1341262
Process for the Synthesis of Vinblastine and Vincristine
Field of the Invention
The present application relates to a further
improvement of a new and improved method for producing dimer
alkaloid compounds especially of the Catharanthus (Vinca)
alkaloid groups and, in particular, is a further improved
method for producing the anti-viral, anti-leukemic
(antineoplastic) compounds, vincristine and vinblastine of
Formula I.
Iorrr.~rr>
o~~ v.~
frr~r.c)
I
The above compound, when R is COOCH3, and Rl is
OCH3, is vinblastine (NSC 49482) and when R is COOCH3, and
Rl is OCH3 and N1 is N-CHO (N-formyl), vincristine (NSC
67574).
The present series of dimeric alkaloids, including
important antitumor agents, are formed from an indole, such
as catharanthine (Formula II, R - COOCH3), and a
dihydroindole unit, e.g., vindoline (Formula III:), in which
1
~ 3 4 ~-z s z ._
the halves are linked via a carbon-carbon bond involving an
aliphatic center Cl~ in the indole unit and an aromatic
carbon C15 in the vindoline portion.
h a
C00C 1i
II III
Summarv of the Invention
The specific improvements in the present applica-
tion are set out below.
Firstly, improvement of the novel reduction
method, in which conversion of the iminium intermediate
(Formula VI) to the enamine (Formula VIII)
2
'~34i2fiZ
_,....
~N_~
1 O
~ ~nR = H..., ~.'-'
V ~doline
;'
ON
Cti3 D ~ " ~t~'~'~'' O~ -CH3
C~i3 R p
vi R=CC Ct~!
~o?N a i
O I ~ ~ _ .,., _~ _..,-
r.'
'~'~
I
O
CH30 ~' ~'' Oi -Cti3
CH3 R p
VI I I R=C02 CH3
~~34~2s2
as described in step (e), post, is achieved by the use of
various new 1,4-dihydronicotinamides (Formula IX, R2 - R4 -
R5 - R6 - H; R3 - CONH2 and R1 may contain different
functional groups ;such as aryl, caqrboxylic esters, sugars,
carboxylic acid, and carboxylate salt, as represented by
Formulae XXVII to XXXII listed in Table 1) and 1,4-
dihydropyridine compounds such as Formulae XXXIII and XXXIV.
Of special importance is the presence of electron rich
functional groups (e. g., carboxylic esters and carboxylate
salts in R1 of Formula IX, capable of coordination with the
positively charged iminium intermediate (Formula VI). Such
coordination increases both the regioselectivity (1,4-
reduction over 1,2-reduction ) and the rate of the reduction
of iminium intermediate (Formula VI), leading to an improved
yield of the enamine (Formula VIII). Further improvement
in the formation of the latter compound is achieved by
performing the reduction of iminium intermediate (Formula
VI) and maintaining the reaction mixture before any
subsequent manipulation at a low temperature (0 to -70°C)
preferably below -40°C.
p
Rs R3
Re R
z
R,
IX
4
'1341262
Specific examples related to the above improve-
ments for the synthesis of enamine (Formula VIII) by the
1,4-reduction of the iminium intermediate (Formula VI) are
given in the Examples 1-9 (Procedures C to K) . Results of
these examples are summarized in Tables 2 and 3. Table 2
indicates reduction procedures (Procedures I and J as in
Examples 7 and 8), employing reducing agents Formula XXXI
and Formula XXXII, respectively, afford the best yields of
the enamine (Formula VIII). Table 3 shows the effect of
temperature in the reduction of iminium intermediate
(Formula VI) by the reducing agent Formula XXXII. Lowering
the reduction temperature to -40°C (Procedure K(ii) Example
9) resulted in an increase in both the ratio of 1,4-
reduction production, enamine (VIII), versus 1,2-reduction
product, 3,4-dehydrovinblastine (VII), (4.2:1), as well as
the overall yield of the reaction (85~).
The second area of improvement relates to the
oxidative transformation of the enamine (Formula VIII) to
the iminium intermediate (Formula XVI).
Various parameters for this
oxidative transformation have been studied to optimize the
yield of vinblastine (Formula I) production.
Table 4 indicates the effect of ferric chloride
concentration. Two equivalents of ferric chloride provide
the highest yield of vinblastine (I).
13~12f~2
'
1
_
'r: E
z - o >
. ~ c
w
.r
G
~rl
w
an
N
v
a
G ..
r1 .,.1
N ~
r
.., .b
E3
w
I
o .J
G
O M
-.1 v
1~
U
'O
1
J
U
~ U~~ M
, O
,
'_" U
1
, I Z ~ U
.- . ~
w ~ 'z~' V '
~'
~ LL
Z1,~~ ~
.
'
_
.
O
U
:c
v
r
6
1341262
w
a
a~ A w
0
0.
N
~~ I
O a
p ~ C~
Z
- ~ .n ~ .n
v V U V V
p v _ H _
Z- U Z~
a
~4
a ~4 L1 fir V jp '" ~,i
0
>w.
a.
'~ ,,~ ,~ : O : v U ~o
v N ~ n ~ ~ < Z
Z O = t,~ c~, .p O c~ ~c
N ~r
p _ Z- ~ ~ ~ ~ o 0
v ...~,. ..,~.. 0G ~ ?e
b II V v II r r r
a
N
r1 r~
H h~l N
w
134122
Table 2. Effect of Reducing Agent on I,~- vs. 1,2-keduction of Iminiuu ~'i
Reduction Procedurel'4 1,4:1,2-Reduction Pzoducts' Yield3 (~)
E~ 1:1 75
1:1 60
0.9:1.0 60
v 1:1 40
1.1:1 60
2:1 65
1.5:1 70
1.1:1 65
I 2.3:1 70
J 2.2:1 70
1 Typical Procedure: 100 mg Iminiua VI in 6 ml methanol to which
reducing aitenta C-J (1-6 eq.) distolved in 6 ml :ethanol weze
added. F~11 details in Fxpeziaental Section.
2 B~ reverse phase HPLC quantitation.
Combined 1,2-reduction (3,4-dehydrovinblastine, VII) + 1,4-reduction
tenamiae VIII) products.
4 All of these reactions were co~ucted at 20° C.
Procedures A and B are resented in Canadian
p application Serial No.
57,897 and presented here for oar~parison.
~3~fizsz
Table 3. Effect of Temperature on 1,4- vs. 1,2-Reduction of Iminiug VI.
Reduction Procedural Teap. (°C) 1,4:1,2-Reduction Products2 Yield3
(t)
D 20 2.2:1 70
K (i) -20 3.2:1 80
X (ii) -40 4.2:1 85
1 Typical procedure as in Table 2.
2
Quantiution by IiPLC.
Cosbined 1,2-reduction (3,4-dehydrovinblastixe, VII) ~ 1,4-reduction
(examine VIII) products.
9
1341262
Table ~. Effect of Ferric C~t~loride on Production of Vinblastine (I) f rou
Ena~ine VIII3
Amount of FeCi; Orxidation Conditions ! Yield of
(Equivalentsl'' (Temp., time) Vinblastine2
p Aizi, Oo C, 5 min 0
I Air , 0 C, 5 min 13.3
l
Air C, S min 19.0
, 0
3 Airl, 0 C, 5 min 10.4
pt a rate of 6U ml/min.
B~ revezee-phase HPLC quantiution, after reductive work-up with NaBH4.
3 Ensmine VIII itenerated at -4D° C (Procedure K (ii)).
1341'262
Table 5. Effect of Time of Oxidation on Production of Vinblastine (I) froa
Enamine VIII3
Timel (min) a Yield of 2
Vinblastine
1 8.2
15.4
15.5
15.7
45 6.5
Reaction conditions: - 2 eq. ferric chlorine added, air bubbled
throu,~ the solution at 60 ~1/min at 0° C.
2 By reverse-phase BpI~C quaatitation aftxr reductive work-up s~ith NaBIi4.
~namine DIII generated at -40° C (Procedure K (ii)).
11
13412fi2
~abie E. Effect of Oxidation TemperaturE on Production of t~inbiastine (I)
f roa Ena~ine VIII3
TemF., ° CI ~ Yield of 2
vinblastine
-40 3..
-23 6.?
0 19.6
20 20.6
45
16.0
Reaction conditions: 2 eq. ferric chloride added, air bubbled
through the solution at 60 sl/~in for 15 mir..
By reverse-phase HPLC quantitation after reductive work-up with NaBP.4.
Enamine generated at -40° C (Procedure K (fi)).
12
1341262
Table 7. Effect of Dilution on Production of Vinblastine (I) frog Enamine
vIIT3
Dilution Factorl'4 1 Yield of 2
Vinblastine
1 19.6
25.2
30.1
29.6
50 24.7
1 iteactian conditions: 2 eq. ferric chloride added, air bubbled
through the solution at 60 slain for 15 :in. at 0° C.
2 By reverse-phase xPI~C quantitation after reductive work-up with NaBIi4.
Enamine Generated at -40° C (Procedure R (ii)).
~' Dilution Factor 1 ~ 100 ~g Isinium 'VI in 6 :1 sethanol to ~rhich
reducing agent Formula III (6 aq.) in 6 st2 ~ethaaol vas added.
(Total voluae ~ 1~ ~1) Dilution Factor 5 ~ Total voltme of b0 ml, etc.
13
13412fi2
Table 5 indicates results relating to yield of
vinblastine versus time of oxidation. The maximum yield of
vinblastine (I) is reached after 5 to 20 minutes of aeration
in the presence of 2 equivalents of ferric chloride.
Table 6 shows the results of various oxidation
temperatures, in the presence of two equivalents of ferric
chloride, on the yield of vinblastine (I). The temperature
range of 0°C to 20°C provides the highest yield of
vinblastine (I) after a reductive work-up with NaBH4, as
quantified by reverse-phase HPLC.
The effect of dilution of the enamine (VIII)
solution on the production of vinblastine (I), is indicated
in Table 7. A dilution factor of 10 to 20 on the enamine
(VIII) solution before aeration in the presence of ferric
chloride (2 equivalents) at 0°C, affords the best yield of
vinblastine (I) as quantified by reverse-phase HPLC after
reductive work-up with NaBH4.
In summary, the specific improvements in the
present application as they relate to the oxidative trans-
formation of the enamine (VIII), involve the dilution (5 to
50 folds) of the enamine (Formula VIII) solution, obtained
in the above reduction of iminium intermediate (Formula
VI), by the same solvent used in the reduction. For
practical purposes, the preferred dilution factor is usually
in the range of 8 to 12 folds. The above procedure is
conducted at a low temperature (0°C to -70°C), preferably
below -40°C and under cover with inert conditions such as
argon or an inert gas of Group Zero of the Periodic Table
(nitrogen, helium, neon, etc.). After this dilution
14
1341262
process, the oxidative transformation of the enamine
(Formula VIII) can be carried out as described above
(aeration at 60 ml/min for 15 min at 0°C in the presence of
ferric chloride (2 eq.)) to afford the corresponding iminium
intermediates (Formula XVI).
OOf l
N...
- _H_ -~.- t7-r
H ,.:
_~ I
vindof ine O OH
CH30 ~ ~O~ "CH3
CH3 R O
Formula RVI
The third area of improvement relates to the
reduction of iminium intermediates (Formula XVI) by alkali
metal borohydride (NaBH4, KBH4, LibH4, etc.) to vinblastine
(Formula I) and/or leurosidine (Formula XXXV). The iminium
intermediates (Formula XVI and Formula XVIa) produced from
the oxidative transformation of enamine (Formula VIII) are
reduced by the addition of alkali metal borohydride at low
temperature (4°C to -20°C), preferably at 0°C. The reduc-
tion is conducted at pH lower than 8.5 and preferably at 7.5
to 8. The total reaction mixture is then concentrated in
vacuo at low temperature (0°C to 10°C) before extraction and
isolation of alkaloid products.
1 347 2fi 2
H
I R N ~ ~ ~t oo~
O RN-
H'w~--~ I 1
_H_.-w ~"'~
I
off
CH30 N~O~ -CH3 O OH
CH3 R 0 CH3 0 ~~ Oy'CH3
R=COy~i3 CH3~ 0
R=C0yCH3
Formula XXXV (Leurosidine) Formula XVIa
For practical purposes, all the above improvements
can be incorporated, as indicated, directly into the overall
process conducted in a one-pot operation from the indole
unit (Formulas II, XXII or XXIII) and the dihydroindole unit
(Formula XXI, R - H) to the final products of Formula XXI.
Isolation of the various intermediates (Formulas XXIV, VI,
VIII and XVI) is omitted as summarized in Scheme 1.
In summary, the present method is applicable to
the production of dimer products from catharanthine and
dihydrocatharanthine with vindoline as starting materials
and phenyl, alkyl and amide derivatives embraced by the
following formulas:
16
<IMG>
1341262
Scneme 1. Optimum procedure for one-pot process - vinblastine from
catharanthine and vindoline.
D
r
ca~c~
XXtV
~C~3CO~Z O l 1
_60 o C !i 0 t OH ~ _Cti~
C,
yr
OKs
o (r
CONIiZ
l
t 3
c
t.Z-Reduction
f
VI11
Enamine Anhydrovinbls~tme
R~tso t.Z:1
18
1 34i 262
Scneu~_ : - cor,:inue~
h'
..
J
CH~O ~''~''~'!~~-Ct~
R ~ Gp2Ct~3
vllt
FeCI 2 a uiv.), 0° C
Air. ~~ mi9min. !5 min
Dilution Factor 10
00 ff
~ ooH
CiisO ~~ ~~~_C~~
XVI
XVIe
NvBii~ N~Bfix
_C+~i
X341262_
Formula XXI is as pictured and in that formula alk
represents a lower alkyl group of Cl-C6 and preferably Cl-
C3; aryl is mono-aryl such as benyzl, styryl, and xylyl; Rl
is a member of the group consisting of hydrogen, alk, CHO
and COR5 where R5 is alkyl or aryl; R2 and R3 are members of
the group consisting of hydrogen and -CO-alk; R4 is a member
of the group consisting of COO-alk, CONH-NH2, CONH2,
CONHR6, and CON(R6)2 where R6 is alkyl; Z is a member of
the group consisting of -CH2-CH2- and -CH-CH- and R is a
member of the indole family represented by Formula XXII
where R~ is a member of the group consisting of hydrogen, or
COO-alk; R8 is a member of the group consisting of hydrogen,
OH, O-alk, OCO-alk or alkyl; R9 is a member of the group
consisting of hydrogen, OH, O-alk, OCO-alk, or alk; R10 is a
member of the group consisting of hydrogen, OH, O-alk, OCO-
alk, or Formula XXIII where R11 is a member of the group
consisting of hydrogen or COO-alk; R12 is a member
consisting of alkyl.
SEecific Description of the Invention
This invention is specifically a process for the
production of compounds represented by the following
formulas:
s
XXI
n
s'
o-~z
iO n ~ i
1W'
13412fi2
xIi
x
XXIII
G
wherein in Formula XXI
Alkyl = CH3 or (CH2)nXH3 where n=1-5
Rl - CH3 or CHO
R2 - H or CO-alk
R3 - H
R4 - COO-alk or CONR13R14 where R13 and R14
can be any member of the group consisting
of hydrogen, alkyl, substituted alkyl,
aryl or substituted aryl functions
Z - -CH=CH- or -CH2-CH2-
R - Formula XXII or XXIII
R~ - or COO-alk
H
R8 - OH, O-alk, OCO-alk or alkyl
H,
R~ - OH, O-alk, OCO-alk or alkyl
H,
R1~ - OH, O-alk, OCO-alk
H,
R11 - or COO-alk
H
R12 - or alkyl
H
wherein the process a dimer derived
is for the synthesis
of
from an indole unit of the natural Iboga alkaloid family
containing an a dihydroindole
aza bicyclo-octane
portion and
21
13~~~62
unit of the natural Aspidosperma and Vinca alkaloid
families, the stereochemistry of the carbon-carbon linkage
between these two units being identical with that of
vinblastine which consists of
(a) forming an N-oxide intermediate in the cold at
a temperature of -70°C to +40°C from said indole unit by
oxidizing the bridge nitrogen and without isolating said
intermediate;
(b) treating said 2~-oxide indole intermediate in
the presence of one member of the group consisting of acetic
anhydride, halogenated acetic anhydride, and acetyl chloride
to effect a Polonovski-type fragmentation reaction;
(c) without isolating the product of step (b),
coupling said reaction product with a dihydroindole unit in
the presence of acetic anhydride, halogenated acetic
anhydride, and acetyl chloride at a low temperature of about
-70°C to +40°C under inert conditions;
(d) the product of step (c) is solated by solvent
evaporation preferably at low temperature in the range of -
20°C to 0°C;
(e) the product of step (d) is reduced by 1,4-
dihydropyridine compounds represented by Formula IX
H Rw
R R,
IX
~Z
Ri
where R3 and R5 in Formula IX are carboxylic esters (COO-
alk) and Rl, R2, R4 and R6 are members of the group
22
1 341 262
consisting of H, alkyl, aryl (Hantzch ester series) or tv-
substituted 1,4-dihydronicotinamides where R1 is a
substituted alkyl or substituted aryl function, for example,
benzyl, and R3 is CONR~RB where R~ and R8 is one member of
the group consisting of hydrogen, alkyl or aryl function;
(f) the product of step (e), an enamine, Formula
VIII, is isolated by solvent evaporation, preferably at low
temperature in the range of -20°C to 0°C;
(g) the product of step (e), a solution of enamine
VIII, is diluted (5 to 50 folds) by the same solvent
employed, preferably in the range of 8 to 12 folds at a low
temperature (0°C to -70°C), preferably below -40°C;
(h) the enamine obtained in step (f) or the
enamine solution obtained in step (g) is used to prepare
iminium intermediates, Formula XVI or XVIa, by a number of
oxidative processes including:
(1) controlled aeration/oxygenation in
which a solution of the enamine is stirred in open air or
with a stream of air/oxygen bubbled through the solution;
(2) as in (1) but with the addition of a
metal ion;
(3) as in (1) but with the addition of a
flavin coenzyme, as represented by Formula XII, to generate,
in situ, the corresponding 1,5-dihydroflavin coenzyme, as
represented by Formula XIII;
(4) as in (1) but with the addition of
hydrogen peroxide and/or hydroperoxides as represented by
the Formula R-OOH where R is alkyl or aryl
(i) the product, an iminium intermediate or
23
1341262
iminium intermediates obtained in step (h) is isolated by
sovlent evaporation, preferably at low temperature in the
range of -20°C to 0°C;
(j) the product obtained in step (i) or the
iminium intermediate solution obtained in step (h) is
converted to the target compounds of Formula XXI, for which
vinblastine and vincristine are examples and leurosidine
(Formula XXXV) by reduction with alkali metal borohydride
(NaBH4, KBH4, LiBH4) in suitable solvents (organic and/or
aqueous) as used in the oxidative processes of step (h).
Of special importance is the presence of electron
rich functional groups (e.g., caqrboxylic esters and
carboxylate salts) in R1 of Formula IX, capable of
coordination with the positively charged iminium
intermediate (Formula VI). Some examples of these 1,4-
dihydropyridine compounds are given in Table 1 (e. g.,
Formulas XXXI and XXXII). Such coordination increases both
the regioselectivity (1,4-reduction over 1,2-reduction) and
the rate of the reduction of iminium intermediate (Formula
VI), leading to an improved yield of the enamine (Formula
VIII).
The reduction is conducted under an inert
atmosphere at -60°C to +60°C but preferably in the
temperature range -20°C to -40°C. The solvents employed are
alcohols, acetonitrile or higher members of this series,
dimetheyl sulfoxide, dimethylformamide, various ethers such
as dioxane and tetrahydrofuran, and chlorinated
hydrocarbons.
The oxidative processes in step (h) (1)-(4) are
24
134122
conducted in organic solvents such as alcohols, acetonitrile
or higher members of this series, dimethyl sulfoxide,
dimethylformamide, various ethers such as dioxane,
tetrahydrofuran, aromatic hydrocarbons such as benzene,
toluene, etc. An aqueous buffer (e. g., phosphate, Tris-HC1,
MES buffers) at pH 5-9, but preferably in the range of 6-8,
can be used as co-solvent. The reaction temperature may
vary from -60°C to +60°C.
Of special importance is the presence of electron
rich functional groups (e.g., caqrboxylic esters and
carboxylate salts) in R1 of Formula IX, capable of
coordination with the positively charged iminium
intermediate (Formula VI). Some examples of these 1,4-
dihydropyridine compounds are given in Table 1 (e. g.,
Formulas XXXI and XXXII). Such coordination increases both
the regioselectivity (1,4-reduction over 1,2-reduction) and
the rate of the reduction of iminium intermediate (Formula
VI), leading to an improved yield of the enamine (Formula
VIII).
The reduction is conducted under an inert
atmosphere at -60°C to +60°C, but preferably in the
temperture range of -20°C to -40°C. The solvents employed
are alcohols, acetonitrile or higher members of this series,
dimethyl sulfoxide, dimethylformamide, various ethers such
as dioxane and tetrahydrofuran and chlorinated hydrocarbons.
The product of step (e) above, an enamine, Formula
VIII, is isolated by solvent evaporation, preferably at low
temperature in the range of -20°C to 0°C.
The process of this invention also prepares an
~3~~2sz
enamine and oxidizes said enamine to the iminium inter-
mediate (Formula XVI and XVIa) by a number of oxidative
processes including:
(a) controlled aeration/oxygenation in which a
solution of the enamine is stirred in open air with a stream
of air/oxygen bubbled through the solution;
(b) as in step (h)(1) but with the addition of a
ferric chloride;
( c ) as in step ( h ) ( 1 ) but with the addition of a
flavin coenzyme, as represented by Formula XII:
..OH
1~ ~1~~0
XII
w
0
to generate, in situ, the corresponding 1,5-dihydroflavin
coenzyme, as represented by Formula XIII
XIII
H
i
NN
f1
0
The oxidative processes are conducted in organic
solvents such as alcohols, acetonitrile or higher members of
26
1341262
this series, dimethyl sulfoxide, dimethylforman;ide, ethers
such as dioxane, tetrahydrofuran, aromatic hydrocarbons such
as benzene, toluene, etc.
EXAMPLES
The following examples (Examples 1 - 8) were
conducted at 20°C.
Example 1
Reduction of Iminium Intermediate (Formula VI) with 1-
Diphenylmethyl (-1,4-dihydronicotinamide (Formula IX, R1 -
Biphenyl methyl; R2 = R4 = R5 = R6 = H; R3 = CONH2) (Formula
XXVII) - Synthesis of Enamine (Formula VIII) - Procedure C)
To a stirred solution of iminium intermediate (VI,
100 mg) in degassed ethanol (6 ml) was added 1-
diphenylmethyl-1,4-dihydronicotinamide (Formula XXVII) (76
mg, 2.5 equivalents) in methanol (6 ml) under a positive
atmosphere of argon, the reducing agent being added
portionwise at the rate of 1 equivalent each 60 min. After
this, reverse phase HPLC analysis (Waters Radial-Pak C18 or
CtJ cartridge, methanol-H20-Et3N as solvent system)
indicated, among other products, formation of enamine (VIII)
and 3',4'-dehydrovinblastine (VII) in a ratio of 0.9:1 (60$
yield).
Example 2
Reduction of Iminium Intermediate (Formula VI) with 1-
Benzyl-3-cyano-1,4-dihydropyridine (Formula IX, Rl - benzyl;
R2 = R4 = R5 = R6 = H; R3 = CN) (Formula XXXIII) - Synthesis
of Enamine (Formula VIII) - (Procedure D).
27
1341262
To a stirred solution of iminium intermediate (VI,
100 mg) in degassed methanol (6 ml) was added 1-benzyl-3-
cyano-1,4-dihydropyridine (Formula XXXIII) (206 mg, 10
equivalents) in methanol (10 ml) under a positive atmosphere
of argon, the reducing agent being added portionwise at the
rate of 1 equivalent each 60 min. After this, reverse phase
HPLC analysis (as described above) indicated, among other
products, formation of enamine (VIII) and 3',4'-
dehydrovinblastine (VII) in a ratio of 1:1 (40~ yield).
Example 3
Reduction of Iminium Intermediate (Formula VI) with 1-
Benzyl-1,4-dihydronicotinyl-(2'-carbamoyl-pyrrolidinyl)-
amide (Formula IX, Rl = benzyl; R2 - R4 - R5 - R6 - H; R3 -
(2' carbamoyl-pyrrolidinyl) carbonyl) (Formula XXXIV) -
Synthesis of Enamine VIII) - (Procedure E).
To a stirred solution of iminium intermediate (VI,
100 mg) in degassed methanol (6 ml) was added 1-benzyl-1,4-
dihydronicotinyl-(2'-carbamoyl-pyrrolidinyl)-amide (Formula
XXXIV) (163 mg, 5 equivalents) in methanol (5 ml) under a
positive pressure of argon, the reducing agent being added
portionwise at the rate of 1 equivalent each 3U min. After
this, reverse-phase HPLC analysis (as described above)
indicated, among other products, formation of enamine (VIII)
and 3',4'-dehydrovinblastine (VII) in a ratio of 1.1:1 (60~
yield).
Example 4
Reduction of Iminium Intermediate (Formula VI) with 1,4-
Dihydro-1-(1-methoxycarbonyl isobutyl)-nicotinamide (Formula
28
1341262
IX, R1 - 1-methoxycarbonyl isobutyl; R~ - R4 - R5 - R6 - H;
R3 - CONH2) (Formula XXVIII) - Synthesis of Enamine (Formula
VIII) - (Procedure F)
To a stirred solution of iminium intermediate (VI,
100 mg) in degassed methanol (6 ml) was added 1,4-dihydro-1-
(1-methoxy carbonylisobutyl)-nicotinamide (Formula XXVIII)
(150 mg, 6 equivalents) in methanol (6 ml) under a positive
pressure of argon, the reducing agent being added
portionwise at the rate of 1 equivalent each 30 min. After
this, reverse-phase HPLC analysis (as described above)
indicated, among other products, formation of enamine (VIII)
and 3',4'-dehydrovinblastine (VII) in a ratio of 2:1 (65~
yield).
Example 5
Reduction of Iminium Intermediate (Formula VI) with 1-
(2',3',4',6'-Tetraacetyl-(beta)-D-glucopyranosidyl)-1,4-
dihydronicotinamide (Formula IX, R1 - (2',3',4',6'-Tetra-
acetyl-(beta)-D-glucopyranosidyl; R2 - R4 - R5 - R6 - H; R3
- CONH2) (Formula XXIX) - Synthesis of Enamine (Formula
VIII) - (Procedure G)
To a stirred solution of iminium intermediate (VI,
100 mg) in degassed methanol (6 ml) ws added 1-(2',3',4',6'-
tetra-acetyl-(beta)-D-glycopyranosidyl)-1,4-dihydronicotin-
amide (Formula XXIX) (238 mg, 5 equivalents) in methanol (10
ml) under a positive atmosphere of argon, the reducing agent
being added portionwise at the rate of 1 equivalent each 60
min. After this, reverse-phase HPLC analysis (as described
above) indicated, among other products, formation of enamine
(VIII) and 3',4'-dehydrovinblastine (VII) in a ratio of
29
1341262
1.5:1 (70~ yield).
Example 6
Reduction of Iminium Intermediate (Formula VI) with 1,4-Di-
hydro-1-(2'-methoxycarbonyl isopropyl)-nicotinamide (Formula
IX, Rl - 2' -methox;~ carbonylisopropyl; R2 - R4 - R5 - R6 -
H; R3 = CONH2) (Formula XXX) - Synthesis of Enamine (Formula
VIII) - (Procedure H)
To a stirred solution of iminium intermediate (VI,
100 mg) in degassed methanol (6 ml) was added 1,4-dihydro-1-
(2'-methoxy-carbonylisopropyl)-nicotinamide (Formula XXX)
(82 mg, 3.5 equivalents) in methanol (7 ml) under a positive
atmosphere of argon, the reducing agent being added
portionwise at the rate of 1 equivalent each 30 min. After
this, reverse-phase HPLC analysis (as described above)
indicated, among other products, formation of enamine (VIII)
and 3',4'-dehydrovinblastine (VII) in a ratio of l.l:l (65~
yield).
Example 7
Reduction of Iminium Intermediate (Formula VI) with 1,4-
Dihydro-1-(1',2'-dimethoxy carbonyl ethyl 1, R2 - R4 - R5 -
R6 - H; R3 - CONH2) (Formula XXXI) - Synthesis of Enamine
(Formula VIII) - Procedure I)
To a solution of iminium intermediate (VI, 100 mg)
in degassed methanol (6 ml) was added 1,4-dihydro-1-(1',2'-
dimethoxy carbonyl ethyl)-nicotinamide (Formula XXXI) (148
mg, 5 equivalents) in methanol (10 ml) under a positive
atmosphere of argon, the reducing agent being added
i 341 2s 2
portionwise at the rate cf 1 equivalent each 30 min. After
this, reverse-phase HPLC analysis (as described above)
indicated, among other products, formation of enamine (VIII)
and 3',4'-dehydrovinblastine (VII) in a ratio of 1.1:1 (70~
yield).
Example 8
Reduction of Iminium Intermediate (Formula VI) with 1,4-
Dihydro-1-(sodium-isobutyl-1-carboxylate)-nicotinamide
(Formula IV, Rl - sodium-isobutyl-1-carboxylate; R2 - R4 -
R5 - R6 - H; R3 - CONH2) (Formula XXXII) - Synthesis of
Enamine (Formula VIII) - (Procedure J)
To a solution of iminium intermediaqte (VI, 100
mg) in degassed methanol (6 ml) wqs added 1,4-dihydro-1-
(sodium-isobutyl-1-carboxylate)-nicotinamide (Formula XXXII)
(130 mg, 5 equivalents) in methanol (6 ml) under a positive
atmosphere of argon, the reducing agent being added
portionwise at the rate of 1 equivalent each 30 min. After
this, reverse-phase HPLC analysis (as described above)
indicated, among other products, formation of enamine (VIII)
and 3',4'-dehydrovinblastine (VII) in a ratio of 2.2:1 (70~
yield).
Example 9
Reduction of Iminium Intermediate (Formula VI) with 1,4-
Dihydro-1-(sodium-isobutyl-1-carboxylate)-nicotinamide
(Formula IX, R1 - sodium-isobutyl-1-carboxylate; R2 - R4 -
R5 - R6 - H; R3 - CONH2) (Formula XXXII) at low temperature
- Synthesis of Enamine (Formula VIII) (Procedure K)
(i) To a solution of iminium intermediate (VI, 100
31
13~12fit
mg) in degassed methanol (6 ml) at -20°C was added 1,4-
dihydro-1-(sodium-isobutyl-1-carboxylate)-nicotinamide
(Formula XXXII) (155 mg, 6 equivalents) in methanol (6 ml)
under a positive atmosphere of argon, the reducing agent
being added in one portion. After 45 min. at this
temperature reverse-phase HPLC analysis (as described
above) indicated, among other products, formation of enamine
(VIII) and 3',4'-dehydrovinblastine (VII) in a ratio of
3.2:1 (80$ yield).
(ii) Carrying out the reaction above at -40°C
gave, after 60 minutes, enamine (VIII) and 3',4'-
dehydrovinblastine (VII) in a ratio of 4.2:1 (85$ yield).
Example 10
Synthesis of Vinblastine (Formula I) by Oxidation of Enamine
(Formula VIII) to Iminium Intermediate (Formula XVI) with
Air in the Presence of Ferric Chloride at High Dilution.
(Method 5)
The solution containing the enamine (VIII),
(Procedure K (ii)) obtained from the iminium intermediate
(VI, 200 mg) was diluted five-fold with methanol before
oxidation (total vol.: 120 ml). Ferric chloride (75 mg, 2
equivalents) was then added, and air bubbled through the
solution, at 0°C, for 20 min. Sodium borohydride (200 mg)
was added, and the solution concentration in vacuo before
adding water (100 ml) and extracting with ethyl acetate (3 x
200 ml). The combined organic extract was dried over Na2S04
and the solvent evaporated in vacuo. The crude product was
purified by column chromatography (silica gel, TLC grade, 15
g). Elution with ether: chloroform (10:7) gave 3',4'-de-
32
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hydrovinblastine (VII, 18 mg, 11~). Further elution with
ether: chloroform: methanol (10:7:0.5) gave vinblastine (I,
62 mg, 37$).
Example 11
One-Pot Conversion of Catharanthine (Formula II) and
Vindoline (Formula III) to Vinblastine (Formula I) and
Leurosidine (Formula XXXV) - Overall Procedure
To a solution of catharanthine (II, 500 mg, 1.5
mmol) in dry dichloromethane (4.5 ml) at -15°C under a
positive atmosphere of argon was added m-chloroperbenzoic
acid (330 mg, 1.9 mmol) in one portion, and the mixture
stirred at -10 to -15°C for 5 minutes. After this time the
reaction mixture was cooled to -40°C and a solution of
vindoline (III, 450 mg, 1 mmol) in dry dichloromethane (1
ml) was added, followed immediately by trifluoroacetic
anhydride (1 ml, 7.1 mmol). After 2 h at -60°C volatiles
were removed in vacuo (high vacuum pump) and dry, degassed
methanol (12 ml) added after flushing the system with argon.
The resulting orange solution was cooled to -40°C and a
solution of 1,4-dihydro-1-(sodium-isobutyl-1-carboxylate)-
nicotinamide (Formula XXXII) (1.5 g, 6 mmol) in dry degassed
methanol (12 ml) was added under a positive atmosphere of
argon. After reduction was complete (by reverse-phase HPLC
monitoring), cold methanol (about 300 ml) was added, keeping
the temperature of the solutionn between -5 and 0°C. Ferric
chloride (330 mg, 2 mmol) was then added and dry air bubbled
through the solution at a rate of 60 ml/min for a period of
20 min. Sodium borohydride (1 g) was added and the solution
33
1 3~~ 2s 2
concentrated in vacuo (water aspirator) before adding water
(100 ml) and extracting with ethyl acetate (3 X 150 ml).
The combined organic extract was dried over Na2S04 and the
solvent evaporated in vacuo to give the crude product which
was purified by chromatography as previously described to
give 3',4'-dehydrovinblastine (VIII, 95 mg, 12~), vin-
blastine (I, 315 mg, 39~) and leurosidine (XXXV, 130 mg,
16$).
34
