������������������� �����
����� �����
���������� �������������� ������������������� ������� ����������������������������������������
���������� �������������������������������������������������������
��� ��!�"#$!�$#$
"!!���%���%&�'"! ��(�)�(����'��)*+�),�-�-��-,,-������&*(%+
����������������������� ������� ����������������������������������������������������������� ������������������������ ���!��"#��������$���%&���&%&����%&'%(�)���#��������)������)�*#����#�+�"#�����������,�����������������-����#+
��������
.�����������*+��&%&+�����/��������0����������1������������2�������"���������������������+�+����������� �������������� ������������������� �������� ������������������������������������3(%+�34���+�+�5.�6�738/7%/((�/38�&/8+
0�����������������2�������#�����#����������������������������#�����)�����������)��������������������#������9��������������������#���������������������������������+"#�����#�����#�����������������������)�,�9�����#��������������#�������,#����������������������)���#������������������������������������������#����+
:��������������������������������(/����������������2����,����������+�"#������������������� ,���� ������ ��������� )��� ���������� ��� �������� ���������� �/�#����������9����+�"#�����#�����)��)�������������������������������������� ������� ������������(/������4/��������2�����)�#���������#�������������������#����������������+"#�� ���������� ,���� ���������� �� �� %(���� �������� ������������ ���� ����/�#������������������#����+�"#��:;6��,��#��#�����)�������,�����������)����������))����������������������������,��#� �#��,����9,�<6:����)����:;6�+�"#�� �#��������������� �#���=���=���������)��#�����������������������������#������������������������)����������#���������������+
>������ ���,��#� �#�� �#����)� ������������� ����/����� ��� )�����������,�����������������#���������)��#������#��������,����������+�*���������������#������������������� ���� �� ������������������������ )��.���#��������/���������������������9����������������������������������������������+
���� ����1������������2�����������������������������
������� ��������!����� ��������"�������� ������# ������������ �!�"��$%&!������������� ����!��'(%$)*+��������!�������
?�*�����.�����������&%&
5..6�%4(%/4�%�5.�6�738/7%/((�/38�&/8��'�'��'��'����/%��74&� #���'@@��+9�+��@������A��B��'�'��'��'����/%��74&!
In Memory of my Mother, dedicatedto my Father.
List of Papers
This thesis is based on the following papers, which are referred to in the text
by their Roman numerals.
(I) Srivastava, P.; Engman, L.*
A radical cyclization route to cyclic imines
Tetrahedron Lett. 2010, 51, 1149-1151.
(II) Srivastava, P.; Barman, J.; Pathmasiri, W.; Plashkevych, O.; Wenska,
M.; Chattopadhyaya, J*
Five- and six-membered conformationally locked 2', 4'-carbocyclic ribo-
thymidines: synthesis, structure, and biochemical studies
J. Am. Chem. Soc. 2007, 129, 8362-8379.
(III) Srivastava, P.; Engman, L.*
Palladium catalyzed Sonogashira cross-coupling of organic tellurides with
alkynes
Manuscript
(IV) Appendix. Experimental procedures
Reprints were made with permission from the respective publishers.
Contents
1. Introduction ............................................................................................... 11 1.1 Radicals ............................................................................................... 11 1.2 Radical Reactions ............................................................................... 13 1.3 The chain concept ............................................................................... 14
1.3.1 Initiators ...................................................................................... 15 1.3.2 The tin hydride method ............................................................... 18 1.3.3 Silanes ......................................................................................... 20
1.4 Radical precursors ............................................................................... 21 1.4.1 Organochalcogens as radical precursors ..................................... 21
1.5 Radical cyclization .............................................................................. 22 1.5.1 5-Hexenyl cyclizations ................................................................ 23 1.5.2 6-Heptenyl cyclizations ............................................................... 24
1.6 Fundamentals of nucleic acids ............................................................ 26 1.7 Nucleic acid based therapeutics and significance of locked nucleosides ................................................................................................ 28
2. A radical cyclization route to �1 - pyrrolines (publication I) .................... 30 2.1 Design ................................................................................................. 30
2.1.1 Synthesis ..................................................................................... 31
3. A radical cyclization route to conformationally constrained nucleosides: Synthesis, structure and biochemical studies (publication II) ....................... 34
3.1 Radical reactions in nucleic acids ....................................................... 34 3.1.1 Synthesis of 2',4'-locked carbocyclic nucleoside (carbocyclic LNA-T; 37a/37b) ................................................................................. 35 3.1.2 NMR characterization of carbocyclic LNA-T (37a/37b) ............ 37 3.1.3 Synthesis of 2',4'-locked carbocyclic nucleoside (carbocyclic ENA-T; 48) .......................................................................................... 38 3.1.4 NMR characterization of the carbocyclic-ENA-T (48) ............... 39
3.2 Thermal denaturation studies of antisense oligonucleotides (AONs) . 40 3.3 Evaluation of target affinity and antisense properties of modified AONs ......................................................................................... 43 3.4 Probable mechanism for the nuclease stability of the modified AONs ......................................................................................... 46 3.5 RNase H digestion studies .................................................................. 47
4. Microwave-induced chalcogen extrusion reactions .................................. 49
5. Palladium (0)-catalyzed Sonogashira cross-coupling of organic tellurides with alkynes (Publication III) ........................................................ 51
5.1 Synthesis ............................................................................................. 53
6. Oxime ethers in 5-hexenyl radical cyclizations ......................................... 55 6.1 Attempted synthesis of a pyrrolidinone .............................................. 55
7. Piperidines from amino acids via radical cyclization ................................ 57 7.1 Attempted synthesis of piperidines ..................................................... 58
8. Summary in Swedish ................................................................................. 61
Acknowledgements ....................................................................................... 64
References ..................................................................................................... 67
Abbreviations
A Adenin-9-yl or adenosine Ac Acetyl AIBN 2,2' -Azobisisobutyronitrile AON Antisense oligonucleotide Aza-ENA 2'-N,4'-C-Ethylene bridged nucleic acid 9-BBN 9- Borabicyclo(3.3.1)nonane BNA Bridged nucleic acid Bn Benzyl BtH 1-H-Benzotriazole C Cytosin-1-yl or cytidine CD Circular dichroism COSY Correlation spectroscopy DCC Dicyclohexylcarbodiimide DCM Dichloromethane DEAD Diethyl azodicarboxylate DIPEA Diisopropylethylamine DMAP 4-Dimethylaminopyridine DMSO Dimethylsulfoxide DMTr 4,4'-Dimethoxytriphenylmethyl DNA Deoxyribonucleic acid ENA 2'-O,4'-C-Ethylene bridged nucleic acid FDA Food and Drug Administration G Guanin-9-yl or guanosine HMBC Heteronuclear multiple bond correlation LNA Locked nucleic acid mRNA Messenger RNA NOE Nuclear Overhauser effect PAGE Polyacrylamide gel electrophoresis Ph Phenyl RNA Ribonucleic acid RNaseH Ribonuclease H RNAi RNA interference SVPDE Snake venom phosphodiesterase T Thymin-1-yl or thymidine TBAF Tetrabutylammonium fluoride TBDMSCl tert-Butyldimethylsilyl chloride
THF Tetrahydrofuran TOCSY Total correlation spectroscopy TTMSS Tris(trimethylsilyl)silane
11
1. Introduction
1.1 Radicals Radicals are atoms or molecules which contain unpaired electrons.
Simple examples are hydrogen atoms and bromine atoms which have one
and seven electrons, respectively, in their valence shell. Almost all
radicals can be described as ‘‘free’’ as they exist independently, without
any need of support from the surroundings. Radicals are inherently
unstable and reactive since their major reactions i.e. dimerization,
hydrogen abstraction and disproportionation, are often favoured
thermodynamically and they occur with little or no activation barrier.
Notable exceptions are the naturally occuring oxygen (O2) and nitrogen
monoxide (NO). In 1849 Kolbe described the product obtained from
electrolysis of potassium acetate as ‘methyl radical’ with a formula C2H3.
It was later proven that the product was in fact ethane formed by
dimerization of two methyl radicals in the electrolysis reaction. The key
break-through, however, came in 1900 when Gomberg1 investigated the
reaction of triphenylmethyl bromide (1) with silver (Figure 1). In the
absence of oxygen the reaction produced a white solid which on
dissolution turned into a yellow solution. Gomberg proposed that the
product was hexaphenyl ethane (3) which could exist in equilibrium with
the coloured triphenylmethyl radical (2). It is another story that Gomberg
in fact had isolated another stable dimerization product 4. This was a
major discovery since it established that radicals are capable of
independent existence. Almost 30 years later, Paneth2,3 showed the
existance of the less stable, more reactive, methyl radical. It was not until
12
1937 that radicals were postulated to be intermediates in a variety of
organic reactions.4 Later works proved the involvement of radicals in
biological systems.5
Br + Ag + AgBr
Ph
PhPh
PhPhPh
Gomberg product
H
PhPhPh
Ph
Ph
Actual product
1 2
3 4
Figure 1. The classic Gomberg experiment
In the mid 1970s important contributions by physical chemists increased the
understanding of kinetics of various radical processes.6,7 With the knowledge
of rate constants for most of the common reactions, it was possible to design
experiments, i.e. select temperature and concentrations of radical precursors
and additives, that produced the desired product in synthetically useful yields.
This was the basis for the use of radical reactions in modern synthetic organic
chemistry.7-9
13
1.2 Radical Reactions Most reactions involving carbon or heteroatom centered radicals involve one
or several of the following steps:10 (Figure 2; A, B, C and D represent atoms
or groups, not necessarily representing carbon).
A + B A B
A B A + B
A + e- A-
A+A e-+
+A B C A B + C
A + B D A B D
A B D A + B D
(i)
(ii)
(iii)
(iv)
(v)
(vi) Figure 2. Common radical reactions.
(i) Homolysis: Thermal or photochemical cleavage of a covalent bond
between atoms or molecules results in formation of radicals.
(ii) Recombination: Radicals may combine by sharing their valence electrons
in a covalent bond in a process termed as recombination.
(iii) Electron transfer: Radicals may accept or donate an electron to form
anions and cations, respectively.
(iv) Atom or group transfer: Radicals may add to an atom accompnied by
expulsion of another radical from that atom (the SH2 type of reaction).
14
(v) Addition: A radical may add across a double bond to form a covalent
bond, e.g. Kharasch reaction.11
(vi) �-fission: Expulsion of a radical �� to a radical center accompanied by
double-bond formation. Such reactions are favoured, e.g. in the ring-opening
of cyclopropylcarbinyl or cyclobutylcarbinyl radicals where the ring strain is
relieved.12
In-spite of the diversity of radical reactions, the actual number of practical
protocols available for executing radical reactions are rather few in
number.13,14
1.3 The chain concept
Unlike cations or anions, radicals react with themselves by combination or
disproportionation at rates approching the diffusion controlled limit. The
maintainence of low concentrations of radicals over the course of reaction is
therefore imperative for a successful synthetic outcome. Chain reactions
involving reactions between radicals and non-radicals are ideally suited to
meet this requirement. Radicals generated via some initiation process
undergo propagation steps to generate new radicals leading ultimately to
termination of the chain via coupling or disproportionation reactions. The
chain reaction thus maintains a steady concentration of the reactive species
while converting the starting material to product. Most of the chain reactions
used in synthesis involve a metal hydride as a mediator. The processes are
illustrated in Figure 3.
15
Initiation
Initiator In
In +M H In H +M Propagation
M + R1
R2
X R1 M X+
R1
R2 +M H R2 H + M Termination
In , R1 , R2 , M non-radicals Figure 3. Processes involved in radical chain reactions.
The initiator undergoes homolytic fission thermally or photochemically to
generate the initiator radical In which abstracts hydrogen from the metal
hydride to give a metal radical M . Reaction of the radical precursor R1�X
with M results in transfer of X to generate an alkyl radical R1 . The alkyl
radical may undergo rearrangement (atom or group transfer, addition or �-
fission) to generate another radical R2 which could react with M-H with
hydrogen transfer thereby regenerating the metal radical M and thus
propagating the reaction. All radicals generated in the sequence may also
produce non-radicals via recombination thus terminating the chain.
1.3.1 Initiators For radical initiation one requires a molecule with weak covalent bonds that
can be broken homolytically under mild conditions. The energy required to
cleave the bond can be provided by heating (thermolysis), by ultraviolet light
(photolysis) or by X-rays (radiolysis). A majority of radical reactions useful
16
in synthetic organic chemistry use either photolysis or thermolysis as triggers
for radical initiation. The following compounds are frequently used:
Azo-compounds: Thermolysis of azo-compounds (R-N=N-R) leads to
cleavage of the two carbon–nitrogen bonds and formation of stable dinitrogen
(N2). The rate of decomposition is determined by the stability of the resulting
radical (R ). One of the favorite initiators with organic chemists is 2,2'-
azobisisobutyronitrile15 (AIBN; 5) which can be cleaved either photolytically
or thermally (Figure 4).
N CNNNC�
CNNC + + NC2N260%
CNNC
40%
t1/2(60oC)= 17 ht1/2(80oC)=1.3 h
5
Figure 4. Thermal decomposition of the initiator AIBN. Only 60 % of the initiator radical is available for initiation in a typical radical chain reaction.
The easy decomposition of AIBN is due to the stability of the 2-cyano-2-
propyl radical. It is stabilized both by resonance (the unpaired electron can be
delocalized to nitrogen) and through bond by electron donating methyl
groups.
NNNC
CN
MeO
OMe
NNNH
HN NH2
H2N
CNNN
NC
6 7 8
O
O
HO
OH
Figure 5. Azo-type radical initiators: (6) 2,2'-Azobis(4-methoxy-2,4-dimethylvaleronitrile); (7) 2,2'-Azobis(2-methylpropionamidine); (8) 4,4'-Azobis(4-cyanopentanoic acid).
17
Recently, azo-type radical initiators that decompose at room temperature
have been used for stereoselective carbon-carbon bond formation, e.g. 2,2'-
azobis(4-methoxy-2,4-dimethylvaleronitrile) (V-70).16(Figure 5; 6)
Hydrophilic radical initiators such as 2,2'-azobis(2-methylpropionamidine)
(Figure 5; 7) as dihydrochloride (V-50)17 and 4,4'-azobis(4-cyanopentanoic
acid) (V-501)18 (Figure 5; 8) have been used at moderate temperature for
initiating radical reactions in aqueous solution.
Peroxides and diacylperoxides: Thermolysis of peroxides has been used in
the study of radical reactions for a long time.11,19,20 On heating or irradiation,
alkoxy and acyloxy radicals, respectively, are formed by cleavage of the O�O
bond. Some commonly used peroxide initiators are shown in Figure 6.
Alkoxyl radicals obtained from di-tert-butyl peroxide (12) abstract hydrogen
atoms at the �- position in amines, ethers and esters.
O
OO
O
O
OO
OO
OO
OO C11H23 O
OO C11H23
O
9 10 11
12 13
Figure 6. Some commonly used peroxide radical initiators. Benzoyl peroxide (9), acetyl peroxide (10), tert-butyl perbenzoate (11), di-tert-butyl peroxide (12) and lauroyl peroxide (13).
Boranes: Trialkylborane21 together with molecular oxygen has been used
in several stereoselective reactions22 since they effectively initiate reactions at
-78 °C.
18
R3B + O2 R
R + O2
+ R3B R2BO2R + R
+ R2BOO
ROO
ROO Figure 7. Alkyl boranes as initiators.
As shown in Figure 7, alkyl radicals (R ) are thought to be the active
initiating species formed as a result of atom transfer. Other boranes such as 9-
BBN have also been used as radical initiators.23
1.3.2 The tin hydride method
To be useful in synthesis, a given chain reaction must generate radicals
site-selectively. A reaction extensively applied in synthesis which meets the
above requirement is the trialkyltin hydride mediated reduction of various
functional groups such as alcohols, halides and various chalcogen containing
R X + Bu3SnH R H + Bu3SnX
X= Xanthate, I, Br, Cl, SPh, SePh, TePh, NO2.
Figure 8. Reduction of halides and pseudohalides with trialkyltin hydride.
compounds (e.g. Barton-McCombie-deoxygenation,24 Ono-reduction25,26)
(Figure 8). Since its introduction by Kuivila27-29 the methodology has also
been extensively used for carbon-carbon bond formation.30-34Since this thesis
is concerned with radical cyclization chemistry, the various steps involved
are outlined in Scheme 1.
Radicals In produced by decomposition of the initiator readily abstract
hydrogen atoms from tributyltin hydride (14) to form the chain carrying
radical Bu3Sn (15). The propagation sequence involves an atom or group
abstraction (step 1) from 16 to provide a 5-hexenyl radical 17.
19
X
19
16 17
Bu3Sn��
X-SnBu3 Step 1
Bu3SnHIn�
In-H
14 15
+
17
Step 3
18
�kc
�
18
Step 4H-SnBu3kH Bu3Sn�
20
�
+
H
17
�
Step 2
Bu3Sn�H-SnBu3kH+
+ +
+
+
Bu3Sn�
Scheme 1. Steps involved in tin-mediated radical cyclization. Depending on concentrations and rates of hydrogen abstraction mixtures of cyclized (20) and reduced (19) products are formed.
Radical 17 may close via a first order 5-exo cyclization to provide
cyclopentylmethyl radical (18) (step 2). In competition with this process,
hydrogen abstraction could produce reduced compound 19 in a bimolecular
reaction (step 3). Finally, radical 18 abstracts a hydrogen from tin hydride to
produce cyclized compound 20 (step 4).
At 25ºC, kc, the rate of cyclization, is approximately 2 x 105 s-1 and kH, the
rate of hydrogen abstraction, is approximately 2 x 106 M-1s-1.35,36 The fate of
radical 17 is determined by the relative rate of cyclization (kc) versus
hydrogen atom abstraction (kH[Bu3SnH]). Thus, the concentration of tin
hydride is a variable by which product distribution can be controlled. At very
high concentrations, reduction product 19 will predominate (kc<kH[Bu3SnH].
At intermediate concentrations mixtures are expected. At low concentrations
20
(<0.05 M), most of radicals 17 will cyclize to 18 prior to hydrogen atom
abstraction (kc>kH[Bu3SnH] to give 20. Although the tin hydride method is
still a favourite among radical organic chemists,37 there is a need for
replacement,38 primarily because of toxicity and environmental concerns.
One such compound that has been in use for more than 20 years is
tris(trimethylsilyl)silane (TTMSS, (Me3Si)3SiH).39
1.3.3 Silanes A comparative study of the reaction of primary alkyl radicals with a
variety of group 14 hydrides revealed that rate constants for
tris(trimethylsilyl)silane and tributyltin hydride are comparable.40 It also
showed that for all kinds of radicals the reactivity of the former is always
lower. In Figure 9, the actual rate constants for hydrogen atom abstraction
from some group 14 hydrides are shown.
105 106 107 108
n-Bu3GeH
(Me3Si)2Si(H)Me (Me3Si)3SiH
Bu3SnH
(Me3Si)3SiH/RSH
(Me3Si)3SiH/ArSH
kH/M-1s-1 Figure 9. Rate constants for hydrogen atom abstraction from a variety of hydrogen donors by primary radicals at 80 ºC.41
Tris(trimethylsilyl)silane was introduced as an alternative to tin hydride in
the late 1980s.42 Toxicity of tin based reducing agents and the problems
associated with removal of tin residues has led to an increasing number of
reports on the use of TTMSS as a reducing agent in radical reactions. Thus,
iodides, bromides, chlorides, selenides, isocyanides, acid chlorides, xanthates
21
and sulphides have all been used as radical precursors in TTMSS mediated
chain reactions.41
1.4 Radical precursors Depending on the ease of preparation, halides, aryl sulphides, aryl selenides,
aryl tellurides, xanthates and other thiocarbonyls, nitro and thiohydroxamate
compounds were the precursors of choice for radical chain reactions.13 Alkyl
chlorides, however, cannot be used as substrates in reactions involving
addition of a radical to a double bond since chlorine atom abstraction by the
organostannyl radical is a slower process than is addition to the electron
deficient olefin. Hydrostannylation of the olefin and recovery of chloride
would result.43
1.4.1 Organochalcogens as radical precursors
Due to their stability and relative ease of preparation, organoselenium
compounds have proven to be very versatile radical precursors. They have
been used in Sn- and Si- mediated radical reactions as well as in group
transfer reactions for the formation of carbon-carbon bonds and carbon-
heteroatom bonds.44 Alkyl aryl selenides have been prepared for generation
of a wide range of carbon-centered radicals. Due to unfavourable expulsion
of phenyl radicals, alkyl phenyl selenides are known to cleanly produce alkyl
radicals. However, a word of caution should be mentioned here. Small
amounts of diphenyl diselenide, present as a result of photochemical
decomposition of the radical precursor or as a carry over from the radical
precursor synthesis, can be detrimental. The explanation to this is the well
established phenomenon of polarity reversal catalysis45,46 wherein the
reduction of the alkyl radical is brought about by benzeneselenol (PhSeH
22
reacts with alkyl radicals faster than Bu3SnH by more than three orders of
magnitude). In fact, the selenol acts as a catalyst for the reduction processes.
The rapid reduction has been taken advantage of in dehalogenation,
deoxygenation and desulphurization reactions47,48 where undesired radical
rearrangements which are sufficiently fast to proceed in the presence of tin
hydride can be avoided.
Phenyl selenides are easily obtained from nucleophilic49 or electrophilic50
selenium reagents.51 Thus, halides, alcohols, lactones, epoxides and aziridines
have been transformed into phenyl selenides and carried like a protecting
group for a radical through several synthetic steps. In fact, in one of the first
tin-mediated addition reactions, phenylselenides were found to give better
yields than the corrosponding iodides.52
The use of organotellurium compounds as precursors for carbon-centered
radicals was first reported by Clive in 1980.53 Ten years later, Barton and
Ramesh54 first applied organotellurium compounds for radical mediated
carbon-carbon bond formation using a thiohydroxamate ester as initiator and
promotor. This work was followed by the first report on organotellanyl group
transfer reactions with alkynes55 and intramolecular group transfer reactions
of acyl tellurides.56 The scope was further extended by intramolecular group
transfer reactions in the synthesis of cyclopentanes and tetrahydrofurans.57-60
The synthetic utility61 reflects the fact that the organochalcogen group
transfer reactions occur as rapidly as those for iodides.62-64
1.5 Radical cyclization It has been known since long that free radicals add to double bonds.11 The
alkyl radical so formed may undergo further reactions such as hydrogen or
halogen abstraction. The peroxide-catalyzed addition of HBr to olefins is an
early example of this type of reaction. Intramolecular addition of radicals to
alkenes and alkynes always proceed more rapidly than the corresponding
23
intermolecular reactions. Radical cyclization reactions have therefore been
extensively used for preparation of both carbocycles and heterocycles.
1.5.1 5-Hexenyl cyclizations
The kinetically controlled rearrangement of variously substituted 5-hexenyl
radicals to cyclopentylmethyl radicals has been extensively
investigated,10,19,20,65,66 both from a mechanistic and synthetic point of view
by Beckwith,10,67-69 Giese70, Curran,32,33 Stork,71 and Rajanbabu.72-75 It
constitutes an excellent method for preparation of 5-membered carbocycles
and heterocycles.31
�
��
5-exo product 6-endo product
k6-endok5-exo
Figure 10. Rearrangement of a 5-hexenyl radical to a cyclopentylmethyl radical (5-exo) or a cyclohexyl radical (6-endo).
In light of Baldwin’s rules76,77 for ring closure, the preference for 5-
exo- over 6-endo- cyclization (Figure 10) is well understood in terms of a
relatively strain-free chair-like transition state78 which fulfills the
stereoelectronic requirements for radical addition to a double bond. For the
unsubstituted 5-hexenyl radical k5-exo is approximately 2 x 105 s-1 and k6-endo is
4 x 103 s-1 thus resulting in formation of cyclopentylmethyl radical as the
major product. The Beckwith-Schiesser, Houk79 model also provides
satisfactory explanations for the observed stereoselectivities in the cyclization
of 2-, 3- and 4-monosubstituted 5-hexenyl radicals43 (Figure 11). This can be
explained assuming the substituent adopts a pseudo-equatorial position in the
24
transition state,74 i.e. , the 2- and 4-methyl-5-hexenyl radicals cyclize with
preferential formation of trans-disubstituted cyclopentanes whereas 1- and 3-
methyl-5-hexenyl radicals predominantly afford cis-disubstituted products.
The efficiency and regiochemistry of intramolecular cyclization addition
reactions12 have been shown to be controlled by (i) the initial radical
structure, (ii) the steric effects resulting from the olefin substitution pattern,10
(iii) the geometric constraints on the chain linking the radical centre and the
tethered double bond and (iv) the presence of a heteroatom in the chain.80-83
major minor
major
major
major
minor
minor
minor
2-substituted 5-hexenyl radical
3-substituted 5-hexenyl radical
4-substituted 5-hexenyl radical
1-substituted 5-hexenyl radical
Figure 11. Diastereoselectivities in the cyclization of 1-, 2-, 3- and 4-substituted 5-hexenyl radicals.43
1.5.2 6-Heptenyl cyclizations
Whereas 5-hexenyl radical cyclization comprises the ultimate tool for making
five membered rings, the use of 6-heptenyl cyclization for making 6-
membered rings is less important. The formation of 6-membered rings by
radical cyclization presents several problems (Figure 12). First, the rate of
cyclization of the parent 6-heptenyl radical is almost two orders of magnitude
slower than that of the 5-hexenyl radical. Second, ring-closure is
25
considerably less regioselective (kexo/kendo~ 7, for the 6-heptenyl radical10,78 vs
kexo/kendo ~ 50, for the 5-hexenyl radical).12,63 Third, the 6-heptenyl radical is
prone to rearrangement via [1,5]-hydrogen atom transfer to give an allylic
radical.12
�
�
�
H � H�
k6-exo
k7-endo
[1,5]-shif t
6-exo product
7-endo product
rearranged product
Figure 12. Rearrangement of a 6-heptenyl radical to a cyclohexylmethyl radical (6-exo) or a cycloheptyl radical (7-endo). Competing [1,5]-hydrogen atom transfer.
Cyclization of methyl substituted 6-heptenyl radicals was recently
studied84 and the poor selectivity was attributed to the conformational
flexibility inherent in the radical. Difficulties associated with cyclization of
6-heptenyl radicals have been overcome by several methods: (i) Restriction
of the degrees of freedom in the heptenyl chain either by incorporation of
rings10 or substituents (Thorpe-Ingold effect85) (ii) Activation of the double
bond. The presence of an electron-withdrawing group at the terminal vinyl
position of the 6-heptenyl radical significantly improved both the rate of
cyclization and the regioselectivity in the ring-closure. Highly regioselective
6-exo cyclizations with moderate to good stereoselectivity proceeding via a
chair-like transition state have been reported.86-88 Our attempts to use 6-
heptenyl cyclizations in nucleoside synthesis and for the preparation of
substituted piperidines will be discussed in chapters 3 and 7.
26
1.6 Fundamentals of nucleic acids The nucleic acids,89,90 i.e. deoxyribonucleic acid (DNA) and ribonucleic acid
(RNA) are the carriers of genetic information in all living organisms. DNA
acts as the storehouse of genetic information in the nucleus while the RNA
acts as the carrier of the genetic information to the ribosome, where it is
processed and translated into proteins (notable exceptions are the retro
viruses where the RNA acts as the carrier of the genetic information). The
nucleic acids are made-up of nucleotide building blocks. Each nucleotide
consists of a nucleobase, a sugar and a phosphate linker. The nucleobase
generally consist of one of the following: 9-adeninyl (A), 9-guaninyl (G), 1-
thyminyl (T) or 1-cytosinyl (C) in DNA while in RNA, 1-thyminyl is
replaced by 1-uracilyl (U). Adenine and guanine are purines, while thymine,
uracil and cytosine are pyrimidines. The major difference between DNA and
RNA is the absence of the 2'-hydroxyl in DNA (2'-�-D-deoxyribofuranosyl)
which is present in RNA (2'-�-D-ribofuranosyl). This subtle change in the
sugar gives rise to a distinctly different structure as well as function of the
two molecules. Each sugar residue is linked to the neighbouring sugar
residues by a 3',5'-phosphodiester bond (3'-carbon atom of a sugar with the
5'-carbon of the neighbouring sugar). While the sugar moiety and the sugar-
phosphate backbone of DNA/RNA primarily play a structural role, the
genetic information is determined by the sequence of the nucleobases. In the
cell, the RNA usually exists in single strands, while the DNA adopts a double
helical91 structure in which the two DNA strands are held together by
noncovalent interactions i.e. intra-strand stacking of nucleobase moieties and
weak hydrogen bonds between intra-strand pairs of nucleobases. The
hydrogen bonding occurs between laterally opposite bases, ‘The base pair’,
of the two strands of the DNA duplex according to Watson-Crick base-
pairing rules91 (A specifically hydrogen bonds to T and G specifically to C).
27
Normally, G�C-pairs (linked by three hydrogen bonds) are stronger than
A=T-pairs (linked by two hydrogen bonds). The double stranded structures of
DNA and RNA form different types of helices with the two strands oriented
in opposite directions (3'�5' of one strand and 5'�3' of the other). The
DNA:DNA duplexes normally form a B-type helix which is characterized by
an extended structure with a distance of ~3.4 Å between adjacent bases and
approximately 10 base-pairs per helical turn, which are nearly perpendicular
to the helical axis. Also, the B-type helix has a narrow and deep minor
groove (~6 Å) and a wide major groove (~12 Å). In contrast, the RNA:RNA
duplexes normally adopt an A-type helix which has a more compressed
structure (the distance between adjacent bases is ~2.6 Å). Thus, A-type
duplexes exhibit approximately 11 base-pairs per helical turn, which are
tilted approximately 20° with respect to the helical axis. Also, unlike the B-
type helix, they have a very wide (~11 Å) and shallow minor groove and a
very narrow (~3 Å) and deep major groove. The DNA:RNA duplexes
typically adopt an intermediate A/B-type structure, with an overall structure
resembling the A-type. It is also noteworthy that A-type duplexes normally
have higher duplex stability as compared to the B-type ones. With identical
nearest neighbours the RNA:RNA is most stable while the order of stability
between DNA:DNA and RNA:DNA depends on the sequence context.92,93 In
general, the order of stability is RNA:RNA> DNA:RNA> DNA:DNA. The
helical geometry described above can also be explained by the furanose
conformation94-96 adopted in the nucleosides and the �/�-angle of the
phosphate backbone.97 The furanose ring of a nucleoside can adopt numerous
conformations in solution due to puckering98 of the sugar ring. In solution,
the major population of 2'-deoxy pentofuranose adopts a 2'-endo (DNA-type)
conformation while in RNA the sugar adopts a 3'-endo conformation (RNA-
type) also known as South- and North- type respectively.
28
1.7 Nucleic acid based therapeutics and significance of locked nucleosides Targeting proteins with small molecules, which is the basis of the traditional
drug discovery approach is difficult. Moreover, it has been estimated that
only 10-14 % of the proteins have appropriate binding sites (druggable) for
small molecules.99 Therefore, instead of targeting the protein itself, if one can
target DNA or messenger RNA (mRNA) by an oligonucleotide
complementary to the target sequence, it might be possible to stop the protein
production.100 Thus, nucleic acid based therapy offers a seemingly simple and
straight-forward mechanism to down-regulate a gene of interest. The
technique used to treat a disease at the genomic level is called anti-gene
approach.101,102 In this method an artificial, short, modified DNA or RNA
sequence is synthesized which forms a stable triple helix (triplex)103,104 with
the double stranded genomic DNA through the major groove via Hoogsteen
hydrogen bonding.105,106 The other popular oligonucleotide based therapy is
called antisense technology.107 The traditional antisense technology exploits
the ability of a single stranded oligodeoxynucleotide (ODN) of 15-25
nucleotides to bind to the accessible regions of the target mRNA via Watson-
Crick base pairing in a sequence specific manner, thereby inhibiting protein
synthesis either by sterically blocking the ribosomal assembly or by RNase H
mediated degradation of the mRNA.108
The antisense technology has found increased acceptance from the
pharmacological perspective which is evident from the recent approval by
FDA of the first oligonucleotide based antisense drug called Vitravene®.
Other oligonucleotide-based therapeutics are in the pipeline.109
Although the natural phosphodiester oligonucleotides are easy to synthesize,
their use is limited as they are degraded by intracellular endo-and
exonucleases. This has warranted chemical modification of the
oligonucleotides in order to utilize them for therapeutic applications. Recent
29
years have seen development of conformationally constrained bicyclic110,111
and tricyclic112,113 nucleotides, in which the sugar is locked in a definite
puckered conformation. Such oligonucleotides show promising properties
with respect to target RNA binding and nuclease resistance. Among several
molecules reported, short nucleotides containing locked nucleic acid
(LNA)114 or bicyclo-nucleic acid (BNA111) modifications have shown
unprecedented thermal stability (+3 to +8 °C per modification depending
upon the sequence context). The enhanced target binding property of the
conformationally constrained bicyclic sugar units in these nucleotides has
been attributed to the improved stacking between the nearest neighbours and
quenching of concerted local backbone motions by LNA nucleotides in
ssLNA so as to reduce the entropic penalty in the free energy of stabilization
for the duplex formation with RNA.115 The unique features of LNA/BNA
have led to the synthesis of a number of closely related analogues, in which
the 2',4'-bridge has been altered.116-119 Studies120 with several sugar modified
nucleotides show that substituents in the modified sugar play an important
role in conformational steering, hydration, hydrophobic/hydrophilic
interactions, electrostatic interactions121 and thus influencing the interaction
of modified oligonucleotides with other nucleotides and/or enzymes present
in the system.122
30
2. A radical cyclization route to �1 - pyrrolines (publication I)
Pyrrolines are important structural motifs in many pharmacologically
important alkaloids and several synthetic methodologies were used for their
preparation.123-129 As mentioned in the introduction (section 1.5), free radical
methodology has been used for synthesis of 5- and 6-membered nitrogen
containing heterocycles.59,130 However radical methodologies for the
synthesis of pyrrolines are few in number and based primarily on generation
of iminyl radicals.131-134
2.1 Design
As presented in Scheme 2, readily available �-phenylselenenyl ketones
could, after imine formation with an appropriate allylic amine, serve as
precursors of carbon-centered radicals which would hopefully cyclize to �1-
pyrrolines under reductive conditions. A variety of methods are available for
introduction of PhSe-groups into the �-position of ketones and aldehydes.
We found the two-step procedure involving reaction of methyl ketones with
PhSeCl3, followed by reduction of the resulting Se,Se-dichloride, convenient
for large scale preparation.135-137 Imine formation was tried using allyl amine
together with TiCl4 as a dehydrating agent. The reactions went smoothly to
completion and we were able to isolate crude phenylselenenyl imines in good
yields.
31
R
OSePh
R
NSePh
R
N
R
N
R
N
+R
NSePh
N
R
Scheme 2. Radical cyclization route to cyclic imines.
Syn/anti-isomerization in imines has been well studied.138,139 For N-alkyl
imines the energy barrier to interconversion is low. We envisaged (Scheme 2)
that under radical cyclization conditions, resonance in the radical
intermediate would allow for conversion of the unreactive (in cyclization) Z-
isomer into the reactive E-isomer. Our attempts to bring about cyclization by
slow addition of tin hydride led mostly to reduced product. Better results
were obtained using tris(trimethylsilyl)silane as a hydrogen donor.140
2.1.1 Synthesis In the representative reaction sequence shown in Scheme 3, acetophenone
(21) was reacted with phenylseleninyl trichloride in dry ether overnight.
O OSePh
NSePh
O
+
N N
SePh+
(i), (ii) (iii)
(iv)
95% 80%
15% 46%
21 22 23
21 24 25
Scheme 3. Reagents and conditions: (i) PhSeCl3, dry ether, r.t., overnight; (ii) Na2S2O5, water, ether; (iii) Allylamine, TiCl4, dry ether, -78°C to r.t. overnight; (iv) AIBN, TTMSS, C6H6, reflux, 8 h.
32
The white precipitate of crystalline Se(IV)-dichloride formed was filtered off
and isolated in quantitative yield.
Reduction with aqueous Na2S2O5 provided selenide 22 in 95 % yield after
two steps. Reaction with allylamine and TiCl4 afforded the crude imine 23 in
80 % yield as a mixture of syn/anti-isomers. Reductive radical cyclization
using TTMSS/AIBN in refluxing benzene afforded cyclic imine 24 in 46 %
yield along with 15 % of reduced product 21, and a trace of group transfer
product 25. Application of this methodology to other methyl ketones afforded
substituted imines in moderate yields (Table 1).
Table 1. Cyclic imines prepared.
Product
Yielda
Crudeb (%)/isolated (%) N 52/47
N
MeO
50 /46
N
S
33/-
N 49/38 cis/trans=82/18
N 18/-
aYield over two steps (imine formation and cyclization). bAs determined by 1H NMR using DMAP as an internal standard.
Low molecular weight pyrrolines such as those prepared from pinacolone and
2-acetylthiophene were quite difficult to isolate. Unfortunately, our efforts to
33
extend the methodology to cyclic �-phenylselenenyl ketones and �-
phenylselenenyl aldehydes failed at the imine-forming step.
34
3. A radical cyclization route to conformationally constrained nucleosides: Synthesis, structure and biochemical studies (publication II)
3.1 Radical reactions in nucleic acids
As early as 1964, Johnson141 reported formation of C-5' to C-7 linked
adenosine upon anaerobic photolysis of vitamin B12 coenzyme. From that
time and on nucleic acids142 and sugars72-75,143-153 have been frequently used
as substrates in radical mediated transformations. Apart from its synthetic
utility, the radical chemistry of sugars and nucleosides have also contributed
to the understanding of several biological phenomenon such as charge
transfer154 and radical induced damage in DNA and RNA.155,156 The
conversion of ribonucleotides to deoxyribonucleotides catalyzed by
ribonucleotide reductase is one of the most fascinating radical reactions in
nature.157,158
Recent developments in the synthesis of conformationally constrained
nucleoside analogues led us to try free radical carbon-carbon bond formation
to constrain sugar pucker in a definite conformation. We were successful in
synthesizing two conformationally constrained carbocyclic analogues of
thymidine by free radical intramolecular carbon-carbon bond formation. Our
results are presented below.
35
3.1.1 Synthesis of 2',4'-locked carbocyclic nucleoside (carbocyclic LNA-T; 37a/37b)
The synthesis starts from a known sugar precursor 26114 which was
selectively benzylated to give 27 using a known procedure.117 The remaining
primary alcohol in sugar 27 was oxidized to the corresponding aldehyde 28
by Swern oxidation.159 A vinyl group at C-4' was then introduced by Wittig
methodology160 using crude aldehyde to give the sugar 29 (87 % in two steps
from 27). Anti-Markovnikov addition of water to olefin 29 was effected by
successive hydroboration – oxidation using 9-BBN/NaOH-H2O2 to give
O
OO
HO
OBnHO
O
OO
BnO
OBnHO
O
OO
BnO
OBnO
(i) (ii) (iii)O
OO
BnO
OBn
O
OAcOAc
BnO
OBn
O
OAc
BnO
OBn
NH
N
O
OO
OH
BnO
OBn
NH
N
O
OO
OCSOPh
BnO
OBn
NH
N
O
O
(vi)
26 27 28 29
O
OO
BnO
OBnHO
O
OO
BnO
OBn
OHO
OH
NH
N
O
O
CH3
ODMTrO
OH
NH
N
O
O
CH3
ODMTrO
O
NH
N
O
O
CH3PNO CN
(iv) (v)
(vii) (viii) (ix)
(x) (xi) (xii) (xiii)
30
31 32 34 35
36a: major (70 %) 7'R36b: minor (30 %) 7'S 37a/37b 38a/38b
66% 87% 95% 70%
80% 72%
73% 76% 74% 80%
39a/39b
3'
5'
7'6'
4' 2' 1'OBnO
OBn
NH
N
O
O
CH3
3'
5'
7'6'
4' 2' 1'
33
Scheme 4. Reagents and conditions: (i) NaH, BnBr, DMF, �5 °C to r.t. overnight; (ii) oxalyl chloride, DMSO,�78 °C, DIPEA, DCM; (iii) PPh3
+CH3Br�,1.6 M butyllithium in hexane, THF, �78 °C to r.t. overnight,;(iv) 9-BBN, THF, 3 N NaOH, 33 % H2O2; (v) oxalyl chloride, DMSO, �78 °C, DIPEA, DCM, PPh3
+CH3Br�, 1.6 M butyllithium in hexane, THF, �78 °C to r.t. overnight; (vi) acetic acid, acetic anhydride, triflic acid; (vii) persilylated thymine, TMSOTf, CH3CN, 0°C to r.t. overnight; (viii) 27 % methanolic NH3, overnight; (ix) DMAP, O-phenylchlorothiocarbonate, pyridine; (x) Bu3SnH, toluene, AIBN, 4 h; (xi) 20 % Pd(OH)2/C, ammonium formate, methanol, reflux, 8 h; (xii) DMTr-Cl, pyridine, overnight; (xiii) 2-cyanoethyl N,N-diisopropylphosphoramidochloridite, DIPEA, dry THF, overnight.
36
alcohol 30 in 95 % yield, which was again subjected to Swern oxidation /
Wittig 160 olefination to give the required sugar 31 (70 % in two steps from
30) with a strategically placed propenyl side chain at C-4' as an accepter for
radical cyclization. Compound 31 was subjected to acetolysis using a mixture
of acetic anhydride, acetic acid and triflic acid to give the corresponding
diacetate 32, quantitatively as �/� anomeric mixture (single spot on TLC and
proven by 1H-NMR). The crude diacetate 32, after bicarbonate workup, was
subjected to a modified Vorbruggen reaction161 involving in situ silylation of
thymine and subsequent trimethylsilyl triflate mediated coupling to give
thymine nucleoside 33 in 80 % yield in two steps from 31. The �-
configuration of product 33 was confirmed by a 1D differential NOE
experiment, which showed 3 % enhancement of H-2', and 1 % enhancement
of H-3' upon irradiation of H-6 (dH6-2' ~ 2.3 Šfor the �� anomer).
Deacetylation of compound 33 using 27 % methanolic ammonia overnight,
and subsequent esterification using O-phenylchlorothiocarbonate yielded the
desired precursor 35 (72 % in two steps from 33) for radical cyclization. The
key free radical cyclization reaction was carried out using Bu3SnH with
AIBN initiation at 115 °C in degassed (N2) toluene. To ensure that the radical
generated has adequate lifetime to capture the double bond before quenching,
the concentrations of Bu3SnH and AIBN were kept low by high dilution and
slow, drop-wise addition. 5-Exo cyclization occurred exclusively to give the
expected,10 5-membered, 2',4'-cis-fused carbocyclic product with a bicyclo
[2.2.1] heptane skeleton as an inseparable diastereomeric mixture of
compound 36a (major compound 70 %, 7'R) and 36b (minor compound 30
%, 7'S). The formation of the bicyclic nucleosides 36a and 36b, with a North
fused conformationally constrained pentofuranosyl moiety was confirmed by
long range 1H-13C NMR correlation (HMBC)162 and 1H-1H (TOCSY)163 for
both isomers. A 1D differential NOE experiment established that the
exocyclic methyl at C-7' is in close proximity to H-1' in the major isomer (d7'-
37
Me/H-1' ~2.8 Å for R and 4.4 Å for S configured C-7') of the bicyclic structure.
The benzyl groups in the bicyclic nucleosides 36a/36b were removed using
Pd(OH)2/C and ammonium formate in methanol to give the corresponding
dihydroxy compounds 37a and 37b respectively. Several attempts to separate
this diastereomeric mixture of 37a/37b failed. We therefore subjected it
directly to 5'-dimethoxytritylation (74 %) to give 38a/38b followed by 3'-
phosphitylation (80 %) to give the phosphoramidite 39a/39b using standard
conditions.119
3.1.2 NMR characterization of carbocyclic LNA-T (37a/37b)
The 1H NMR spectrum at 600 MHz of the ring-closure products 36a/36b
revealed a diastereomeric mixture of sugar-fused 5-membered bicyclic 3', 5'-
di-O-benzyl protected nucleosides (Scheme 4). However, because of the
overlap of the H-7' and H-2' peaks, firm NMR evidence could not be
obtained for the carbon-carbon ring closure between C-2' and C-7'. The H-7'
and H-2' peaks in the deprotected compounds 37a/37b were however fully
resolved, and hence could be used for full NMR characterization.
The 1H spectrum showed the presence of two diastereomers, a major 37a
as well as a minor isomer 37b in ca. 7:3 ratio. The upfield H-2' at � 2.43 ppm
along with H-7' at � 2.65 ppm and their proton-proton couplings, proven by
detailed double decoupling and by COSY experiments, showed that the C-2'
substituent is the C-7' methine-carbon. The NOE enhancement (~12 %,
corresponds to ca. 2.6 Å) between H-6 (thymine) and H-3' of 37a/37b, in
addition to 3JH1',H2' = 0 Hz, further confirmed that the sugar is indeed locked
by the fused carbocycle in the North conformation as observed for other
North-locked nucleosides such as ENA164, LNA114, and aza-ENA117. This
further showed that the 1-thyminyl moiety is in the �-configuration with an
anti-conformation across the glycoside bond. The fact that the NOE
38
enhancement of 6.5 % for H-1' upon irradiation on CH3 at C-7' of 37a was
found, showed that the methyl group on C-7' is in close proximity of H-1' (ca.
2.8 Å), thereby confirming the R configuration for C-7'. The NOE
enhancement of 4.5 % for H-7' in 37b upon irradiation on H-1', on the other
hand, confirmed that the H-7' is in close proximity of H-1' (ca. 2.2 Å) and
hence the S configuration was assigned for C-7'. HMBC correlations between
H-7' and C-2' for compounds 37a/37b unequivocally proved that the oxa-
bicyclo[2.2.1]heptane ring system was formed in the ring closure reaction.
3.1.3 Synthesis of 2',4'-locked carbocyclic nucleoside (carbocyclic ENA-T; 48)
For synthesis of a carbocyclic-ENA-T analog, a 6-exo heptenyl type
radical cyclization was desired, and thus the C-4' allylated sugar 31 described
in Scheme 4 was subjected to another sequence of hydroboration-oxidation,
Swern oxidation159 and Wittig160 reactions described previously (yields of 40
and 42 were 90 % and 72 %, respectively). The sugar 42 was subjected to
acetolysis followed by a modified Vorbruggen-type161 coupling to give 44
with a �-configured thymine in 70 % yield over two steps. Deacetylation
followed by esterification using O-phenylchlorothiocarbonate yielded the
ester 46 (60 % yield after two steps). Purified ester was then subjected to free
radical cyclization. Although 7-endo cyclization usually occurs only 6-7
times more slowly than 6-exo cyclization, we were able to isolate exo-product
47 in a decent 76 % yield.
39
O
OO
BnO
OBn
O
OO
BnO
OBnHO
O
OO
BnO
OBnO
O
OO
BnO
OBn
O
OAcOAc
BnO
OBn
O
OAc
BnO
OBn
NH
N
O
OO
OH
BnO
OBn
NH
N
O
OO
OCSOPh
BnO
OBn
NH
N
O
O
OHO
NH
N
O
O
OH CH3
ODMTrO
NH
N
O
O
OH CH3
ODMTrO
NH
N
O
O
O CH3P
N
ONC
47: (8'S)
31 40 41 42 43
44 45 46
48 49 50
(i) (ii) (iii) (iv)
(v) (vi) (vii) (viii) (ix)
(x) (xi)
90% 72%
70% 60% 76% 82%
80% 84%1'2'3'4'
5'
6'7' 8'
OBnO
NH
N
O
O
OBn CH3
1'2'3'4'
5'
6'7' 8'
Scheme 5. Reagents and conditions: (i) 9-BBN, THF, overnight, 3 N NaOH, 33 % H2O2; (ii) oxalyl chloride, DMSO, �78°C, DIPEA, CH2Cl2; (iii) PPh3
+CH3Br�, THF, 1.6 M butyllithium in hexane, �78 °C to r.t. overnight; (iv) acetic acid, acetic anhydride, triflic acid; (v) persilylated thymine, TMSOTf, CH3CN, 0 °C to r.t. overnight; (vi) 27 % methanolic NH3, overnight; (vii) DMAP, O-phenylchlorothiocarbonate, pyridine, r.t. overnight; (viii) Bu3SnH, toluene, AIBN, 4 h, reflux; (ix) 20 % Pd(OH), ammonium formate, methanol, reflux,12 h; (x) DMTr-Cl, pyridine, r.t. overnight; (xi) 2-cynoethyl N,N-diisopropylphosphoramidochloridite, DIPEA, dry THF.
The bicyclic nucleoside 47 was deprotected using Pd(OH)2/C and
ammonium formate in methanol to give the corresponding diol 48 in 82 %
yield. Dimethoxytritylation (80 %) followed by phosphitylation using
standard conditions119 gave the fully protected phosphoramidite 50 in 84 %
yield.
3.1.4 NMR characterization of the carbocyclic-ENA-T (48)
For compound 48, the upfield H-2' shift at � 2.26 ppm along with H-8' at �
2.20 ppm and their vicinal proton-proton couplings, proven by the double
decoupling as well as COSY experiments, shows that the C-2' substituent is
the C-8' methine-carbon. Strong NOE enhancement (8.6 %, corresponding to
40
ca. 2.6 Å) between H-6 (thymine) and H-3' in compound 48, in addition to 3JH1',H2' = 0 Hz, further confirms that the sugar is indeed locked in the North-
conformation and that the 1-thyminyl moiety is in the �-configuration with an
anti conformation across the glycoside bond. The NOE enhancement of 3.0
% for H-1' upon irradiation at CH3(C-8') proved that the CH3(C-8') group is
in close proximity of H-1', hence the C-8' chiral center is R-configured. The
vicinal coupling of H-2' with H-8' as evidenced by double decoupling
experiments and COSY spectra also unequivocally showed that the bicyclo
[3.2.1] octane ring system had indeed been formed in the ring-closure
reaction (Scheme 5). This conclusion was further corroborated by the
observation of a long range 1H-13C connectivity of H-8' with C-2', C-3', and
C-1', that of H-7' with C-2' and that of H-2' with CH3(C-8'), C-7', and C-8' in
an HMBC experiment.
3.2 Thermal denaturation studies of antisense oligonucleotides (AONs) The phosphoramidites 39a/39b and 50 were incorporated by mono-
substitution into a 15-mer DNA sequence through automated synthesis on an
Applied Biosystems 392 RNA/DNA synthesizer for further studies. The
stepwise coupling yields of the modified phosphoramidite were 96 % and 98
%, respectively. Dicyanoimidazole was used as the activating agent for
39a/39b, whereas tetrazole was used to activate 50 with 10 min coupling
time for modified phosphoramidites, followed by deprotection of all base-
labile protecting groups with 33 % aqueous ammonia at 55 °C to give AONs
1-17 (Table 2).
41
Table 2. Thermal denaturation of native and modified AONs in the duplexes with complementary RNA or DNA targets.a
AON Sequence Tm/°C with RNA
b�Tm Tm/°C with DNA
c�Tm
1. 3'-d(CTTCTTTTTTACTTC)-5' 44 45
2. 3'-d(CTT(LNA)CTTTTTTACTTC)-5' 48 +4 47 +2
3 3'-d(CTTCT T(LNA)TTTTACTTC)-5' 49 +5 46.5 +1.5
4 3'-d(CTTCTTTT(LNA)TTACTTC)-5' 49 +5 45.0 0.0
5 3'-d(CTTCTTTTTT(LNA)ACTTC)-5' 49 +5 46 +1
6 3'-d(CTT(5-carbo)CTTTTTTACTTC)-5' 47.5 +3.5 45 0.00
7 3'-d(CTTCTT(5-carbo)TTTTACTTC)-5' 49 +5 44 -1.00
8 3'-d(CTTCTTTT(5-carbo)TTACTTC)-5' 48 +4 44 -1.00
9 3'-d(CTTCTTTTTT(5-carbo)ACTTC)-5' 47.5 +3.5 43.0 -2.00
10 3'-d(CTT(6-carbo)CTTTTTTACTTC)-5' 45.5 +1.5 43.5 -1.5
11 3'-d(CTTCTT(6-carbo)TTTTACTTC)-5' 45.5 +1.5 39.5 -5.5
12 3'-d(CTTCTTTT(6-carbo)TTACTTC)-5' 45.5 +1.5 40.0 -5.0
13 3'-d(CTTCTTTTTT(6-carbo)ACTTC)-5' 45.5 +1.5 39.5 -5.5
14 3'-d(CTT(aza-ENA)CTTTTTTACTTC)-5' 48 +4 44.5 -0.5
15 3'-d(CTTCTT(aza-ENA)TTTTACTTC)-5' 46.5 +2.5 42.5 -2.5
16 3'-d(CTTCTTTT(aza-ENA)TTACTTC)-5' 47.5 +3.5 42 -3
17 3'-d(CTTCTTTTTT(aza-ENA)ACTTC)-5' 48 +4 42 -3
42
a Tm values measured as the maximum of the first derivative of the melting curve (A260 vs. temperature) and are average of at least three runs recorded in medium salt buffer (60 mM Tris-HCl at pH 7.5, 60 mM KCl, 0.8 mM MgCl2 and 2 mM DTT) with temperature range 20 to 70 oC using 1μM concentrations of the two complementary strands; b�Tm = Tm relative to RNA compliment; c�Tm = Tm relative to DNA compliment.
The sequence is targeted to the coding region of the SV40 large T-antigen
(TAg)165,166 and has been used in the study of antisense activity of (N)-
Methanocarba-T substituted oligonucleotide167 and as well as in the study of
antisense and nuclease stability assays of oxetane, 118 azetidine,119 and aza-
ENA117 modified oligonucleotides. Thermal denaturation studies of AONs
(Table 2) containing carbocyclic-LNA-T or carbocyclic-ENA-T showed an
increase in Tm of 3.5 to 5.0 °C/modification. (AONs 6-9 in Table 2) and 1.5
°C/modification (AONs 10-13 in Table 2), respectively, for the AONs
duplexes with complementary RNA. However, a net decrease of up to 5.5
°C/modification has been observed for the AON duplexes with
complementary DNA. The DNA/DNA duplex is known to have a shallow
minor groove and the presence of a water backbone all along the
oligonucleotide further contributes to the stability of the duplex90. A decrease
in the thermodynamic stability of a modified AON/DNA duplex may be due
to an increase in steric crowding in the shallow minor groove. Since the
DNA/RNA heteroduplex is known to have a wider but shallow minor groove,
it is evident that the exocyclic methyl is well tolerated in the modified
AON/RNA heteroduplex. The 6-membered carbocyclic-ENA-T counterpart
did not show any sequence dependent change in Tm (+1.5 °C/modification).
A higher decrease in the stability of the 6-membered carbocyclic-ENA-T
modified AONs with DNA further suggests that an additional hydrophobic
methylene linker introduces more steric crowding in the minor groove of the
duplex in addition to disrupting the extensive backbone water network.
However, since 5-membered carbocyclic-LNA-T was used as a
43
diastereomeric mixture of R- and S- isomers oriented differently in the minor
groove, we might be observing averaged steric effects. Further studies with
pure diastereomers are warranted in addition to X-ray crystallographic
evidence of hydration disruption by the exocylic methyls oriented differently
in the minor groove space.
3.3 Evaluation of target affinity and antisense properties of modified AONs
The stability of AONs towards nucleases is necessary in order to develop
any therapeutic oligonucleotides (antisense,108 RNAi,168, microRNA169,170 or
triplexing agents171). In order to study the nuclease stability, if any, of the
carbocyclic-LNA (37a/37b) and carbocyclic-ENA (48) modified AONs a
single modification was introduced at the 3' end and were digested in snake
venom phosphodiesterase (SVPDE) and human blood serum which mainly
comprises 3'-exonucleases. In order to study the effect of a hydrophobic 2'
modification a comparative nuclease stability assay was done with LNA114
and aza-ENA117 modified AONs as shown in the PAGE autoradiograms
(insets A, B, C and D in Figures 13 and 14).
44
Figure 13. Autoradiograms of 20 % denaturing PAGE showing degradation patterns of 5'-32P-labeled AONs in human blood serum (see Table 2 for AON sequences). Inset A: AON 1 and LNA-modified AONs. Inset B: Carbocyclic LNA-modified AONs, Inset C: Carbocyclic–ENA modified AONs, and Inset D: aza-ENA-modified AONs. Time points are 0 h, 0.5 h 1 h, 2 h, 5 h, 7 h, 9 h and 12 h after incubation with blood serum.
Investigation by using circular dichroism (CD) did not reveal any change
in the global structure of the hetero or homoduplex. Since only singly
modified oligonucleotides were used in the CD studies, we assumed that
there is little effect of the structural modification on the structure of the 15-
mer sequence and that there must be minor structural changes adjacent to the
modification which would be reflected in the enzyme digestion studies.
45
Figure 14. Autoradiograms of 20 % denaturing PAGE showing degradation pattern of 5�-32P-labeled AONs in SVPDE (see Table 2 for AON sequence). Inset A: AON 1 and LNA-modified AONs. Inset B: Carbocyclic-LNA modified AONs. Inset C: Carbocyclic-ENA modified AONs. Inset D: aza-ENA modified AONs. Time points are 0 h, 1 h, 2 h, 24 h, 48 h and 72 h after incubation with the enzyme.
The comparison of the modified AON stability in blood serum and
SVPDE, owing to various modifications, becomes very clear as we compare
the digestions of AONs 2, 6, 10 and 14 with T(LNA), T(5-carbo), T(5-carbo) and
T(aza-ENA) each having a specific single modification at the position 3 from the
3'-end: The site of 3'-exonuclease promoted hydrolysis was dictated by the
site of the incorporation of the modification in the AON (Figures 13 and 14).
The isosequential parent LNA modified AONs were found to be rapidly
digested within 30 min similarly to the native heteroduplex. On the other
hand, the 3'-terminal nucleotide is hydrolyzed by 3'-exonuclease (SVPDE
and human serum) in AON substituted by the 5- or 6-membered carbocyclic
residue (AONs 6 and 10) at position 3 to give only the n-1 fragment.
46
3.4 Probable mechanism for the nuclease stability of the modified AONs
The nuclease digestion studies with modified AONs show that a single
modification of AON with carbocyclic-LNA-T or with carbocyclic-ENA-T
nucleotides at position 3 from the 3'-end in AONs 6 and 10 has provided
unprecedented nuclease stability. It has been suggested that replacement of
substituents involved in natural enzyme-substrate complex results in poor
recognition and processing by the nucleolytic enzymes, thereby resulting in
the nuclease stability.172 It was also shown that the native ribonucleoside with
a 2'-O-alkyl substituent either by its bulk173 or by its stereoelectronic
modulation120,122,173,174of hydration can bring about nucleolytic stability. Egli
et al.175 have demonstrated that charge effects and hydration properties are
important factors in influencing the nuclease stability of AONs with a normal
phosphodiester backbone.
AONs containing 5-membered carbocyclic-LNA and 6-membered
carbocyclic-ENA show enhanced, but similar, blood serum stability, thereby
indicating that the steric bulk is relatively unimportant.173 This was in sharp
contrast to the conclusion drawn by comparison of the LNA versus ENA176
modified AONs, in that the latter was found to be 2.5-3 times more stable
than the former,164 wherein the extra methylene linker was suggested to be
the structural cause of observed stability.
Since the carbocyclic AONs were completely devoid of polar effect at C-
2',177 the explanations invoking charge or steric effects is not applicable to
explain the nuclease stability of the carbocyclic AONs in the blood serum.
The enhanced stabilities of the carbocyclic-AONs with respect to their
bicyclic 2'-O-(LNA and ENA) and aza-ENA analogs suggests that either the
accessibility of water or the active site of the enzyme to the 2'-O- in LNA or
to the 2'-N- in aza-ENA substituent in a modified nucleotide is probably more
important in order to cleave the vicinal 3'-phosphodiester bond by the
47
exonucleases of the blood serum. A calculation of the solvation energy for
various 2'-modified nucleosides suggests that the 5-membered and 6-
membered carbocyclic nucleosides, on account of their hydrophobic nature,
are not as well solvated as compared to their LNA, ENA and aza-ENA
counterparts, thereby showing that hydration around a scissile phosphate is
probably important for the nuclease promoted hydrolysis in carbocyclic
modified AONs devoid of charge effects.
3.5 RNase H digestion studies
In the antisense strategy, RNase H recruitment by the modified AON, with
long nuclease stability as exhibited by the carbocyclic modifications, and the
target RNA heteroduplexes is an important step in the design of potential
therapeutics for engineering specific gene silencing effects.108
Hence, RNase H mediated cleavage of the RNA strand complementary to
the modified AON strands (with carbocyclic-LNA, carbocyclic-ENA, aza-
ENA and LNA modifications) was studied, using the native AON/RNA
duplex as a reference, in order to address the issue if the recruitment of
RNase H by the heteroduplexes is in any way compromised as a result of
carbocyclic modifications incorporated in the AON strand. We used E. coli
RNase H in the studies because of its availability and the fact that its
properties are known to be similar to the mammalian enzyme.178 The RNA
complementary to AONs formed duplexes and were found to be good
substrates for RNase H but with varying cleavage sites (Figure 15),
depending on the site of modification in the AONs.
48
Figure 15. The RNase H1 cleavage pattern of AON/RNA heteroduplexes. Vertical arrows show the RNase H cleavage sites, with the relative length of the arrow showing the extent of the cleavage. The square boxes around a specific sequence shows the stretch of the RNA, which is resistant to RNase H cleavage.
49
4. Microwave-induced chalcogen extrusion reactions
The carbon-chalcogen bond is weakened as we traverse the chalcogen
group of the periodic table from top to bottom. Extrusion reactions resulting
in carbon-carbon bond formation are feasible but many of them require harsh
conditions such as flash vacuum pyrolysis used to induce formation of
benzocyclobutenes from dihydrobenzotellurophenes179 or pyrolysis to effect
deselenation180 and desulphurization181 during cyclophane synthesis.
Sometimes extrusion had to be assisted in order for carbon-carbon bonds to
form. For example, degassed Raney nickel was found to produce biaryls from
diaryl tellurides.182 We were curious to see if microwave-heating would
induce such chalcogen extrusion reactions. Indeed, we observed that certain
6- or 7-membered cyclic chalcogenoanhydrides when heated with
microwaves afforded diketones as a result of chalcogen extrusion
accompanied by carbon-carbon bond formation (Table 3). Thus, when
selenodiphenic anhydride (51) was heated in a sealed vial in chlorobenzene at
250 ºC for 2 h, phenanthroquinone (52) was quantitatively formed. Seleno-
and telluroanhydride starting materials were easily prepared from acid
chlorides and Na2Se and Na2Te generated in situ.183-185 Tellurodiphenic
anhydride decomposed to give phenanthroquinone already during attempted
recrystallization. Telluroglutaric anhydride (53) afforded 1,2-
cyclopentanedione (54) in 46 % yield when heated at 210 ºC in the
microwave cavity. However, we were unable to produce
benzocyclobutanedione from seleno- or tellurophthalic anhydride at 250 ºC
under similar reaction conditions. Our results are summarized in Table 3.
50
Table 3. Microwave-assisted chalcogen extrusion reactions.
Chalcogenohydride Reaction conditions Product Yield
Se
O
O 51
Microwaves, chlorobenzene,
250 ºC, 2 h O
O
52
100 %
M
O
O
M=Se, Te
Microwaves, chlorobenzene,
250 ºC, 4 h
No reaction
SeO O
Microwaves, chlorobenzene,
250 ºC, 4 h
No reaction
Te OO 53
Microwaves, chlorobenzene,
210 ºC, 2 h OH
O
54
46 %
Attempts to extend the chalcogen extrusion methodology to the preparation
of biaryls and alkylated aromatics was met with little success. We therefore
turned the attention to metal catalyzed coupling reactions.
51
5. Palladium (0)-catalyzed Sonogashira cross-coupling of organic tellurides with alkynes (Publication III)
The rare element tellurium has a surprisingly rich organic chemistry. A
variety of organotelluriums with well-defined stereochemistry are available
and they are often stable enough to be taken through several synthetic
transformations. In addition to their application as radical precursors (section
1.4.1), organotelluriums have been extensively used in other carbon-carbon
bond forming reactions.186-189 Synthetically useful carbodetelluration was
demonstrated in the carbonylation of aryltellurium chlorides with Ni(CO)4.190
Carbodetelluration was also shown to occur when aryltellurium (IV)-
chlorides were reacted with olefins in the presence of a stoichiometric
amount of palladium.191 In the absence of olefins, biaryls were obtained in
good yields. In the presence of carbon monoxide, carbonylation occured to
yield aromatic acids in good yields using Pd(II) in stoichiometric amount.192
In 1988 Barton193 reported homocoupling of diaryl and alkyl aryl tellurides in
the presence of a stoichiometric amount of Pd(OAc)2. Diaryl ditellurides
which are the precursors to alkyl aryl and diaryl tellurides were also shown to
undergo homocoupling. Until now, most of the palladium assisted
carbodetelluration reactions occured with stoichiometric amounts of rather
expensive palladium reagents. It was not until 1996 that Uemura reported
catalysis of palladium in cross-coupling reactions.194,195 Under Fujiwara-Heck
conditions in the presence of a suitable oxidant, aryl tellurides were coupled
with alkenes to give aryl alkenes in good yields. Whereas diaryl tellurides did
not form any homocoupling products, alkenyl tellurides did react in the
52
presence of a catalytic amount of palladium to provide 1,3-dienes. The cross-
coupling of organic tellurides using catalytic palladium has been extensively
used in Stille,196 Heck,197 Negishi,198 Suzuki,199 and Sonogashira type
reactions of vinylic,197,200,201 and heteroatomic202,203 tellurides. Recently
Pd(0)-catalyzed coupling stimulated by ultrasound was successful for biaryl
synthesis using butyl aryl tellurides as starting materials.204 In our hands this
reaction did not proceed as described. However, from the previous literature
it is obvious that the success of these coupling reactions depends rather
critically on the choice of palladium catalyst, base, solvent and additives. We
therefore decided to find out more about the suitability of diaryl and alkyl
aryl tellurides as coupling partners in transition metal-catalyzed reactions.
Te
(i) +55
56 570%44%
Scheme 6. Reagents and conditions: (i) Pd(OAc)2 (2.0 eq.), Et3N (2.0 eq.), CH3CN, reflux overnight.
When butyl 2-naphthyl telluride (55) was allowed to react (Scheme 6) with a
2-fold molar excess of Pd(OAc)2, 2,2'-binaphthyl (56) was isolated in low (44
%) yield and none of the homocoupling product 57 was formed. This is in
contrast to the early Barton193 paper which claims formation of such products.
When diaryl tellurides were tried under these conditions, biaryls were formed
in similarly low yields (e.g. bis(4-methoxyphenyl) telluride gave 4, 4'-
dimethoxybiphenyl in 28 % yield). Catalytic reactions carried out in the
presence of an oxidant (CuCl2) did not produce any of the desired coupling
products. Rather, the organotellurium compounds were oxidised to the
corresponding tellurium (IV)-species which could be reduced to regenerate
the starting materials. We therefore turned to Sonogashira type cross-
coupling reactions205 which are known to proceed in a catalytic fashion in the
absence of an oxidant.
53
5.1 Synthesis Alkyl aryl and diaryl telluride starting materials were prepared by standard
methods206-208 from the corresponding diaryl ditellurides. Thus, diphenyl
ditelluride (58) 209 was reduced with NaBH4 with careful exclusion of air and
n-butyl bromide was added to produce butyl phenyl telluride (59) in 97 %
yield. In another experiment, diphenyl ditelluride (58) was refluxed with Cu
powder in benzene for 12 h to produce diphenyl telluride (61) in 95 % yield.
TeTe
Tei ii
iii
Te iv
59
61
95 %
68 %
36 %
97 %
58 60
60 Scheme 7. Reagents and conditions: (i) NaBH4, n-butyl bromide, EtOH; (ii) 10 mole % Pd(PPh3)4, 1.5 equivalents of phenylacetylene and CuI, NEt3, reflux, 8 h; (iii) C6H6, Cu powder, reflux, 12 h; (iv) As in (ii) but 2.0 equivalents each of phenylacetylene and CuI were used.
For Sonogashira-type reactions it was found that heating at reflux with 10
mole-% of Pd(PPh3)4, CuI (1.5-2.0 eq) and phenylacetylene (1.5-2.0 eq) with
NEt3 as solvent produced fair to good yields of diarylacetylene coupling
products. Ideally, diaryl tellurides would give rise to two equivalents of
coupling products for every equivalent of telluride used. However, judging
from the low yields of diphenylacetylenes (Table 4) isolated when diphenyl
telluride (61) and other diaryl tellurides were used as coupling partners, only
one of the aryls seems to be transferable. The fate of the remaining aryl group
is as yet unclear. n-Butyl phenyl telluride (59) produced diphenyl acetylene
(60) in 68 % yield as shown in Table 4. Other n-butyl aryl tellurides also
furnished coupling products in fair yields (Table 4). It would therefore seem
more atom economic to use alkyl aryl tellurides as coupling partners rather
than diaryl tellurides. It remains to be seen if the butyl group that we have
54
used so far is optional or if a smaller alkyl such as methyl or ethyl would be
an even better choice. Table 4. Sonogashira cross-coupling of organic tellurides.
Telluride Alkyne Product Yielda
Te
68 %b
Te
36 %c
Te
67 %
Te
35 %c
Te
H3CO H3CO
78 %
Te
OCH3H3CO
H3CO
40 %c
Te
Me2N NMe2
Me2N
19 %c
a Isolated yield; bYield determined by 1H NMR using an internal standard; cYield calculated on the basis that 1.0 mmol of reactant would give 2.0 mmol of product. Two equivalents each of phenylacetylene and CuI were used.
55
6. Oxime ethers in 5-hexenyl radical cyclizations
5-Hexenyl radical cyclization in oxime ethers has been used for
construction of lactones,210 carbocyclic amine derivatives211 and
pyrrolidinones (Figure 16).210 The approach to construct pyrrolidinones from
amino acid precursors has, to the best of our knowledge, not been tried using
radical cyclization chemistry.
N
NOBn
O N
NHOBn
O
EtEt3B (8 eq.)toluene, ref lux
OHOH
47%
Figure 16. Tandem radical addition-cyclization to oxime ethers for pyrrolidinone synthesis.
6.1 Attempted synthesis of a pyrrolidinone
Our approach, exemplified with the commercially available phenylalaninol
(62), is shown in Scheme 8. TBDMS protection of the hydroxyl group in
pyridine afforded 63 in 98 % yield. Coupling with the O-benzyl oxime of
glyoxylic acid212 with dicyclohexylcarbodiimide afforded amide 64 with an
appropriately positioned double bond for 5-exo radical cyclization (70 %
yield). Deprotection with tetrabutylammonium fluoride gave alcohol 65 in 96
% yield. Attempted thiocarbamoylation of the alcohol with
thiocarbonyldiimidazole was unsuccessful in our hands. However, treatment
56
with O-phenylchlorothiocarbonate afforded the desired radical precursor 66
in moderate yield (62 %).
NH2
HO
NH2
OTBDMS
NH
NTBDMSO
O
OBn
NH
NHO
O
OBn
NH
NO
O
OBnSPhO
NH
O
NHOBn
i ii
iii iv v
O
HO NOBn
63 64
65 66
62
98% 70%
96% 62%
Scheme 8. Reagents and conditions: (i) TBDMSCl, DMAP, pyridine, overnight, (ii) DCC, DCM, overnight, (iii) TBAF, THF, 4 h, (iv) O-phenylchlorothiocarbonate, DMAP, pyridine, overnight, (v) TTMSS, AIBN, toluene, reflux. Radical cyclization was tried with tris(trimethylsilyl)silane and AIBN
initiation in refluxing toluene. The reaction product turned out to be a
complex mixture from which it was not possible to identify any cyclized
material.
57
7. Piperidines from amino acids via radical cyclization
The piperidine ring is an important structural motif in natural products and
synthetic pharmaceuticals and hence there is a need for development of
stereocontrolled methodologies for piperidine synthesis.213 The problem of
diastereocontrol in radical reactions has received considerable attention.214
Several excellent reviews215-220 are available on the topic including the radical
cyclization approch.87,221-228
Taking clue from previous work229,230 in our group (Scheme 9) with 2-
substituted 3-aza-5-hexenyl radicals, wherein it was demonstrated that bulky
N-substituents direct cyclization to occur preferentially with formation of cis-
2,4-disubstituted pyrrolidines, we set out to investigate the effect of the N-
substituent in cyclizations of 2-substituted 3-aza-6-heptenyl radicals.
N
PhSe
R
P=H
P= Ph2PO
(i)
(i)
NR
PNR
P
NR
PNR
P
P
trans:cis= 3.3:1
trans:cis= 1:10
Scheme 9. Control of diastereoselectivity by the N-substituent during radical cyclization. Reagents and conditions (i) n-Bu3SnH, AIBN, C6H6, h� (15 °C)
58
7.1 Attempted synthesis of piperidines The initial plan for radical precursor preparation involved homoallylation of
an appropriate amine with 4-bromo-1-butene in analogy with previous
work.229,230 However, we found alkylation difficult, probably due to the low
reactivity of homoallyl bromide and the presence of a bulky �-substituent in
the amino acid derived amines. We were able to circumvent the problem as
shown in Scheme 10.
HN
R O
OMe
Bt
NH
R
HO
NH
R
O OMe
N
RNH
R
PhSe
NR
PhSe
R
67a, R= Ph
i ii
iv
iii
v
67b, R= Bn68a, R= Ph, (57 %)68b, R= Bn, (62 %)
69a, R= Ph, (85 %)69b, R= Bn, (61 %)
70a, R= Ph, (80 %)70b, R= Bn, (85 %)
71a, R= Ph, (73 %)71b, R= Bn, (70 %)
72a, R= Ph, (97 %)72b, R= Bn, (0 %)
Scheme 10. Reagents and conditions: (i) Allyltrimethylsilane, BF3-etherate, THF, (ii) NaBH4, EtOH, reflux, (iii) PPh3, DEAD, DCM, (iv) PhSeSePh, NaBH4, TFA, EtOH, (v) Diphenylphosphinoyl chloride, DMAP, NEt3. Bt= Benzotriazolyl. Starting materials were previously prepared amino acid esters 67a and
67b,231,232 where the amine had been alkylated with
formaldehyde/benzotriazole (BtH), and an allyl group installed by treatment
with allyltrimethylsilane/BF3-etherate giving 68a and 68b. Reduction with
NaBH4 or LiAlH4 afforded the corresponding alcohols 69a and 69b which
under Mitsunobu conditions (DEAD and PPh3) provided aziridines 70a and
70b in 80 % and 85 % yields, respectively. Ring-opening with
59
benzeneselenol gave the radical precursors 71a and 71b in 73 % and 70 %
yields, respectively.
Radical cyclization of these compounds with TTMSS and AIBN-initiation
gave complex mixtures of reduced and cyclized products.
Introduction of a bulky diphenylphosphinoyl group before cyclization was
also attempted. However, whereas N-phosphinoylation of phenylalanine
derived radical precursor 71b failed under standard conditions
(diphenylphosphinoyl chloride, DMAP, Et3N), the corresponding
phenylglycine derivative 71a was readily phosphinoylated to give 72a (97 %
yield).
TTMSS-mediated cyclization of 72a resulted in a complex mixture of
products. Cyclization was also tried with Bu3SnH/AIBN. TFA treatment to
remove the diphenylphosphinoyl group, followed by basic workup, yielded
3-butenyl 2-phenylethylamine (73), as the only product (92 %). This material
has probably formed via a neophyl rearrangement233 (Scheme 11).
N
PhSei, ii,iii
72a 73R
N N N
NH
Bu3SnH
R R R Scheme 11. Formation of amine 73 via a neophyl rearrangement. Reagents and conditions: (i) Bu3SnH, AIBN, toluene, reflux, (ii) TFA, reflux, (iii) NaOH (aq). Obviously, 6-heptenyl cyclization onto an unactivated double bond does not
60
occur rapidly enough for the reaction to be synthetically useful for piperidine
synthesis. Instead, other processes such as reduction and rearrangement
reactions will be responsible for product formation.
61
8. Summary in Swedish
Som forskare inom området organisk kemi är man inte bara intresserad av att
studera hur atomer och molekyler reagerar utan även hur
reaktionsprodukterna kan användas för att förbättra livskvaliteten för
människor på olika sätt. Traditionellt sett framställs nya kol-kol bindningar
genom reaktioner mellan reagens som har utpräglat positiv (katjonisk)
karaktär och sådana som har en utpräglat negativ (anjonisk) karaktär.
Därutöver finns emellertid även reaktioner där reagensen saknar dessa
utpräglade negativa eller positiva laddningar. Här kan fria radikaler nämnas
som ett exempel. Radikaler är intressanta både ur ett kemiskt och biologiskt
perspektiv. Radikalreaktioner kan genomföras vid mycket låga temperaturer.
De sker på samma sätt oberoende av vilket lösningsmedel man väljer. På
senare tid har man utvecklat radikalreaktioner som uppvisar hög selektivitet
både vad gäller regiokemi och stereokemi. Detta har bidragit till att radikal-
medierade reaktioner numer ingår i den organiske kemistens vertygslåda för
framställning av nya föreningar.
Grundämnena selen och tellur brukar ofta förknippas med giftighet och
illaluktande organiska föreningar. Den organiska kemin för dessa två ämnen
är dock förvånansvärt rik och en mycket stor mängd selen- och
tellurorganiska föreningar finns beskrivna i litteraturen. I denna avhandling
har vi omvandlat lätttillgängliga selenorganiska föreningar på ett sådant sätt
att den radikal som bildas vid homolytisk klyvning av kol-selen bindningen
undergår en cykliseringsreaktion och ger upphov till kväveheterocykliska
föreningar – pyrroliner (Figur 9). Pyrroliner påträffas naturligt som
biosyntetiska mellanprodukter och som en del av strukturen hos feromoner,
62
alkaloider, hem- och klorofyllföreningar samt diverse marina toxiner. Dessa
kväveinnehållande molekyler är p.g.a. sin biologiska aktivitet av stort
intresse.
N
Figur 17. En substituerad pyrrolin.
Vi har även försökt skapa nya kol-kol bindningar från lättillgängliga selen-
och tellurorganiska föreningar genom att hetta upp dem m.h.a. mikrovågor.
Det har lyckats ibland att termiskt extrudera heteroatomen men metodiken
kan inte anses generellt användbar. Vi har därför även undersökt
möjligheterna att katalysera processen med hjälp av övergångsmetaller. Vi
har sålunda beskrivit reaktionsbetingelser där den aromatiska delen av
diaryltellurider och alkyl aryl tellurider kan kopplas till acetylener i närvaro
av katalytisk mängd av Pd(PPh3)4 (Sonogashirareaktionen).
Under det senaste årtiondet har man försökt hitta nya tillvägagångssätt för
behandling sjukdomar. ”Antisensterapi”, en metod baserad på den
biofysikaliska kemin hos DNA och RNA, har rönt stor uppmärksamhet där-
OO N
NH
O
O
CH3O
OO N
NH
O
O
OCH3
OO N
NH
O
O
OO
OO N
NH
O
O
O O
A B C D Figur 18. Nukleosidanaloger med intressanta biofysikaliska egenskaper. (A) Locked nucleic acid (LNA); (B) Etylene bridged nucleic acid (ENA); (C) Karbocyklisk LNA. (D) Karbocyklisk ENA. Strukturerna C och D är nyligen framtagna och diskuteras i denna avhandling.
63
vidlag, även om de terapeutiska tillämpningarna förmodligen ligger en bit
fram i tiden. I denna avhandling beskriver vi syntes av två nya
nukleosidanaloger (C och D; Figur 18) som har visat intressanta egenskaper
både vad gäller stabilitet och aktivitet. Nyckelsteget i syntesen av de båda
nya nukleosidanalogerna är en radikalcyklisering. De nya analogerna C och
D utvärderades med avseende på stabilitet och specificitet i jämförelse med
de redan kända strukturerna A och B. Den förlängda livslängden av dessa
karbocykliska analoger C och D i blodserum är en intressant farmakologisk
egenskap som borde kunna medföra att de kan doseras i mindre mängd.
Förhoppningsvis kan vår forskning bidra till att påskynda utvecklingen av
nya antisens- eller RNAi-terapeutiska läkemedel.
64
Acknowledgements
In the words written below I take the opportunity to thank everybody who
stood by me and supported me in this journey. I am already at loss of
words.‘Words’ mean much more than they say.
I would like to express my sincere gratitude to my supervisor Professor Lars
Engman for agreeing to be my supervisor, for good science and above all for
treating me like a human being.
Professor Adolf Gogoll, you were my pillar of support during these years. I
think I ran to you everytime I felt like giving up and had nowhere to go. A
big reason why I could reach the brighter side of the tunnel is YOU. I am
forever indebted.
Professor Lars Balzer for helping me out and showing me a different world.
Professor Helena Grenneberg for moral support and an ever smiling face.
Professor Joseph Samec for tolerating me and my smelly compounds. I wish
I had an iota of enthusiasm that you have.
I also wish to thank Prof. Henrik Ottoson, Prof. Olle Mattson and Prof. Pher
G. Anderson for being kind to me.
Thanks to Prof. Jyoti Chattopadhyaya and department of biorganic
chemistry for a fascinating chemistry.
Special thanks to Dr. Per Lowdin for excellent support and help. I am
indebted.
I would like to express my gratitute to department secretaries, Eva Pylvänen
and Johanna Johansson for helping me out with everything in the department.
65
Henrik Johansson the lab king. You have been very kind to me and
tolerated all my sayings and all that blah blah blah…, It seems I had a saying
for every occasion. I wish all the best for your future. Be happy.
Bobo for all the help and friendly disposition. Your sincerity, deligence and
calm are your virtues. Keep up the good spirit. Lycka till.
Jonas Rydfjord for being a friendly, honest and a ‘cool coworker’. For all the
help, encouraging discussions and for a memorable movie night with kids.
Your Dalarna souvenir will be cherished forever. I wish you a bright and
happy future ahead. Lycka till.
Zeyed Abdulkarim, the hardworking late night guy. For letting a sur gubbe
accompany the kids to a nice movie. I wish you a bright and happy future
ahead. Lycka till
Supaporn Sawadjoon for giving me nice and peaceful company. You are a
hard and sincere worker. I wish that your hard-work is rewarded. Good luck.
Sawadee
Dr. Khadijeh Bakhtiari the Dokhtar for being a such a good soul, for Hafiz,
Sadi, kalle pacche, kufteh-tabrizi, Shiraz, Isfahan, NMR, tellurium, fresh air,
coffee……the list continues. I wish happiness for you in future.
Anwar Merzae the Peshmerga birader for tolerating me and for being a nice
coworker also for cakes, chips……fresh air…...the list continues. I wish
good luck for your future.
I wish to thank Adam, Andreas, Alexander, Alvi, Christian, Diana, Johan,
Julius, Rikard, Sarah and special thanks to my NMR teacher Claes Henrik. I
thanks all other people from the department whose name I have missed, for
being friendly and kind to me.
I take this opportunity to express my love to my family in Uppsala.
Smita, Nimesh and Adwait; Pratima, Radha Raman and Prashant; Julie,
Varghese and Evleyn; Madhuri, Pradeep and Mayank; Navya and Sandeep,
Fatima, Faizan and Zidan; Kali and Sulena. The ‘bacchas’ in the family
66
Gaurav, Tanmoy and Prasoon. Special thanks to Jitendra the mamu of
‘bacchas’.
I will not be able to pen down my emotions here for each of you. You
have tolerated me over the years, assuaged my pain and insecurities and
above all changed me by providing so much of affection. I wish a happy and
fulfilling life to all the members.
Special Thanks to Jharna, the angry girl for all the discussions, food and
caring attitude.
To Subhrangshu for all the help and caring for me even when you are far
away. To Wimal for a great collaboration and his wife Ayesha for great food
and family atmosphere.
To my world, my mummy (this event came too late for you) and papa and my
sisters Nutan, Ushma and Mili for all the little amount of happiness that is
destined for me. To Madhukarjijaji, Ashwani Jijaji and Rahul for love
affection and care. Love to my kids Kislaya, Ananya, Arnav and Dhruv.
67
References
(1) Gomberg, M. J. Am. Chem. Soc. 1900, 22, 752-757.
(2) Paneth, F. A.; Loleit, H. J. Chem. Soc. 1935, 366-371.
(3) Paneth, F.; Hofeditz, W. Ber. Dtsch. Chem. Ges. B 1929, 62B, 1335-1347.
(4) Hey, D. H.; Waters, W. A. Chem. Rev. 1937, 21, 169-208.
(5) Halliwell, B.; Gutteridge, J. M. C. Free Radicals in Biology and Medicine;
Oxford University Press: New York, 2007.
(6) Griller, D.; Ingold, K. U. Acc. Chem. Res. 1980, 13, 317-323.
(7) Walling, C.; Cooley, J. H.; Ponaras, A. A.; Racah, E. J. J. Am. Chem. Soc. 1966,
88, 5361-5363.
(8) Hart, D. J. Science 1984, 223, 883-887.
(9) Giese, B. Radicals in Organic Synthesis: Formation of Carbon-Carbon Bonds;
Pergamon: Oxford, 1986.
(10) Beckwith, A. L. J. Tetrahedron 1981, 37, 3073-3100.
(11) Kharasch, M. S.; Potts, W. M. J. Org. Chem. 1937, 2, 195-197.
(12) Beckwith, A. L. J.; Moad, G. J. Chem. Soc., Chem. Commun. 1974, 472-473.
(13) Renaud, P.; Sibi, M. P. Radicals in Organic Synthesis; Vol. 1; Wiley-VCH:
Weinheim, 2001.
(14) Renaud, P.; Sibi, M. P. Radicals in Organic Synthesis; Vol 2; Wiley-VCH:
Weinheim, 2001.
(15) Waits, H. P.; Hammond, G. S. J. Am. Chem. Soc. 1964, 86, 1911-1918.
(16) Matsugi, M.; Gotanda, K.; Ohira, C.; Suemura, M.; Sano, A.; Kita, Y. J. Org.
Chem. 1999, 64, 6928-6930.
(17) Yorimitsu, H.; Wakabayashi, K.; Shinokubo, H.; Oshima, K. Bull. Chem. Soc.
Jpn. 2001, 74, 1963-1970.
(18) Nambu, H.; Anilkumar, G.; Matsugi, M.; Kita, Y. Tetrahedron 2003, 59, 77-85.
(19) Julia, M. Acc. Chem. Res. 1971, 4, 386-392.
68
(20) Julia, M. Pure Appl. Chem. 1974, 40, 553-567.
(21) Brown, H. C.; Midland, M. M. Angew. Chem., Int. Ed. Engl. 1972, 11, 692-700.
(22) Ollivier, C.; Renaud, P. Chem. Rev. 2001, 101, 3415-3434.
(23) Perchyonok, V. T.; Schiesser, C. H. Tetrahedron Lett. 1998, 39, 5437-5438.
(24) Barton D, H. R.; Mccombie, S. W. J. Chem Soc. Perkin Trans. 1 1975, 1574-
1585.
(25) Ono, N.; Kaji, A. Synthesis 1986, 693-704.
(26) Ono, N.; Miyake, H.; Tamura, R.; Kaji, A. Tetrahedron Lett. 1981, 22, 1705-
1708.
(27) Kuivila, H. G.; Menapace, L. W.; Warner, C. R. J. Am. Chem. Soc. 1962, 84,
3584-3586.
(28) Menapace, L. W.; Kuivila, H. G. J. Am. Chem. Soc. 1964, 86, 3047-3051.
(29) Kuivila, H. G. Acc. Chem. Res. 1968, 1, 299-305.
(30) Giese, B. Angew. Chem., Int. Ed. 1985, 97, 555-567.
(31) Ramaiah, M. Tetrahedron 1987, 43, 3541-3676.
(32) Curran, D. P. Synthesis 1988, 417-439.
(33) Curran, D. P. Synthesis 1988, 489-513.
(34) Jasperse, C. P.; Curran, D. P.; Fevig, T. L. Chem. Rev. 1991, 91, 1237-86.
(35) Chatgilialoglu, C.; Ingold, K. U.; Scaiano, J. C.; Woynar, H. J. Am. Chem. Soc.
1981, 103, 3231-3232.
(36) Johnston, L. J.; Lusztyk, J.; Wayner, D. D. M.; Abeywickreyma, A. N.;
Beckwith, A. L. J.; Scaiano, J. C.; Ingold, K. U. J. Am. Chem. Soc. 1985, 107, 4594-
4596.
(37) Rowlands, G. J. Tetrahedron 2009, 65, 8603-8655.
(38) Studer, A.; Amrein, S. Synthesis 2002, 835-849.
(39) Chatgilialoglu, C. Acc. Chem. Res. 1992, 25, 188-194.
(40) Chatgilialoglu, C.; Newcomb, M. Adv. Organomet. Chem. 1999, 44, 67-112.
(41) Chatgilialoglu, C. Chem. Eur. J. 2008, 14, 2310-2320.
(42) Chatgilialoglu, C.; Griller, D.; Lesage, M. J. Org. Chem. 1988, 53, 3641-3642.
(43) Motherwell, W. B.; Crich, D. Free radical chain reactions in organic synthesis;
Academic Press: London, 1992.
69
(44) Scaiano, J. C.; Schmid, P.; Ingold, K. U. J. Organomet. Chem. 1976, 121, C4-
C6.
(45) Crich, D.; Yao, Q. W. J. Org. Chem. 1995, 60, 84-88.
(46) Roberts, B. P. Chem. Soc. Rev. 1999, 28, 25-35.
(47) Cole, S. J.; Kirwan, J. N.; Roberts, B. P.; Willis, C. R. J. Chem. Soc., Perkin
Trans. 1 1991, 103-12.
(48) Crich, D.; Grant, D.; Krishnamurthy, V.; Patel, M. Acc. Chem. Res. 2007, 40,
453-463.
(49) Iwaoka, M.; Tomoda, S. Top. Curr. Chem. 2000, 208, 55-80.
(50) Tiecco, M. Top. Curr. Chem. 2000, 208, 7-54.
(51) Renaud, P. Top. Curr. Chem. 2000, 208, 81-112.
(52) Burke, S. D.; Fobare, W. F.; Armistead, D. M. J. Org. Chem. 1982, 47, 3348-
3350.
(53) Clive, D. L. J.; Chittattu, G. J.; Farina, V.; Kiel, W. A.; Menchen, S. M.;
Russell, C. G.; Singh, A.; Wong, C. K.; Curtis, N. J. J. Am. Chem. Soc. 1980, 102,
4438-4447.
(54) Barton, D. H. R.; Ramesh, M. J. Am. Chem. Soc. 1990, 112, 891-892.
(55) Han, L. B.; Ishihara, K.; Kambe, N.; Ogawa, A.; Ryu, I.; Sonoda, N. J. Am.
Chem. Soc. 1992, 114, 7591-7592.
(56) Chen, C.; Crich, D.; Papadatos, A. J. Am. Chem. Soc. 1992, 114, 8313-8314.
(57) Ericsson, C.; Engman, L. J. Org. Chem. 2004, 69, 5143-5146.
(58) Berlin, S.; Ericsson, C.; Engman, L. J. Org. Chem. 2003, 68, 8386-8396.
(59) Berlin, S.; Engman, L. Tetrahedron Lett. 2000, 41, 3701-3704.
(60) Gupta, V.; Besev, M.; Engman, L. Tetrahedron Lett. 1998, 39, 2429-2432.
(61) Yamago, S. Synlett 2004, 1875-1890.
(62) Curran, D. P.; Martin-Esker, A. A.; Ko, S. B.; Newcomb, M. J. Org. Chem.
1993, 58, 4691-4695.
(63) Newcomb, M. Tetrahedron 1993, 49, 1151-1176.
(64) Schiesser, C. H.; Skidmore, M. A. J. Organomet. Chem. 1998, 552, 145-157.
(65) Lamb, R. C.; Ayers, P. W.; Toney, M. K. J. Am. Chem. Soc. 1963, 85, 3483-
3486.
(66) Walling, C.; Cioffari, A. J. Am. Chem. Soc. 1972, 94, 6059-6064.
70
(67) Beckwith, A. L. J.; Phillipou, G.; Serelis, A. K. Tetrahedron Lett. 1981, 29,
2811-2814.
(68) Beckwith, A. L. J. Chem. Soc. Rev. 1993, 22, 143-151.
(69) Beckwith, A. L. J.; Blair, I. A.; Phillipou, G. Tetrahedron Lett. 1974, 2251-
2254.
(70) Giese, B. Angew. Chem., Int. Ed. 1983, 95, 771-782.
(71) Stork, G.; Baine, N. H. J. Am. Chem. Soc. 1982, 104, 2321-2323.
(72) RajanBabu, T. V. J. Org. Chem. 1988, 53, 4522-4530.
(73) RajanBabu, T. V. Acc. Chem. Res 1991, 24, 139-145.
(74) RajanBabu, T. V.; Fukunaga, T. J. Am. Chem. Soc. 1989, 111, 296-300.
(75) RajanBabu, T. V.; Fukunaga, T.; Reddy, G. S. J. Am. Chem. Soc. 1989, 111,
1759-1769.
(76) Baldwin, J. E. J. Chem. Soc., Chem. Comm. 1976, 18, 734-736.
(77) Baldwin, J. E. J. Chem. Soc., Chem. Comm. 1976, 18, 738-741.
(78) Beckwith, A. L. J.; Schiesser, C. H. Tetrahedron 1985, 41, 3925-3941.
(79) Spellmeyer, D. C.; Houk, K. N. J. Org. Chem. 1987, 52, 959-74.
(80) Wilt, J. W. Tetrahedron 1985, 41, 3979-4000.
(81) Padwa, A.; Nimmesgern, H.; Wong, G. S. K. J. Org. Chem. 1985, 50, 5620-
5627.
(82) Beckwith, A. L. J.; Page, D. M. J. Org. Chem. 1998, 63, 5144-5153.
(83) Beckwith, A. L. J.; Page, D. M. Tetrahedron 1999, 55, 3245-3254.
(84) Bailey, W. F.; Longstaff, S. C. Org. Lett. 2001, 3, 2217-2219.
(85) Jung, M. E.; Piizzi, G. Chem. Rev. 2005, 105, 1735-1766.
(86) Hanessian, S.; Dhanoa, D. S.; Beaulieu, P. L. Can. J. Chem. 1987, 65, 1859-
1866.
(87) Gandon, L. A.; Russell, A. G.; Gueveli, T.; Brodwolf, A. E.; Kariuki, B. M.;
Spencer, N.; Snaith, J. S. J. Org. Chem. 2006, 71, 5198-5207.
(88) Curran, D. P.; Chang, C. T. J. Org. Chem. 1989, 54, 3140-3157.
(89) Saenger, W. Principles of Nucleic Acid Structure; Springer-Verlag: Berlin,
1984.
(90) Bloomfield, V. A.; Crothers, D. M.; Tinoco, I., Jr. Nucleic Acids: Structures,
Properties and Functions; University Science Books: Sausalito, CA, 2000.
71
(91) Watson, J. D.; Crick, F. H. Nature 1953, 171, 737-738.
(92) Sugimoto, N.; Nakano, S.; Yoneyama, M.; Honda, K. Nucleic Acids Res. 1996,
24, 4501-4505.
(93) Sugimoto, N.; Nakano, S.; Katoh, M.; Matsumura, A.; Nakamuta, H.; Ohmichi,
T.; Yoneyama, M.; Sasaki, M. Biochemistry 1995, 34, 11211-11216.
(94) Sundaralingam, M. J. Am. Chem. Soc. 1965, 87, 599-606.
(95) Altona, C.; Sundaralingam, M. J. Am. Chem. Soc. 1973, 95, 2333-2344.
(96) Altona, C.; Sundaralingam, M. J. Am. Chem. Soc. 1972, 94, 8205-12.
(97) Dupouy, C.; Iche-Tarrat, N.; Durrieu, M.-P.; Rodriguez, F.; Escudier, J.-M.;
Vigroux, A. Angew. Chem., Int. Ed. 2006, 45, 3623-3627.
(98) Kilpatrick, J. E.; Pitzer, K. S.; Spitzer, R. J. Am. Chem. Soc. 1947, 69, 2483-
2488.
(99) Hopkins, A. L.; Groom, C. R. Nat. Rev. Drug Discovery 2002, 1, 727-730.
(100) Scherer, L. J.; Rossi, J. J. Nat. Biotechnol. 2003, 21, 1457-1465.
(101) Giovannangeli, C.; Helene, C. Nat. Biotechnol. 2000, 18, 1245-1246.
(102) Giovannangeli, C.; Helene, C. Curr. Opin. Mol. Ther. 2000, 2, 288-296.
(103) Roberts, R. W.; Crothers, D. M. Science 1992, 258, 1463-1466.
(104) Roberts, R. W.; Crothers, D. M. Proc. Natl. Acad. Sci. USA 1991, 88, 9397-
9401.
(105) Hoogsteen, K. Acta Crystallogr. 1963, 16, 907-916.
(106) Hoogsteen, K. Acta Crystallogr. 1959, 12, 822-823.
(107) Zamecnik, P. C.; Stephenson, M. L. Proc. Natl. Acad. Sci. USA 1978, 75, 280-
284.
(108) Crooke, S. T. Annu. Rev. Med. 2004, 55: , 61-95.
(109) Opalinska, J. B.; Gewirtz, A. M. Nat. Rev. Drug Discovery 2002, 1, 503-514.
(110) Wengel, J. Acc. Chem. Res 1999, 32, 301-310
(111) Obika, S.; Nanbu, D.; Hari, Y.; Morio, K.-I.; In, Y.; Ishida, T.; Imanishi, T.
Tetrahedron Lett. 1997, 38, 8735-8738.
(112) Leumann, C. J. Bioorg. Med. Chem. 2002, 10, 841-854.
(113) Steffens, R.; Leumann, C. J. J. Am. Chem. Soc. 1997, 119, 11548-11549.
(114) Koshkin, A. A.; Singh, S. K.; Nielsen, P.; Rajwanshi, V. K.; Kumar, R.;
Meldgaard, M.; Olsen, C. E.; Wengel, J. Tetrahedron 1998, 54, 3607-3630
72
(115) Petersen, M.; Nielsen, C. B.; Nielsen, K. E.; Jensen, G. A.; Bondensgaard, K.;
Singh, S. K.; Rajwanshi, V. K.; Koshkin, A. A.; Dahl, B. M.; Wengel, J.; Jacobsen, J.
P. J. Mol. Recogn. 2000, 13, 44-53.
(116) Singh, S. K.; Kumar, R.; Wengel, J. J. Org. Chem. 1998, 63, 10035-10039.
(117) Varghese, O. P.; Barman, J.; Pathmasiri, W.; Plashkevych, O.; Honcharenko,
D.; Chattopadhyaya, J. J. Am. Chem. Soc. 2006, 128, 15173-15187.
(118) Pradeepkumar, P. I.; Chattopadhyaya, J. J. Chem. Soc., Perkin Trans. 2 2001,
2074-2083.
(119) Honcharenko, D.; Varghese, O. P.; Plashkevych, O.; Barman, J.;
Chattopadhyaya, J. J. Org. Chem. 2006, 71, 299-314.
(120) Teplova, M.; Minasov, G.; Tereshko, V.; Inamati, G. B.; Cook, P. D.;
Manoharan, M.; Egli, M. Nat. Struct. Biol. 1999, 6, 535-539.
(121) Prakash, T. P.; Pueschl, A. P.; Lesnik, E.; Mohan, V.; Tereshko, V.; Egli, M.;
Manoharan, M. Org. Lett. 2004, 6, 1971-1974.
(122) Kanazaki, M.; Ueno, Y.; Shuto, S.; Matsuda, A. J. Am. Chem. Soc. 2000, 122,
2422-2432.
(123) Melhado, A. D.; Luparia, M.; Toste, F. D. J. Am. Chem. Soc. 2007, 129,
12638-12639.
(124) Savarin, C. G.; Grise, C.; Murry, J. A.; Reamer, R. A.; Hughes, D. L. Org. Lett.
2007, 9, 981-983.
(125) Peddibhotla, S.; Tepe, J. J. J. Am. Chem. Soc. 2004, 126, 12776-12777.
(126) Shvekhgeimer, M. G. A. Chem. Heterocycl. Compd. 2003, 39, 405-448.
(127) Narasaka, K. Pure Appl. Chem. 2003, 75, 19-28.
(128) Kondo, T.; Okada, T.; Mitsudo, T. J. Am. Chem. Soc. 2002, 124, 186-187.
(129) Ahn, Y.; Cardenas, G. I.; Yang, J.; Romo, D. Org. Lett. 2001, 3, 751-754.
(130) Jasperse, C. P.; Curran, D. P.; Fevig, T. L. Chem. Rev. 1991, 91, 1237-86.
(131) Bowman, W. R.; Bridge, C. F.; Brookes, P. Tetrahedron Lett. 2000, 41, 8989-
8994.
(132) Fallis, A. G.; Brinza, I. M. Tetrahedron 1997, 53, 17543-17594.
(133) Zard, S. Z. Synlett 1996, 1148-1154.
(134) Boivin, J.; Fouquet, E.; Zard, S. Z. Tetrahedron Lett. 1990, 31, 85-88.
73
(135) Houllemare, D.; Ponthieux, S.; Outurquin, F.; Paulmier, C. Synthesis 1997,
101-106.
(136) Engman, L. J. Org. Chem. 1988, 53, 4031-4037.
(137) Engman, L. Tetrahedron Lett. 1985, 26, 6385-6388.
(138) Curtin, D. Y.; Grubbs, E. J.; McCarty, C. G. J. Am. Chem. Soc. 1966, 88, 2775-
2786.
(139) Ostrogovich, G.; Kerek, F.; Simon, Z. J. Chem. Soc. (B), 1971, 541-544.
(140) Chatgilialoglu, C.; Dickhaut, J.; Giese, B. J. Org. Chem. 1991, 56, 6399-6403.
(141) Johnson, A. W.; Oldfield, D.; Rodrigo, R.; Shaw, N. J. Chem. Soc. 1964, 4080-
4082.
(142) Wu, J. C.; Xi, Z.; Gioeli, C.; Chattopadhyaya, J. Tetrahedron 1991, 47, 2237-
2254.
(143) Dickson, J. K., Jr.; Tsang, R.; Llera, J. M.; Fraser-Reid, B. J. Org. Chem. 1989,
54, 5350-5356.
(144) Tsang, R.; Dickson, J. K., Jr.; Pak, H.; Walton, R.; Fraser-Reid, B. J. Am.
Chem. Soc. 1987, 109, 3484-3486.
(145) Tsang, R.; Fraser-Reid, B. J. Am. Chem. Soc. 1986, 108, 2116-2117.
(146) Fraser-Reid, B.; Tulshian, D. B.; Tsang, R.; Lowe, D.; Box, V. G. S.
Tetrahedron Lett. 1984, 25, 4579-4582.
(147) Hansen, S. G.; Skrydstrup, T. Top. Curr. Chem. 2006, 264, 135-162.
(148) Giese, B.; Zeitz, H.-G. In Preparative Carbohydrate Chemistry; Hanessian, S.,
Ed.; Marcel. Dekker Inc.: New York, 1997, p 507-525.
(149) Ferrier, R. J.; Middleton, S. Chem. Rev. 1993, 93, 2779-2831.
(150) Ferrier, R. J.; Prasit, P. Pure Appl. Chem. 1983, 55, 565-576.
(151) Wilcox, C. S.; Gaudino, J. J. J. Am. Chem. Soc. 1986, 108, 3102-3104.
(152) Wilcox, C. S.; Thomasco, L. M. J. Org. Chem. 1985, 50, 546-547.
(153) Rajanbabu, T. V. In Preparative Carbohydrate Chemistry; Hanessian, S., Ed.;
Marcel Dekker Inc.: New York, 1997, p 545-568.
(154) Giese, B.; Carl, B.; Carl, T.; Carell, T.; Behrens, C.; Hennecke, U.; Schiemann,
O.; Feresin, E. Angew. Chem., Int. Ed. 2004, 43, 1848-1851.
(155) Giese, B. Acc. Chem. Res. 2000, 33, 631-636.
74
(156) Hong, I. S.; Ding, H.; Greenberg, M. M. J. Am. Chem. Soc. 2006, 128, 485-
491.
(157) Stubbe, J.; van der Donk, W. A. Chem. Rev. 1998, 98, 705-762.
(158) Reichard, P. Science 1993, 260, 1773-1777.
(159) Mancuso, A. J.; Brownfain, D. S.; Swern, D. J. Org. Chem. 1979, 44, 4148-
4150.
(160) Wittig, G.; Schlosser, M. Angew. Chem. 1960, 72, 324.
(161) Vorbrüggen, H.; Höfle, G. Chem. Ber. 1981, 114, 1256.
(162) Bax, A.; Summers, M. J. Am. Chem. Soc. 1986, 108, 2093-2094
(163) Bax, A.; Davis, D. G. J. Magn. Reson. 1985, 65, 355-360.
(164) Koji Morita, M. T., Chikako Hasegawa, Masakatsu Kaneko, Shinya Tsutsumi,
Junko Sone, Tomio Ishikawa, Takeshi Imanishi and Makoto Koizumi Bioorg. Med.
Chem. 2003, 11, 2211-2226.
(165) Graessmann, M.; Michaels, G.; Berg, B.; Graessmann, A. Nucleic Acids Res.
1991, 19, 53-59.
(166) Wagner, R. W.; Matteucci, M. D.; Lewis, J. G.; Gutierrez, A. J.; Moulds, C.;
Froehler, B. C. Science 1993, 260, 1510-13.
(167) Marquez, V. E.; Siddiqui, M. A.; Ezzitouni, A.; Russ, P.; Wang, J.; Wagner, R.
W.; Matteucci, M. D. J. Med. Chem. 1996, 39, 3739-3747.
(168) Grunweller, A.; Wyszko, E.; Bieber, B.; Jahnel, R.; Erdmann Volker, A.;
Kurreck, J. Nucleic Acids Res. 2003, 31, 3185-3193.
(169) Ambros, V. Cell 2001, 107, 823-826.
(170) Ruvkun, G. Science 2001, 294, 797-799.
(171) Buchini, S.; Leumann, C. J. Current Opin. Chem. Biol. 2003, 7, 717-726.
(172) Altmann, K.-H.; Bevierre, M.-O.; De Mesmaeker, A.; Moser, H. E. Bioorg.
Med. Chem. Lett. 1995, 5, 431-436.
(173) Monia, B. P.; Johnston, J. F.; Sasmor, H.; Cummins, L. L. J. Biol. Chem. 1996,
271, 14533-14540.
(174) Pattanayek, R.; Sethaphong, L.; Pan, C.; Prhavc, M.; Prakash, T. P.;
Manoharan, M.; Egli, M. J. Am. Chem. Soc. 2004, 126, 15006-15007.
75
(175) Egli, M.; Minasov, G.; Tereshko, V.; Pallan, P. S.; Teplova, M.; Inamati, G. B.;
Lesnik, E. A.; Owens, S. R.; Ross, B. S.; Prakash, T. P.; Manoharan, M.
Biochemistry 2005, 44, 9045-9057.
(176) Morita, K.; Hasegawa, C.; Kaneko, M.; Tsutsumi, S.; Sone, J.; Ishikawa, T.;
Imanishi, T.; Koizumi, M. Bioorg. Med. Chem. Lett. 2001, 12, 73-76.
(177) Teplova, M.; Wallace, S. T.; Tereshko, V.; Minasov, G.; Symons, A. M.;
Cook, P. D.; Manoharan, M.; Egli, M. Proc. Natl. Acad. of Sci. of the USA 1999, 96,
14240-14245.
(178) Kurreck, J.; Wyszko, E.; Gillen, C.; Erdmann, V. A. Nucleic Acids Res. 2002,
30, 1911-1918.
(179) Cuthbertson, E.; MacNicol, D. D. Tetrahedron Lett. 1975, 1893-1894.
(180) Higuchi, H.; Misumi, S. Tetrahedron Lett. 1982, 23, 5571-5574.
(181) Boekelheide, V. Acc. Chem. Res. 1980, 13, 65-70.
(182) Bergman, J. Tetrahedron 1972, 28, 3323-3331.
(183) Engman, L.; Cava, M. P. J. Org. Chem. 1981, 46, 4194-4197.
(184) Bergman, J.; Engman, L. Org. Prep. Proced. Int. 1978, 10, 289-290.
(185) Koketsu, M.; Nada, F.; Hiramatsu, S.; Ishihara, H. J. Chem. Soc., Perkin Trans.
1 2002, 737-740.
(186) Petragnani, N.; Comasseto, J. V. Synthesis 1986, 1-30.
(187) Petragnani, N.; Comasseto, J. V. Synthesis 1991, 897-919.
(188) Petragnani, N.; Comasseto, J. V. Synthesis 1991, 793-817.
(189) Petragnani, N.; Stefani, H. A. Tetrahedron 2005, 61, 1613-1679.
(190) Bergman, J.; Engman, L. J. Organomet. Chem. 1979, 175, 233-237.
(191) Uemura, S.; Wakasugi, M.; Okano, M. J. Organomet. Chem. 1980, 194, 277-
283.
(192) Ohe, K.; Takahashi, H.; Uemura, S.; Sugita, N. J. Org. Chem. 1987, 52, 4859-
4863.
(193) Barton, D. H. R.; Ozbalik, N.; Ramesh, M. Tetrahedron Lett. 1988, 29, 3533-
3536.
(194) Nishibayashi, Y.; Cho, C. S.; Uemura, S. J. Organomet. Chem. 1996, 507, 197-
200.
76
(195) Nishibayashi, Y.; Cho, C. S.; Ohe, K.; Uemura, S. J. Organomet. Chem. 1996,
526, 335-339.
(196) Kang, S.-K.; Lee, S.-W.; Ryu, H.-C. Chem. Commun. 1999, 2117-2118.
(197) Braga, A. L.; Ludtke, D. S.; Vargas, F.; Donato, R. K.; Silveira, C. C.; Stefani,
H. A.; Zeni, G. Tetrahedron Lett. 2003, 44, 1779-1781.
(198) Alves, D.; Schumacher, R. F.; Brandao, R.; Nogueira, C. W.; Zeni, G. Synlett
2006, 1035-1038.
(199) Cella, R.; Cunha, R. L. O. R.; Reis, A. E. S.; Pimenta, D. C.; Klitzke, C. F.;
Stefani, H. A. J. Org. Chem. 2006, 71, 244-250.
(200) Zeni, G.; Alves, D.; Pena, J. M.; Braga, A. L.; Stefani, H. A.; Nogueira, C. W.
Org. Biomol. Chem. 2004, 2, 803-805.
(201) Singh, F. V.; Amaral, M. F. Z. J.; Stefani, H. A. Tetrahedron Lett. 2009, 50,
2636-2639.
(202) Zeni, G.; Ludtke, D. S.; Nogueira, C. W.; Panatieri, R. B.; Braga, A. L.;
Silveira, C. C.; Stefani, H. A.; Rocha, J. B. T. Tetrahedron Lett. 2001, 42, 8927-
8930.
(203) Zeni, G.; Nogueira, C. W.; Panatieri, R. B.; Silva, D. O.; Menezes, P. H.;
Braga, A. L.; Silveira, C. C.; Stefani, H. A.; Rocha, J. B. T. Tetrahedron Lett. 2001,
42, 7921-7923.
(204) Singh, F. V.; Stefani, H. A. Synlett 2008, 3221-3225.
(205) Sonogashira, K.; Tohda, Y.; Hagihara, N. Tetrahedron Lett. 1975, 4467-4470.
(206) Engman, L.; Persson, J. Synth. Commun. 1993, 23, 445-458.
(207) Engman, L.; Stern, D. Organometallics 1993, 12, 1445-1448.
(208) Cunha, R. L. O. R.; Omori, A. T.; Castelani, P.; Toledo, F. T.; Comasseto, J. V.
J. Organomet. Chem. 2004, 689, 3631-3636.
(209) Haller, W. S.; Irgolic, K. J. J. Organomet. Chem. 1972, 38, 97-103.
(210) Miyabe, H.; Ueda, M.; Fujii, K.; Nishimura, A.; Naito, T. J. Org. Chem. 2003,
68, 5618-5626.
(211) Clive, D. L. J.; Pham, M. P.; Subedi, R. J. Am. Chem. Soc. 2007, 129, 2713-
2717.
(212) Kolasa, T.; Sharma, S. K.; Miller, M. J. Tetrahedron 1988, 44, 5431-5440.
77
(213) Gandon, L. A.; Russell, A. G.; Snaith, J. S. Org. Biomol. Chem. 2004, 2, 2270-
2271.
(214) Curran, D. P.; Porter, N. A.; Giese, B.; Editors Stereochemistry of Radical
Reactions: Concepts, Guidelines, and Synthetic Applications; VCH Publishers, Inc:
New York, 1996.
(215) Buffat, M. G. P. Tetrahedron 2004, 60, 1701-1729.
(216) Katritzky, A. R.; Rachwal, S. Chem. Rev. 2010, 110, 1564-1610.
(217) Pearson, M. S. M.; Mathe-Allainmat, M.; Fargeas, V.; Lebreton, J. Eur. J. Org.
Chem. 2005, 2159-2191.
(218) Felpin, F.-X.; Lebreton, J. Eur. J. Org. Chem. 2003, 3693-3712.
(219) Weintraub, P. M.; Sabol, J. S.; Kane, J. M.; Borcherding, D. R. Tetrahedron
2003, 59, 2953-2989.
(220) Laschat, S.; Dickner, T. Synthesis 2000, 1781-1813.
(221) Prevost, N.; Shipman, M. Org. Lett. 2001, 3, 2383-2385.
(222) Sjöholm, A.; Hemmerling, M.; Pradeille, N.; Somfai, P. J. Chem. Soc., Perkin
Trans. 1 2001, 891-899.
(223) Gottlich, R. Synthesis 2000, 1561-1564.
(224) Hemmerling, M.; Sjöholm, A.; Somfai, P. Tetrahedron: Asymmetry 1999, 10,
4091-4094.
(225) Parsons, A. F.; Pettifer, R. M. J. Chem. Soc., Perkin Trans. 1 1998, 651-660.
(226) Parsons, A. F.; Pettifer, R. M. Tetrahedron Lett. 1997, 38, 5907-5910.
(227) Naito, T.; Tajiri, K.; Harimoto, T.; Ninomiya, I.; Kiguchi, T. Tetrahedron Lett.
1994, 35, 2205-2206.
(228) Yoo, S. E.; Yi, K. Y.; Lee, S. H.; Jeong, N. Synlett 1990, 575-576.
(229) Besev, M.; Engman, L. Org. Lett. 2000, 2, 1589-1592.
(230) Besev, M.; Engman, L. Org. Lett. 2002, 4, 3023-3025.
(231) Katritzky, A. R.; Kirichenko, N.; Rogovoy, B. V.; He, H.-Y. J. Org. Chem.
2003, 68, 9088-9092.
(232) Katritzky, A. R.; He, H.-Y.; Jiang, R.; Long, Q. Tetrahedron: Asymmetry 2001,
12, 2427-2434.
(233) Franz, J. A.; Barrows, R. D.; Camaioni, D. M. J. Am. Chem. Soc. 1984, 106,
3964-3967.
78
$������������������.������������������ �������������� ������������������� ������� ����������������������������������������
����-��/��0���1��/�����,����1�!������������/�2�
$�������������������1����/�����,����1�!�����������/�2�3��..��������������3����,�,��������,������1��,�����1�.�.���4�$�1�5��.����1��/����.�������������������6�.�������7��!5����/��������/����������3�5/�����/��,��������������������,�����������������/�,2/�/���������0�2�������.��/������!,��������1��..����0�����������1����/�����,����1�!������������/�2�48�������9�,���3�&++�3��/���������5���.,����/���,�����/�������:��.��/������!,��������1��..�����0����������1����/�����,����1�!������������/�2�;4<
0������,��-�.,��������4,,4��,�-�-��-,,-������&*(%+
������������������� �����
����� �����