under each
experimental condition was compared to that found in
the control wells and the amount of inhibition of the
HIV-RT was calculated for each analogue and, for
certain experiments, the inhibition found with each
combination of analogue and AZT-TP. A more detailed
description of reaction conditions are found in the
Experimental Section. The concentration of drug which
inhibited 50% of the enzyme activity (IC50)was deter-
mined by using the computer software program Cal-
cusyn, which plots the dose-effect curves from the
inhibition data and calculates the IC50 for each drug.
The results of this screen clearly show that several of
these analogues have significant anti-HIV RT activity
at levels considerably below the level which is cytotoxic
as measured either with murine lymphocytes or human
lymphocytic cell lines H-9 and CEM (Table 3). Ana-
logues 12, 19, and 20 were the most effective HIV-RT
inhibitors with IC50’s below 8 µM. One of the two more

5776 J ournal of Medicinal Chemistry, 2003, Vol. 46, No. 26
Behforouz et al.
ported to be 12.8 mg/kg, streptonigrin, which also has
anti-HIV RT activity, is lethal to mice at doses as low
as 0.4 mg/kg. The substantially lower level of animal
toxicity of lavendamycins as compared to stretonigrin,
another quinolinedione, may be due to the presence of
the â-carboline moiety in the lavendamycins. Other
quinonoid agents such as mitomycin C and adriamycin
are toxic at levels far below those we have observed with
the lavendamycins. In one reported tumor treatment
trial in mice, the maximum tolerated dose of mitomycin
C was less than 2 mg/kg when given for five consecutive
days.17 Likewise, adriamycin (doxorubicin) reportedly
has a maximally tolerated dose below 10 mg/kg when
given to mice once a week for three weeks.18
Com bin ed Activity of An a logu es w ith Azid oth y-
m id in e Tr ip h osp h a te. As combined drug therapy is
an essential feature of AIDS treatment today, it is
critical to assess the effect that any new drug may have
on the activity of other mainstay anti-HIV treatments
such as AZT-TP. Thus, we examined in a checkerboard
fashion the combined effect of the lavendamycin ana-
logues with AZT-triphosphate (AZT-TP) over a range
of concentrations for both drugs. The results of this
evaluation are presented in Table 4 and represent two
separate trials for each of the analogues 11, 12, 16, 18-
21. The results were analyzed by the method of Chou
and Talalay19 based on the median-effect principle and
represented by combination indices (CI) determined for
each of the two drug combinations. This method involves
the plotting of dose-effect curves for each agent and
for each of the different combinations of the agents. The
slope of the median-effect plot, which signifies the shape
of the dose-effect curve, and the x intercept of the plot,
which signifies the potency of each compound alone and
each combination, were then used for a computerized
calculation of a combination index. Concentrations of
AZT-TP were used which were both slightly below or
slightly above the IC50 (0.012 µM) and concentrations
of the lavendamycins were well below the CC50 of each
of the analogues. Many combinations of the lavenda-
mycins with AZT-TP were found to be either synergistic
(CI e 0.7) or moderately or slightly synergistic (CI g
0.7e 0.9), particularly at the lower concentrations of the
analogues. Additive effects were also seen for many of
combinations and some antagonism for some combina-
tions, particularly at the higher concentrations of both
drugs (e.g. 18 at 1.5 µM together with AZT-TP at 0.02
µM). The greatest amount of synergism over a wide
range of concentrations was observed with compounds
18-20. Interestingly, the increased water solubility of
compounds 18 and 19 may play a role in the increased
synergism with AZT -TP (see Experimental Section for
solubilization of analogues). Clearly, this entire series
of lavendamycin analogues have the ability to inhibit
the HIV-RT together with AZT in either additive or
synergistic manner. To derive the dose reduction index
of several of these compounds, the method of Chou and
Talalay was again employed using the classic isobolo-
gram technique. This method also involves the plotting
of dose-effect curves for each agent as well as for
multiply diluted fixed-ratio combinations of the agents.
The inhibitory capacity of 12, 18-20 with AZT-TP were
evaluated by this method, and the determination of CI’s
at 50% and 70% effect level as well as the dose reduction

Novel Lavendamycin Analogues as HIV-RT Inhibitors
J ournal of Medicinal Chemistry, 2003, Vol. 46, No. 26 5777
Ta ble 5. Two Drug Combination of AZT and Lavendamycin
Analogues at Constant Molar Ratios
lavendamycin analogues, the final reaction mixture contained
100 mM Tris-HCl (pH 8.0), 5mM MgCl2, 5 mM dithiothreitol,
60 mM NaCl, 0.2 mM [3H] thymidine triphosphate (5 µCi well),
4 µg/mL poly (rA)-oligo(dT)12-18 (Amersham Pharmacia Bio-
tech, Piscataway, NJ ), and 1 unit /ml of HIV-RT (Worthington
Biochemical Corp., Freehold, NJ ) in a total volume of 300 µL.
For each assay, the reaction mixture containing 8×buffer,
double distilled H2O, 3HTTP, template primer, and HIV-RT
was premixed in a 15 mL sterile tube in quantities required
for the particular experiment. After addition of the enzyme,
the reaction mixture was mixed and kept on ice. The inhibitors,
either AZT (Moravek Biochemicals, Brea, CA), analogues, or
both were added to appropriate wells in a 24-well microtiter
plate and ddH2O was added to each well to bring the volume
of all test wells to 100 µL. Two hundred microliters of the
chilled reaction mixture was then added to each of the test
wells, mixed, and incubated for 1 h at 37 °C. At least 9 control
wells with no inhibitors were included on each test plate. After
stopping the reactions with 25 µL of 0.1 M EDTA, triplicate
samples of 15 µL from each test well were spotted on DE81
ion exchange filter paper squares (Whatman Paper, Maidstone,
England). After drying, each filter was washed for 10 min three
times with 5% TCA-NaH2PO4, three times with 0.6 M NaCl-
0.06 mM Na citrate, one time with ddH2O, and one time with
95% ethanol. After drying, the individual filter papers were
added to 5 mL Scintiverse E (Fisher Scientific, Chicago, IL)
scintillation fluid and the amounts of 3H-TTP uptake mea-
sured as counts per minute on a Beckman liquid scintillation
counter. Mean uptake obtained with triplicate wells containing
inhibitors were compared with the mean uptake found in at
least three sets of triplicate control wells containing enzyme
plus substrate, and percent inhibition for each experimental
condition was calculated. The results were then analyzed using
Calcusyn, a windows software program for dose effect analysis
designed by T.-C.Chou and M. P. Hayball and published by
Biosoft, UK.
dose reduction indexb
CI at HIV-RT inhibition of:a
(50%)
molar ratio
AZT:analogue
50%
70%
AZT-TP analogue
12
18
19
20
1:600c
1:857d
1:600
1:857
1:600
1:857
1:214e
1:600
0.45 ( 0.06
0.39 ( 0.06
1.03 ( 0.29
1.14 ( 0.49
0.32 ( 0.08
0.32 ( 0.09
1.35 ( 0.64
1.7 ( 1.49
3.0
3.9
1.5
1.7
8.2
7.3
2.6
2.0
<0.1 ( 0.10 <0.1 ( 0.16
>100
>40
4.08
2.4
>100
>15
4.63
13.7
<0.1 ( 0.15
0.46 ( 0.11
0.49 ( 0.17
0.21 ( 0.18
0.56 ( 0.22
0.40 ( 0.09
a
Combination index at the effect level of 50% and 70% as
calculated by the method of Chou and Talalay using a computer
software program on an IBM PC.19 Additive effect (CI ) 1),
synergism (CI < 1), or antagonism (CI > 1) b Dose reduction index
is a measure of how much (how many times less) the dose of each
drug in a synergistic combination may be reduced at the 50% effect
level compared with the doses of each drug alone.19b c At least
four different combinations of the analogues and AZT-TP were
tested for HIV-RT inhibition whereby the ratio of the two drugs
was kept constant at 1:600 but the concentrations increased from
0.005 µM AZT-TP and 3 µM analogue to 0.04 µM AZT-TP and 24
d
µM analogue. At least four different combinations of the ana-
logues and AZT-TP were tested for HIV-RT inhibition whereby
the ratio of the two drugs was kept constant at 1:857, but the
concentrations increased from 0.0035 µM AZT-TP and 3 µM
analogue to 0.28 µM AZT-TP and 24 µM analogue. e Four different
combinations of analogues were tested whereby the ratio of the
two drugs was kept constant at 1:214, but the concentrations
increased from 0.0035 µM AZT-TP and 0.75 µM to 0.028 µM AZT-
TP and 6 µM analogue.
index (DRI) at 50% inhibition is shown in Table 5. The
dose reduction index is a measure of how much the dose
of each drug in a synergistic combination may be
reduced at a given effect level compared with the doses
of each drug alone.19 The combination of AZT-TP and
each of the four analogues 12, 18, 19, and 20 lead to
considerable dose reduction with the highly soluble
analogue 19 showing both the highest level of synergism
at the 50th and 70th percent inhibition level and
greatest dose reduction index in this in vitro system.
In summary, a new family of biologically active
lavendamycins including water soluble derivatives were
synthesized through short and practical methods. Sev-
eral of these lavendamycins have low toxicity, potent
anti-reverse transcriptase activity themselves, and are
synergistically inhibitory in combination with AZT-TP.
In particular, the water soluble analogue 19 had the
greatest inhibitory activity both alone and with AZT-
TP and was remarkably nontoxic in animals. It may be
that these compounds, particularly those with greater
water solubility, will also show similar activity toward
the replication of HIV in tissue culture and these or
other derivatives may become useful adjuncts to our
nonnucleoside anti-HIV arsenal.
Cytotoxicity Assa ys. Normal murine spleen cells were
washed in HBSS, cultured at 2.5 × 106 or 5 × 106 cells per
mL in RPMI 1640 (with Hepes) complete medium with 2mM-
L-glutamine, 1 mM sodium pyruvate, 0.05 mM â-mercaptoet-
hanol, penicillin, streptomycin, and amphotericin B with 10%
fetal calf serum and concanavalin A (ConA) at 2.5 µg/mL in
96-well microtiter plate. A range of diluted analogues were
added to triplicate culture wells, and the cells were incubated
at 37 °C in 5% CO2. Wells containing cells, media, and ConA
were used as controls. At 48 h postincubation, 20 µL of the
vital dye MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetra-
zolium bromide; thiazolyl blue) at 5 mg/mL was added to each
well, and the cultures were reincubated for 4-6 h. Following
centrifugation, the supernatants were decanted, and the
stained cells were resuspended in 100 µL of 0.4% HCl acidified
2-propanol. The optical densities of each well were read at 600
nm using a Cambridge Technologies series 700 microplate
reader. The human lymphocytic cell lines CCRF-CEM
(ATCC: HTB 176) and H-9 (ATCC: CCL 119) were subcul-
tured from recently cultured cells grown in complete RPMI
1640 medium as described above but without the â-mercap-
toethanol and ConA at 6.25 × 105 and 2 × 106 per mL in a
96-well culture plate. These cell numbers represented log
phase and early stationary phase of the freshly cultured cells.
As these cell lines rapidly reproduce without mitogenic stimu-
lation, Con A was not used. A range of analogues or media
alone was added in triplicate to the cultures and the cultures
incubated, pulsed with MTT, and harvested, and their growth
was estimated as described above for the murine cultures. The
CC50 or the concentration of drug which inhibited cell growth
as estimated by MTT formazan production to 50% of that of
untreated control cells was determined by linear interpolation
for each lavendamycin analogue. As the results from the
cytotoxicity assays of the two human cell lines were not
significantly different, the data was pooled and analyzed
together.
Exp er im en ta l Section
HIV-R T Assa y. Adapted from Okada et al.14 Except for
analogue 19, the lavendamycin analogues were initially dis-
solved in DMSO at a concentration of 1 mg/mL. Occasionally
mild sonication and heat were used to dissolve the less soluble
compounds. As 19 had greater water solubility, it was initially
dissolved in 2.5 mM HEPES buffer (pH 7.2) at a concentration
of 0.05 mg/mL. Compound 18 also had higher solubility readily
dissolving in DMSO at 1 mg/mL without any heating or
sonication. The dissolved analogues were then serially diluted
in distilled water to achieve concentrations in the final reaction
mixture ranging from 0.5 to 15 µM. In addition to the
In Vivo Toxicity. Analogue 19 was first solubilized in 10%
dry DMSO, diluted in water, and given to the mice intraperi-

5778 J ournal of Medicinal Chemistry, 2003, Vol. 46, No. 26
Behforouz et al.
toneally either in one dose or in two doses per day. Mice were
weighed and observed for 3 days following treatment.
Hydrogenator, this mixture was shaken under 30 psi of
hydrogen for 15 h. The catalyst was removed, and the dark
red solution containing the dihydrochloride salt of 5,7-diamino-
8-hydroxy-2-methylquinoline was placed in a 250 mL round-
bottomed flask equipped with a magnetic bar. To the stirred
solution, were added as quickly as possible, sodium sulfite (12
g), sodium acetate (16 g), and butyric anhydride (65 mL). The
thick whitish solid that continued to form over a 3-h period
was filtered, washed with water, and dried under vacuum (7.4
g, 93%). Attempts to recrystallized 26 from methanol-water
caused it to hydrolyze to the corresponding 5,7-dibutyramido-
8-hydroxyquinoline-5,8-dione. However, NMR showed the
crude 26 to be relatively pure and was used as such for the
preparation of 28. Analytical data for 26: mp 195-200 °C; Rf
) 0.26 (0.1/5 MeOH/EtOAc); 1H NMR (DMSO-d6) δ 0.92 (t,
3H, J ) 8.0), 0.96 (t, 3H, J ) 8.0), 1.08 (t, 3H, J ) 8.0), 1.52-
1.72 (m, 4H), 2.58 (s, 3H), 2.70 (t, 2H, J ) 8.0), 7.37 (d,1H, J
) 8.8), 8.21 (d, 1H, J ) 8.8), 8.24 (s, 1H), 9.65 (br s, 1H), 9.94
(br s, 1H).
5,7-Dibu tyr a m id o-8-h yd r oxy-2-m eth ylqu in olin e. This
compound was obtained by dissolving 26 in a minimum
amount of hot methanol-water (1/1).10b Pale white crystals
were filtered and dried: mp 200-208 °C (dec); Rf ) 0.32 (0.1/5
MeOH/EtOAc); 1H NMR (DMSO-d6) δ 0.93 (t, 3H, J ) 7.0),
0.96 (t, 3H, J ) 7.0), 2.3-2.55 (m, 4H), 2.7 (s, 3H), 7.3 (d, 1H,
J ) 8.8), 8.03 (3, 1H), 7.37 (d, 1H, J ) 8.8), 9.45 (br s, 1H),
9.63 (br s, 1H); EIMS, m/z, 329 (95), 286 (20), 259(98), 241
(17), 216 (6), 188 (100); HRMS, m/z calculated for C18H23N3O3
329.173942, found 329.173043; Analysis calculated C, 65.63;
H, 7.04; N 12.76; found C, 65.80; H, 7.05; N, 12.75.
Ch em istr y. Gen er a l Meth od s. Chemical reagents and
solvents were purchased from Aldrich, Sigma, and Fisher
Chemical companies and used as received unless otherwise
noted. Dioxane, xylene, anisole, and THF were distilled from
sodium/benzophenone. DMF was dried and distilled over
calcium hydride. Ammonium formate was dried in a vacuum
desiccator over calcium sulfate. Analytical TLC was performed
on 0.1 mm Eastman Kodak and Baker silica gel 60 F254 plates.
Baker silica gel (particle size 4.0 µm) was used for flash column
chromatography. NMR spectra were recorded on Varian
Gemini 200 and J EOL 400 spectrophotometers and calibrated
by using the residual undeuterated solvent as internal stan-
dard. Chemical shifts (δ) are in ppm, and coupling constants
(J ) are in Hz. Infrared spectra were recorded on a Perkin-
Elmer 1000 series FT-IR spectrometer. Low resolution mass
spectra and HRMS were recorded on Kratos MS 80, and the
relative peak intensities are given in parentheses. Elemental
analyses were performed by the Midwest Microlabs. Ltd.
Melting points are uncorrected. Each new compound showed
a single TLC spot (SiO2, unless indicated otherwise), was pure
as shown by NMR, and gave excellent elemental analyses or
HRMS data.
7-Aceta m id o-2-for m ylqu in olin e-5,8-d ion e (3). This is a
known aldehyde and was prepared from dinitro 24 according
to our previously reported method (see Scheme 3).10b
7-Bu tyr a m id o-2-for m ylqu in olin e-5,8-d ion e (4). In a 25
mL round-bottomed flask equipped with a magnetic bar, a
condenser, and an argon filled balloon, 7-butyramido-2-meth-
ylquinoline-5,8-dione (516 mg, 2 mmol), selenium dioxide (255
mg, 2.3 mmol), 12 mL of dried distilled 1,4-dioxane, and 0.25
mL of water were stirred and slowly heated to reflux over 2 h.
The reaction mixture was refluxed for 13 h, the black selenium
metal was allowed to settle, and the supernatant was pipetted
off and filtered. Dioxane (10 mL) was added to the solid
residue, stirred, and refluxed for 5 min. The entire mixture
was filtered, and the selenium was washed with dichlo-
romethane (10 mL). To the combined filtrates was added 50
mL dichloromethane, and they were washed with 3% sodium
bicarbonate solution (2 × 50 mL), dried (MgSO4), and evapo-
rated in vacuo to give 356 mg, (65%) of a pale yellow solid.
The product was recrystallized from ethyl acetate: mp 208-
210°C; Rf ) 0.39 (1/1 EtOAc/CH2Cl2); 1 H NMR (CDCl3): δ 1.02
(t, 3H, J ) 7.4), 1.70-1.89 (m, 2H), 2.52 (t, 2H, J ) 7.4), 8.06
(s,1H), 8.31 (d, 1H, J ) 8.0), 8.39 (br s, 1H), 8.61 (d, 1H, J )
8.0), 10.28 (s,1H); EIMS, m/z, 272 (54), 202 (36), 175 (9);
Analysis for C14H12N2O4 calculated C, 61.76; H, 4.44; N, 10.29;
found C, 61.31; H, 4.36; N, 9.94.
Tr yp top h a n â-Hyd r oxyeth yl Ester (8). In a 250 mL two-
necked round-bottomed flask was dissolved compound 23
(2.088 g, 5.4 mmol) in 100 mL dry DMF under an argon
balloon. Dry ammonium formate (1.124 g, 17.82 mmol) and
964 mg of 10% Pd/C were added, and the mixture was stirred
for 4.5 h at room temperature. The reaction mixture was
filtered through a layer of Celite, and the filter cake was
washed with 3 × 8 mL of EtOAc. The light orange solution
was evaporated in vacuo and dried under a vacuum pump at
50-60 °C for two or more days until no trace of DMF was
observed in the product NMR. The product was a gel and
1
weighed 1.164 g (87%); Rf ) 0.25 (0.25/5 MeOH/CH2Cl2); H
NMR (DMSO-d6) δ 2.95 (dd, 1H, J ) 14.1, 6.2), 3.04 (dd, 1H,
J ) 14.1, 6.0), 3.60-3.70 (m, 2H), 3.63 (dd, 1H, J ) 6.0, 6.2),
3.90-4.10 (m, 2H), 6.97 (dd, 1H,J ) 8.1, 7.0), 7.05 (dd, 1H, J
) 8.1, 7.0), 7.14 (s, 1H), 7.33 (d, 1H, J ) 8.1), 7.51 (d, 1H, J )
8.1), 7.95 (s, 1H),10.86 (br s, 1H); HRMS(EI) calculated for
C
13H16N2O3 248.1161, found 248.1158.
N-(Ben zyloxycar bon yl)tr yptoph an â-Hydr oxyeth yl Es-
ter (23). To partially dissolved CBZ-tryptophan (22, 1.02 g, 3
mmol) in 18 mL of n-butyl ether in a 50 mL three-necked
round-bottomed flask under argon were added 2-chloroethanol
(483 mg, 6 mmol) and triethylamine (0.4 mL, 291 mg, 2.9
mmol) via a syringe. The reaction mixture was refluxed
overnight for 5 h. The two-layered mixture was evaporated in
vacuo, and the residue was dissolved in 12 mL of EtOAc and
placed in the refrigerator overnight. The white solid material
was filtered and rinsed with EtOAc, and the filtrate was
washed with 5 mL of a 5% solution of sodium bicarbonate
followed by water until the pH of the aqueous layer reached 7
(5 × 3 mL). The organic solution was dried (MgSO4), filtered,
and evaporated in vacuo and then dried on a pump at 40-50
°C to give a dark orange gel (880.4 mg, 78%); Rf ) 0.44 (1/5
7-Bu t yr a m id o-2-m et h ylq u in olin e-5,8-d ion e (28). In a
500 mL round-bottomed flask equipped with a magnetic bar,
5,7-dibutyramido-8-Butyroxy-2-methylquinoline (26, 3.29 g,
8.25 mmol) was suspended in 122 mL of glacial acetic acid. A
solution of potassium dichromate (8.8 g, 30 mmol) in 115 mL
of water was added and stirred at room temperature. Dichlo-
romethane (70 mL) was added to promote solution, and the
resulting two-phase mixture was stirred overnight. The or-
ganic layer was separated, and the aqueous portion was
extracted with dichloromethane (12 × 50 mL). The combined
extracts were washed with 200 mL of 3% sodium bicarbonate
solution, dried (MgSO4), evaporated, and dried under vacuum
to give an orange yellow solid (1.65 g, 77%). The product was
recrystallized from ethyl acetate: mp 188-189 °C; Rf ) 0.54
(1/1 EtOAc/CH2Cl2); 1H NMR (CDCl3): δ1.00 (t, 3H, J ) 7.4),
1.69-1.82 (m, 2H), 2.48 (t, 2H, J ) 7.4), 2.74 (s, 3H), 7.53 (d,
J ) 8.0), 7.90 (s, 1H), 8.28 (d, 1H, J ) 8.0), 8.36 (br s, 1H);
EIMS, m/z, 258 (81), 215 (7), 188 (100), 161 (66); Analysis for
1
EtOH/EtOAc, Al2O3); H NMR (CDCl3) δ 3.20-3.40 (m, 2H),
3.55-3.70 (m, 2H), 4.00-4.25 (m, 2H), 4.60-4.80 (m, 1H),
5.00-5.15 (m, 2H), 5.25-5.45 (m, 2H), 7.02 (s, 1H), 7.05-7.15
(m, 1H), 7.17-7.25 (m, 1H), 7.25-7.4 (m, 5H), 7.50-7.60 (m,
1H), 8.09 (br s, 1H). HRMS(EI) calculated for C21H22N2O5
382.1528, found 382.1523.
C
14H14N2O3 calculated C, 65.11; H, 5.46; N, 10.85; found C,
65.22; H, 5.51; N, 10.90.
5,7-Dibu tyr a m id o-8-bu tyr oxy-2-m eth ylqu in olin e (26).
In a 500 mL heavy-walled hydrogenation bottle, 5.00 g (0.2
mmol) of finely powdered 8-hydroxy-2-methyl-5,7-dinitro-
quinoline10a and 5% Pd/C (1.5 g) were suspended in 100 mL of
water and 12 mL of concentrated hydrochloric acid. In a Parr
7-N-Acetyld em eth ylla ven d a m ycin â-Hyd r oxyeth yl Es-
ter (14). In a 500 mL three-necked round-bottomed flask
equipped with a Dean-Stark trap, a condenser, a dropping
funnel, and a magnetic bar, 7-acetamido-2-formylquinoline-
5,8 -dione (3, 146.4 mg, 0.6 mmol) in 250 mL dry anisole was

Novel Lavendamycin Analogues as HIV-RT Inhibitors
J ournal of Medicinal Chemistry, 2003, Vol. 46, No. 26 5779
stirred and heated under argon to 80 °C over 30 min. Nearly
all of aldehyde 3 was solubilized. To this mixture was dropwise
added a solution of tryptophan â-hydroxyethyl ester (8,198.64
mg, 0.8 mmol) in 12 mL of dry DMF over 40 min at 95 °C,
and the temperature was slowly raised to 160 °C over 4.5 h
and then heated at this temperature for an additional 1 h.
The reaction mixture was allowed to cool for 10 min and
then filtered to remove a small amount of colloidal material.
The filtrate was evaporated in vacuo to dryness, and then 15
mL acetone was added and allowed to stand at room-temper-
ature overnight. The solid material was filtered and washed
with acetone followed by ether to give 177 mg (94%) of yellow
orange solid 14: mp 270.5-271 (dec); Rf ) 0.69 (0.2/5 MeOH/
CH2Cl2); 1H NMR (DMSO-d6): δ 2.32 (s, 3H), 3.80-3.90 (m,
2H) 4.40-4.50 (m, 2H), 7.45 (t, 1H, J ) 7.3), 7.72 (d, 1H,J )
7.3), 7.70-7.75 (m, 1H), 7.83 (s, 1H), 8.53 (d, 1H, J ) 8.0),,
8.61 (d, 1H, J ) 8.4), 8.98 (d, 1H, J ) 8.4) 10.31 (s, 1H), 12.01
(br s, 1H); HRMS(EI) calculated for C25H18N4O6 470.1226,
found 470.12406.
Diben zyl P h osp h a te 29. To a stirred mixture of 14 (135.7
mg, 0.289 mmol), dibenzyl hydrogen phosphate (241.2 mg, 0.87
mmol), and triphenyphosphine (228.2 mg, 0.87 mmol) in 8 mL
dry THF, under Ar, was dropwise added a solution of diethyl
azodicarboxylate (151.52 mg, 0.87 mmol) in 2 mL dry THF via
a syringe over a course of 12 min. The mixture was stirred for
4 h at room temperature, and the resulting solid was filtered
off. The filtrate was evaporated to dryness, and the solid was
washed with ethyl ether to give 179 mg (85%) of orange solid
29. Silica gel chromatography (1% MeOH in CH2Cl2) gave 142
mg of pure 29: mp 183.5-184°C; Rf ) 0.46 (EtOAc); 1H NMR
(CDCl3) δ 2.36 (s, 3H), 4.45 (br s, 2H), 4.64 (br s, 2H), 5.07 (s,
2H), 5.12 (s, 2H) 7.20-7.34 (m, 10H), 7.40 (dd, 1H, J ) 8.0,
7.3), 7.68 (dd, 1H, J ) 8.0, 7.3), 7.76 (d, 1H, J ) 8.0), 8.01 (s,
1H), 8.20 (d, 1H, J ) 8.0), 8.45 (s, 1H), 8.48 (d, 1H, J ) 8.3),
8.99 (s, 1H), 9.18 (d, 1H, J ) 8.3), 11.85 (br s, 1H); 31P NMR
(CDCl3): δ 1.69; HRMS(EI) calculated for C39H34N4O9P (M+3)
733.2065, found 733.2065.
7-N-Acetyld em eth ylla ven d a m ycin Ben zyl Ester (13).
In a 500 mL three-necked round-bottomed flask under argon
was dissolved 7-acetamido-2-formylquinoline-5,8-dione (3, 146.4
mg, 0.6 mmol) in 300 mL of dry anisole and heated to 100 °C.
To this mixture was dropwise added tryptophan benzyl ester
(7, 176.4 mg, 0.6 mmol) over 30 min. The yellow lemon solution
was gradually heated to 152 °C over 2 h and kept at this
temperature for 18 h. To convert any generated hyroquinone
intermediate to the desired product, a flow of oxygen gas was
passed through the lemon yellow hot solution for 20 min. The
resulting golden yellow mixture was filtered hot to remove
some impurities, evaporated in vacuo, and then left overnight
in a vessel open to the air. The small amount of the brownish
solid was removed, and the solution was concentrated to near
dryness. Acetone (10 mL) was added, and the resulting solid
material was filtered and washed with a small portion of
acetone and then petroleum ether to give a pure golden
crystalline solid (222 mg. 72%): mp 174.6-175 °C; Rf ) 0.68
(0.1/5 MeOH/CH2Cl2); 1H NMR (CDCl3) δ 2.36 (s, 3H), 5.55 (s,
2H), 7.35-7.48 (m, 5H), 7.58-7.62 (m, 2H), 7.64-7.7 (m, 1H),
7.72-7.78 (m, 1H), 8.00 (s, 1H), 8.23 (d, 1H, J ) 8.0), 8.43 (br
s, 1H), 8.56 (d, 1H, J ) 8.4), 8.98 (s, 1H), 9.21 (d, 1H, J ) 8.4),
11.83 (br s, 1H); HRMS(EI) calculated for C30H20N4O5 516.14347,
found 516.14340.
7-N-Acetyld em eth ylla ven d a m ycin (18). To a stirred
mixture of benzyl ester 13 (125.8 mg, 0.24 mmol) in dichlo-
romethane (186 mL), methanol (14 mL), water (0.01 mL) was
added 42 mg of palladium black, and the mixture was
hydrogenated under
a hydrogen filled balloon for 20 h.
Palladium was removed, and the solution was stirred under
an oxygen-filled balloon for 96 h and then evaporated in vacuo.
The solid was washed with acetone followed by ether and then
dried under vacuum to give 94.2 mg (92%) of brown orange
13: mp > 270 °C; Water solubility (0.05 mg/1 mL HEPES
buffer solution; Rf ) 0.34 (1/2 MeOH/CH3CN); 1H NMR
(DMSO-d6) δ 2.32 (s, 3H), 7.43 (t, 1H, J ) 7.7), 7.66-7.88 (m,
2H), 7.82 (s, 1H), 8.54 (d, 1H, J ) 7.7), 8.58 (d, 1H, J ) 8.4),
9.18 (d, 1H, J ) 8.4), 10.28 (s, 1H), 11.99 (br s, 1H); HRMS-
(EI) calculated for C23H14N4O5 426.0964, found 426.0959.
Dih yd r ogen P h osp h a te 19. To a solution of 29 (38. 8 mg,
0.053 mmol) in 7.4 mL of a mixture of CH2Cl2/EtOH/H2O (5/
2/0.4) was added 14.3 mg of palladium black, and the mixture
was stirred under a balloon full of hydrogen for 16 h. The
balloon was removed, the mixture was filtered, the filter cake
was thoroughly washed with 22 mL of a 1/1 mixture of EtOH/
CH2Cl2, and then the filtrate was stirred in the presence of
air for 2 h. The resulting filtrate was evaporated in vacuo to
dryness to yield 19.6 mg (84%) of pure orange brown 19: mp
179 °C (dec); Water solubility (0.05 mg/1 mL HEPES buffer
solution); Rf ) 0.54 (Eastman Kodak SiO2, 8/3/0.5 MeOH/H2O/
7-N-Acetyld em eth ylla ven d a m ycin n -Octyl Ester (15).
This compound was prepared according to a method similar
to that of 17. The final product was obtained by the evapora-
tion of the reaction mixture in vacuo, and the brown solid was
washed with a mixture of ethyl acetate-hexane to give the
orange solid 15 in 48%: mp 210.5-211 °C; Rf ) 0.41 (0.06/5
1
MeOH/CH2Cl2); H NMR (DMSO-d6) δ 0.86 (t, 3H, J ) 7.3),
1.25-1.6 (m, 10H), 1.83 (m, 2H), 2.32 (s, 3H), 4.41 (t, 2H, J )
6.6), 7.44 (dd, 1H, J ) 8.0, 7.3), 7.65-7.8 (m, 2H), 7.82 (s, 1H),
8.54 (d, 1H, J ) 8.0), 8.57 (d, 1H, J ) 8.2), 8.93 (d, 1H, J )
8.2), 9.09 (s, 1H), 10.29 (s, 1H), 11.96 (br s, 1H); EIMS, m/z,
538 (M+, 55), 382 (100), 366 (5), 340 (16); HRMS(EI) calculated
for C31H30N4O5 538.2216, found 538.2220.
1
EtOAc); H NMR (DMSO-d6): δ 2.32 (s, 3H), 4.27 (br s, 2H),
4.57 (br s 2H), 7.41 (dd, 1H, J ) 8.0, 7.6), 7.64 (d, 1H, J )
8.0), 7.71 (dd, 1H, J ) 8.0, 7.6), 7.79 (s, 1H), 8.47 (d, 1H, J )
7.7), 8.53 (1H, d, J ) 8.0), 8.91 (d, 1H J ) 8.0), 9.08 (s, 1H),
1026 (s, 1H), 11.89 (br s, 1H); 31P NMR (CDCl3); δ - 0. 02.
Dem eth ylla ven d a m ycim n -Octyl Ester (20). 7-N-Acetyl-
demethyllavendamycin n-octyl ester (15, 76.3 mg, 0.142 mmol)
was added to 7 mL of a 70% solution of sulfuric acid and under
an argon balloon was stirred and heated at 60 °C for 45 min.
The reaction mixture was carefully basified with a saturated
solution of sodium carbonate to pH ) 8 and then extracted
with chloroform (4 × 45 mL). The combined extracts were
washed with water (2 × 20 mL), dried (MgSO4), and concen-
trated to a small volume. The solution was cooled in the
refrigerator, and the red solid was filtered off. Silica gel flash
chromatography using chloroform and then methanol-chlo-
roform (1.6 mL/100 mL) as the solvent system gave the pure
orange red product 20 (45 mg, 64%): mp 175-176 °C (dec); Rf
) 0.12 (0.06/5 MeOH/CH2Cl2); 1H NMR (CDCl3) δ 0.96 (t, 3H,
J ) 7.3), 1.25-1.40 (m, 10H), 1.93 (m, 2H), 4.59 (t, 2H, J )
6.9), 7.10-7.30 (m, 1H), 7.35-7.43 (m, 1H), 7.51 (d, 1H, J )
8.0), 7.74 (br s,1H), 7.82 (d, 1H, J ) 8.0), 8.11 (d, 1H, J ) 8.0),
8.15 (s, 1H), 9.10 (d, 1H, J ) 8.0), 9.16 (s, 1H), 11.84 (br s,
1H); HRMS(EI) calculated for C29H28N4O4 496.2110, found
496.2099
HRMS(FAB) calculated for
550.0912.
C25H19N4O9P 550.0089, found
7-N-n -Bu t yr yld em et h ylla ven d a m ycin n -Bu t yl E st er
(12). Tryptophan n-butyl ester (6, 78.1 mg, 0.3 mmol) was
dissolved in 180 mL of dry anisole. To this solution under an
argon balloon was added 7-n-butyramido-2-formylquinoline-
5,8-dione (81.6 mg, 0.3 mmol), and the mixture was stirred
and heated to reflux over a course of 3 h. The reflux was
continued for another 3.5 h, and the solution was evaporated
in vacuo to dryness. The residue was dissolved in acetone and
allowed to stir at room temperature in the presence of air for
24 h. The yellow-orange product was filtered and dried under
vacuum (82.3 mg, 54%): mp 220.8-221 °C (dec); Rf ) 0.54
(0.5/5 EtOAc/CH2Cl2); 1H NMR (CDCl3) δ 0.82 (t, 3H, J ) 7.3),
0.95 (t, 3H, J ) 7.5), 1.42-1.6 (m, 4H), 1.75-1.86 (m, 4H),
4.56 (t, 2H, J ) 6.6), 7.15-7.35 (m, 2H), 7.56 (d, 1H, J ) 8.1),
7.85 (d, 1H,J ) 8.1), 7.89 (br s, 1H), 8.14 (d, 1H, J ) 8.1), 8.22
(s, 1H), 9.15 (d, 1H, J ) 8.1), 9.19 (s, 1H), 11.96 (br s, 1H);
EIMS, m/z, 510 (M+, 69), 446 (12), 417 (11.5), 410 (100), 394
(36), 340 (39), 217 (13); HRMS calculated for C29H26N4O5
510.1903, found 510.1886.
7-N-Acetyld em eth ylla ven d a m ycin Am id e (16). This
compound was prepared according to a method similar to the

5780 J ournal of Medicinal Chemistry, 2003, Vol. 46, No. 26
Behforouz et al.
Romero, D. L.; Busso, M.; Tan, C.-K.; Reusser, F.; Palmer, J .
R.; Poppe. S. M.; Aristoff, P. A.; Downey, K. M.; So, A. G.;
Resnick, L.; Tarpley, W. G. Non-Nucleoside Reverse Tran-
scriptase Inhibtors That Potently and Specifically Block Human
Immunodeficiency Virus Type 1 Replication. Proc. Natl. Acad.
Sci. U.S.A. 1991, 88, 8806-8810. (c) De Clerq, E. The Role of
Nonnucleoside Reverse Transcriptase Inhibitors (NNRTIs) in the
Therapy of HIV-1 Infection. Antiviral Res. 1998, 38, 153-179.
(6) J oly V.; Descamps, D.; Yeni, P. NNRTI Plus PI Combinations
in the Perspective of Nucleoside-Sparing or Nucleoside-Failing
Antiretroviral Regimens. AIDS Rev. 2002, 4, 128-139.
(7) (a) Take, Y.; Kubo, T.; Takemori, E.; Inouye, Y.; Nakamura, S.;
Nishimura, T.; Suzuki, H.; Yamaguchi, H. Biological Properties
of Streptonigrin Derivatives III. In Vitro and In Vivo Antiviral
and Antitumor Activities. J . Antibiot. 1989, 42, 968-976(b)
Balitz, D. M.; Bush, J . A.; Bradner, W. T.; Doyle, T.W.; O’Herron,
F. A.; Nettleton, D. E. Isolation of Lavendamycin, a New
Antibiotic from Streptomyces lavendulae. J . Antibiot. 1982, 35,
259-265.
procedure used for the synthesis of 17. The final product was
obtained as a lemon yellow solid in 62% yield by cooling the
reaction mixture in the refrigerator, filtration of the solid, and
washing with hexane: mp > 280 °C (dec); Rf ) 0.65 (0.2/5
MeOH/CH2Cl2); 1H NMR (DMSO-d6) δ 2.32 (s, 3H), 7.41 (d,
1H, J ) 7.5, 7.0), 7.6-7.75 (m, 3H), 7.82 (s, 1H), 8.48 (dd, 1H,
J ) 8.4), 8.52 (d, 1H,J ) 8.0), 8.61 (br s, 1H), 9.07 (s, 1H),
10.29 (s, 1H), 11.95 (br s, 1H); HRMS(EI) calculated for
C
23H15N5O4 425.1124, found 425.1115.
7-N-n -Bu tyr a m id od em eth ylla ven d a m ycin Am id e (17).
To a stirred solution of tryptophan amide (10, 243.6 mg, 1.2
mmol) in 480 mL of dry anisole under argon balloon was added
7-N-butyramido-2-formylquinoline-5,8-dione (4, 326.4 mg,1.2
mmol), and the mixture was heated to reflux over a 3-h period.
The reaction mixture was refluxed for 13 h and then allowed
to stand at room-temperature overnight. The yellow solid was
filtered off and washed with a small volume of acetone followed
by ether to give 343.3 mg (63%) of pure 17: mp 244 °C (dec);
Rf ) 0.58 (0.2/5 MeOHCH2Cl2); 1H NMR (DMSO-d6) δ 0.95 (t,
3H, J ) 7.3), 1.64 (quintet, 2H, J ) 7.3), 2.64 (t, 2H, J ) 7.3),
7.41 (dd, 1H, J ) 8.2, 7.9), 7.70-7.76 (m, 2H), 7.83 (s, 1H),
8.47 (d, 1H, 8.2), 8.51 (d, 1H, J ) 7.9), 9.05 (s, 1H), 9.45 (d,
1H, 8.2),10.18 (s, 1H), 11.92 (br s, 1H). HRMS(EI) calculated
for C25H20N5O4 (M + 1) 454.15153, found 454.15129.
(8) Take, Y.; Inouye, Y.; Nakamura, S. Allaudeen, H. S.; Kubo, A.
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on Human Immunodeficiency Virus and Avian Myeloblastosis
Virus Reverse Transcriptases and Cellular DNA Polymerases.
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Thompson, S. C. Streptonigrin and Lavendamycin Partial
Structures. Probes for the Minimum, Potent Pharmacophore of
Streptonigrin, Lavendamycin, and Synthetic Quinoline-5,8-
diones. J . Med. Chem. 1987, 30, 1918-1928. (b) Kende, A. S.;
Ebetino, F. H. The Regiospecific Total Synthesis of Lavenda-
mycin Methyl Ester. Tetrahedron Lett. 1984, 25, 923-926.
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A Highly Concise Synthesis of Lavendamycin Methyl Ester. J
Org. Chem. 1993, 58, 7089-7091. (b) Behforouz, M.; Haddad,
J .; Cai, W.; Arnold, M. B.; Mohammadi, F.; Sousa, A. C.; Horn,
M. A. Highly Efficient and Practical Syntheses of Lavendamycin
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Dem eth ylla ven d a m ycin Am id e (21). 7-N-n-Butyryldem-
ethyllavendamycin amide (17, 147.8 mg, 0.32 mmol) was
placed in a 50 mL two-necked round-bottomed flask under
argon balloon. A 70% solution of sulfuric acid (7.5 mL) was
dropwise added, and the homogeneous mixture was stirred and
heated at 60 °C in an oil bath for 6 h. The dark red solution
was cooled to 0 °C and then added to 75 mL of ice-water. The
mixture was carefully basified with a saturated solution of
sodium carbonate to about pH ) 8. The solution was evapo-
rated in vacuo to dryness and then water (75 mL) was added
and stirred. The orange red crystals were washed with water
and dried under vacuum (115.4 mg, 96.5%): 154.2 °C
(dec); Rf ) 0.45 (0.2/5 MeOH/CH2Cl2); 1 H NMR (DMSO-d6) δ
5.96 (s, 1H), 7.41 (t, 1H, J ) 7.3), 7.60-7.74 (m, 2H), 7.79 (d,
1H, J ) 8.0), 8.46 (d, 1H, J ) 8.0), 8.52 (d, 1H, J ) 7.3), 8.60
(s, 1H), 9.06 (s, 1H), 9.44 (d, 1H, J ) 8.0), 12.00 (br s, 1H);
HRMS(EI) calculated for
383.1026.
C21H13N5O3 383.1018, found
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Ack n ow led gm en t. Financial support by the Na-
tional Institutes of Health, Grant No. R01 CA74245 is
appreciated. We also thank Professor David Williams
and Mr. Mark Herbert of Indiana University for their
assistance in obtaining mass spectral data.
Su p p or tin g In for m a tion Ava ila ble: NMR spectra for
compounds 26, 8, 23, 14, 29, 19, 12, 13, 18, 15, 20, 16, 17,
and 21 are available free of charge via the Internet at http://
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