lw masters seminar
TRANSCRIPT
Linshu WangAdvisor: Prof. Jakobsche
Towards Hydrazine-Functionalized Peptides as Potential Lysyl Oxidase Inhibitors and a Four-Step
Amine to Alcohol Conversion via N-Nitrosoamides
Clark University Master’s Seminar
08/12/2016Master’s Seminar
Overview
a potent and selective inhibitor metastasis-related
enzyme Lysyl Oxidase (LOX)
Goal:
1. Synthesis of Small Molecules and Amino Acids for LOX Inhibition
Projects:
NH
NH
NH2
O
O
O
NH
NH2
semicarbazide carbazate
H2N OH
O
NHNH2
H2N OH
O
O
NHNH2
O
HN N
H
O
HNNH2
PeptidePeptide H2N OH
O
NH2Lysine
PgHN OH
O
OH6-Hydroxynorleucine
Key conversion
MeO
O
NH29 MeO
O
OH9model molecule
2. Study of the Amine to Alcohol Conversion via N-Nitrosoamides
MeO
O
NH9 R
O
MeO
O
N9 R
O
N O
MeO
O
O9 R
O
NaNO2AcOHAc2O
K2CO3MeOH
Introduction
Why are we interested in Lysyl oxidase (LOX)?upregulated during cancer metastasis
down regulated in other cases
Lung adenocarcinoma invasion and metastasis
silenced in human gastric cancers
What is LOX?HN Peptide
O
H
PeptideHN Peptide
O
NH3
Peptidean enzymeoxidases
in elastin and collagentwo major structural proteinsextracellular matrix (ECM)
peptidyl lysine
peptidyl AAS
O2 H2O NH4 H2O2
O
-aminoadipic--semialdehyde
Introduction
HN Peptide
O
N
Lysine
HN Peptide
O
H
Peptide
PeptideHN Peptide
O
H
Peptide
AAS AAS
HN Peptide
O
NH2
Peptide
HN Peptide
O
Peptide
NH
O
H
O
Peptide
Peptide
Formation of Schiff base
Aldol condensation
Cross linkage reactions of ECM proteins
O O
NHO
Peptide
Peptide
Introduction
amino acid sequences of LOX has been reported3D structure is still unknown
crystalization of LOX has not been successful
Lysyl tyrosylquinone (LTQ) cofactorfunctional part of LOX catalysis
O
O
H2C
NH
NH
O
HN
OLys 314
Tyr 349
ON
NH
Lysyl Oxidase
RCH2NH2
H2O
ROH
N
NH
R:B
OHNH2
NH
Lysyl Oxidase
ONH
NH
Lysyl Oxidase
O2
H2O2
H2O
NH3
H2O
R H
O
Substrate lysine
Catalytic Cycle
Lysyl Oxidase
OH
NH
Lysyl Oxidase
NN
Ph
azo tautomer(favored)
InhibitionMechanism
PhHN NH2
O
NH
Lysyl Oxidase
NNH
Ph:B
Phenylhydrazine inhibits the LTQ cofactor covalently and irreversibly
Introduction
OH
Tyr 349NH2
Lys 314
Lysyl Oxidase
Cu2+
oxidation
OO
NH
Lysyl Oxidase
Introduction
To better understand LOX's effects in cancer metastasis and develop new treatments
Small molecule inhibitors
HO O
O
HNN
H
O
NH
O
O
O NHNH2
H2N OH
O
NHNH2
HO O
O
HNN
H
O
NH
O
NHNH2
O
HNN
H
O
NH
O
NHNH2
Amino acids building blocks that mimic lysine
Peptide inhibitors
H2N OH
O
NH2
Lysine
H2N OH
O
O
NHNH2
O
Selective and potent inhibitors
Our approach:
Synthesis
Analytical methods:1H NMR, 13C NMR, COSY NMRLC-MS
HN
NH2 NH
NH
NH2
O
O NHNH2
O
O NH
NH2
S ONH2N
HN
HN
NH2
O
NH
NH2
phenylhydrazine semicarbazide
carbazate thiocarbonyl hydrazide pyrrolidine semicarbazide
alkyl hydrazine acyl hydrazine
Syntheses of Small molecule inhibitors
H2N NHBocCuI 0.05 eq, DMSO 1M,
Cs2CO3 1.5 eq, 50 oC 18 h.
NNH2
Boc
I
Column24% y
99% y
HN
NH2 HCl
Synthesis of phenyl hydrazine hydrochloride salt
Ullmann type coupling reaction
1.2 eq
1 eqHCl 17 eq
dioxane 0.16 MN2, RT 4 h
Model for more complex systems
NH
NH
NHBocO
NC
ONH2
column11% y
DCM 0.5 MNaHCO3 (sat. in H2O)
0 °C 30min
H2N NHBoc
0 °C 15min then RT 15h
NH
NH
NHBocO
NH
NH
NH2
O
97% yHCl
HCl / dioxane
O O Cl
O
Cl
Cl Cl Cl Cl
H2N NH2 H2O
Synthesis of semicarbazide hydrochloride salta modified one-pot reaction
The known methodology usedhydrazine monohydrate
by Ashley Burke
isocyanate intermediateseperate the org. phase
reported 4 h reaction time for the second step
0.33 eq
1 eq
Syntheses of Small molecule inhibitors
Synthesis of pyrrolidine semicarbazide trifluoroacetic acid salt
NHN
DCM 0.36 M
N2, 0 C then RT 1hy 46%
O O Cl
O
Cl
Cl Cl Cl Cl0.39 eq
2 eq
N2H4H2O in EtOH N NH
NH2
O
N O
O
N Cl
O
N2H4H2O in THFnot clean, can't be purified by column
O
CF3ON NH
NHBocO
N Cl
O
y 24%
H2N NHBoc
N
reflux 1.5 h
column y 12%
ONH3N
HN
TFA 0.16 eq, DCM 0.12 M
3.5 h RT
1 eq
16 eq 10 M
the reaction does not occur without pyr
O NH
NH2
SC SS
EtOH 3.2 eqKOH 1.1 eq, H2O 13 M,
RT 2 hN2H4H2O 1.1 eq
40 C 1.5 hO S
S
61% mass recovery
Synthesis of thiocarbonyl hydrazide one-pot reaction
xanthate intermediateyellow
colorless crude product,become pink overtimemight be light sensitivedecomepose overtimevia radical reactions
Syntheses of Small molecule inhibitors
OH
O
N N NN
O
DCM 0.48 MN2, RT,30 min
O
N
N
N
HN
y 70%
byproduct
O
O
HN
NHBocH2N NHBoc
NEt3 1.5 eq, 100 °Covernight, Tol 0.025 M
Attempt of synthesis of boc-protected carbazate
Conclusion: a different intermediate was needed.
CDI, similar to acid chlorideless reactive
1.2 eq1.1 eq
reduced reactivity compare to
hydrazine monohydrate
Syntheses of Small molecule inhibitors
O NHNH2
O
O Cl
O
N2H4H2O THF 0.39 M
0 C then RT overnight
Attempt of synthesis of hydrazine ethyl ester
4 eqThe more reactive chloroformate was tested
Direct conversion to hydrazide was tested first
column purificatrion
confirmed by LC-MS
O NH
O HN O
O61% y
O Cl
OO
H2N NHBoc
NEt3 1.5 eq, THF 0.53 Mwater bath then RT overnight,
column purificationy 21%, can be optimized
confirmed by LC-MS
Boc-protection was consideredInitial attempted reaction of boc-hydrazide substitution.
1 eq
Conclusion: the conversion was proved to be possible
HN
ONH
O
O
Syntheses of Small molecule inhibitors
O O Cl
O Cl
Cl
ClClCl
0.39 eq
OH O
O
HN
NHBocO Cl
O
O O
O
N2 , DCM 0.12MN 2 eq
H2N NHBoc 1 eq
NEt3 1.5 eq THF , RT overnight,
mostly productjust filtration of white solids0 C then RT overnight
partially volatilereacts with water
aqueous work up
Final reaction conditions:
OHO Cl
OO O Cl
O Cl
Cl
ClClCl
0.52 eq
DMF 0.035 eqNa2CO3 1 eq
Tol 0.25MN2 , 0 °C 8h
Na2CO3 removed by filtration
N
THF 0.61M , RT overnight
O
O
HN
NHBoc
H2N NHBoc 1.5 eq
1.4 eqcolumn purification
y 39%can be optimized
The second step reaction with extra DMF was tested with ethyl chloroformate.The reaction was not affected.
DMF is known to activate triphosgene to prepare liquid phosgene
one-pot reaction
Syntheses of Small molecule inhibitors
O
CF3OO
O
HN
NHBoc
TFA, DCM 3.5h RT
O
O
HN
NH3
O
O
HN
NHBocO
O
HN
NH2
HCl / dioxane
HCl
Attempts of deprotection reactions:
Conclusion: a different group on the hydrazide is needed to achieve a clean deprotection
Alternative: Cbz protecting group
O Cl
O
O NH
O HN O
O
Na2CO3 removed by filtration
H2NHN O
O
N
THF, RT overnight
O NH
ONH2
Pd/C, H2
H2NHN O
OO Cl
O
N2H4H2O 1.5 eqTHF 0.23M
Na2CO3 2 eq
2h 0°C
Exact Mass: 166.07
NH
HN O
OO
O
Exact Mass: 300.11
O O
O
Exact Mass: 242.09
Possible byproduct
reaction not cleanmostly product with
unidentified byprodect LC-MS m/z 236 (ESI)
Syntheses of Small molecule inhibitors
Summary
HN
NH2 NH
NH
NH2
O
O NHNH2
O
O NH
NH2
S ONH2N
HN
HN
NH2
O
NH
NH2
phenylhydrazine semicarbazide
carbazate thiocarbonyl hydrazide pyrrolidine semicarbazide
alkyl hydrazine acyl hydrazine
Syntheses by Ashley Burke
Initial inhibition data collected by Ashley Burke, Maria Solares and Lizzy Severson
More characterization and synthesis needed
Not stable upon storageDeprotection reactions not clean
Part 2 of project 1Syntheses of hydrazine functionalized amino acid
and dipeptides
Clark University Master’s Seminar
08/12/2016Master’s Seminar
Syntheses of hydrazine functionalized amino acid and dipeptides
HO O
O
HNN
H
O
NH
O
O
O NHNH2
HO O
O
HNN
H
O
NH
O
NHNH2
O
HNN
H
O
NH
O
NHNH2
Same length side chain as lysine
Compare affinity and see if the side chain charge is a factor
H2N OH
O
NBocNHBoc
NH
O
N
O
O
O
O
Key coupling reactions of succinimide esters
tBu O O
NH
O
N
O
O
O
O
H2NOH
OH
O
L-serine
tBu O O
O
OHH2N
tBu O O
O
HNN
H
O
NH
O
OH
boc-protected glutamic acid
Couple firstthen the conversion of functional group
O
OHH2N
L-alanine
H2N OH
O
OH
H2NOH
O
OHOL-glutamic acid
Key conversion
Syntheses of hydrazine functionalized amino acid and dipeptides
H2N OH
O
OH
Key conversion
previously reported by Majumdar group in 201224% y over 5 steps
Bn2NOBn
O
NBocNHBoc
H2NOH
O
OHO
H2N OH
O
NBocNHBoc
Bn2NOBn
O
OH
H2NOH
O
OHO
Bn2N OBn
O
OBn
BnBr, H2ODiBAl-H
THFreflux 1 hcolumny 61%
Ice bath 2 hcolumny 58%
K2CO3, KOH
O
selective reduction
Bn2NOBn
O
NBocNHBoc
BocN=NBocBocNH-NHBoc
PPh3,THF
N2, RT filtrationcolumn
One step Mitsunobu-like reactiondeveloped by Nicholas S. MacArthur
H2N OH
O
NBocNHBoc
H2Pd / C (500mg/mmol)
MeOH
triphenyl phosphene oxide byproduct
residue can interact with Pd catalyst loses reactivity
successful only once in over 10 attempts
Syntheses of hydrazine functionalized amino acid and dipeptides
Bn2NOBn
O
OH
Bn2NOBn
O
OMs
Bn2NOBn
O
NBocNHBoc
Pyr(0.84 eq)
MsCl(0.56 eq)
DCM, N2, Ice bath 2 h
Bochydrazine(1.5 eq)
DMF(0.3M) RT 24 h
Cs2CO3(2 eq)
Bn2NOBn
O
Br
Bn2NOBn
O
OH
PBr3 2 eqDCM 0.23 M
Two intermediates were tested
RT
decomposes during 2D-TLCConversion is almost complete
over 24 h at RT
N
OOBn
Ph
Ph
N
OOBn
Ph
Ph
RT
The only product recovered
COSY-1H NMR
2 steps y 9%
To avoid by productused upon synthesis
Conclusion: these two strategies were not successful
N
OO
Ph
Ph
ba
Hc
d
ef
g
Ph
Ar d
b
d/b
a/ca
g g e fe/f
OMs
NO
O
OMs
Ph
Ph
ab
a bd
Ph
dHce f
g
OMsAr
pyr pyr g c
EtOAc
e/f EtOAc
deshielded because of positively charged N, shifted downfieldN
OO
Ph
HN
OO
Ph
H
d-db-b
a-a
g-g
e-c e-c
g-f g-f
g-f
f-e f-e
f-e
db
d/b
a/c a
g g e f e/f
N
OO
Ph
Ph
ba
Hc
d
ef
g
Ph
Syntheses of hydrazine functionalized amino acid and dipeptides
H2NOH
O
OHO
Bn2NOBn
O
OH
Bn2NOBn
O
OBn
BnBr, H2O DiBAl-H
THFreflux 1 hcolumny 61%
Ice bath 2 hcolumny 58%
K2CO3, KOH
O
H2N OH
O
NBocNHBoc
H2Pd / C (500mg/mmol)
MeOH
85% mass recoveryover 2 steps,70% purity
Bn2NOBn
O
NBocNHBoc
BocN=NBocBocNH-NHBoc
PPh3,THF
N2, RT overnightfiltrationcolumn
monitored by TLCadditional hydrazone and PPh3 were added after 5h.
Removal of triphenyl phosphene oxide byproduct:the bulk of by product was removed by addition of Et2O and gravity filtration of visible solids consecutively.column chromatography.
The bochydrazine has similar polarity as the product. Some reamined in the mixture after the purifications. Bochydrazine did not affect the hydrogenolysis reaction.
Conclusion: optimization is still needed.
Syntheses of hydrazine functionalized amino acid and dipeptides
HO O
O
HNN
H
O
NH
O
NHNH2
O
HNN
H
O
NH
O
NHNH2
NH
O
N
O
O
O
O
tBu O O
NH
O
N
O
O
O
O
H2N OH
O
NBocNHBoc
H2N OH
O
NBocNHBoc
Succinimide ester coupling reactionscommon in peptide synthesis
activated esters
O
OHH2NO
OHNH
O
N
O
O
OF3C
OAc2O
MeOH reflux
recrystalization twice in EtOAc
y 62%
Pyr , THF N2, dark, RT2 h 40 min
tBu O O
O
OHH2N
tBu O O
O
OHNH
O
N
O
O
OF3C
OAc2O
H2O, Na2CO3H2O bath, ph 10
RT 24 hy 99%
Pyr THF N2 , Dark , 2 h 40 min
>100% mass recoverymostly product
three timesnot clean
similar impurity
N
O
O
OF3C
ON
O
O
HO TFAA, THF
RT 2 hy 79%
Syntheses of hydrazine functionalized amino acid and dipeptides
HO O
O
HNN
H
O
NH
O
O
O NHNH2
Carbazate
HO O
O
HNN
H
O
NH
O
O NHNH2
Acyl hydrazide
tBu O O
O
HNN
H
O
NH
O
OH
tBu O O
O
HNN
H
O
NH
O
O
O NHNHBoc
O
N N NN
H2N NHBoc
35% y over 4-stepswithout optimization
Carbazate model reactions didn't work.
Syntheses of hydrazine functionalized amino acid and dipeptides
HO O
O
HNN
H
O
NH
O
O
O NHNH2
H2NOH
OH
O
L-serine
H2N OH
O
NBocNHBoc
tBu O O
NH
O
N
O
O
O
O
HO O
O
HNN
H
O
NH
O
NHNH2
tBu O O
O
HNN
H
O
NH
O
OH
NH
O
N
O
O
O
O
O
HNN
H
O
NH
O
NHNH2
H2NOH
O
OHOL-glutamic acid
Summary
21% y over 4 stepsoptimization of removalof byproduct is needed
coupling reactions of succinimide esterswere not suitable HOBt/ EDC coupling
was cleanbut the conversion to
carbazate functional groupwas not successful
Project 2Study of the Amine to Alcohol Conversion via
N-Nitrosoamides
Clark University Master’s Seminar
08/12/2016Master’s Seminar
Study of amine-alcohol conversion via N-nitrosoamide
HN N
H
O
HNNH2
PeptidePeptide H2N OH
O
NH2
Lysine
PgHN OH
O
OH
6-HydroxynorleucineHydrazine functionalizedpeptide inhibitor
Key conversion
Synthesis of a peptide inhibitor
MeO
O
NH29 MeO
O
OH9model molecule
MeO
O
NH9 R
O
MeO
O
N9 R
O
N O
MeO
O
O9 R
O
NaNO2AcOHAc2O
K2CO3MeOH
Amine-alcohol 4-step sequence
CH3 CF3
Cl
Cl Cl
Cl
A scope of R groups based on previous study
to find the best substrate amide for this conversion
branched aromatic electron-withdrawing
Study of amine-alcohol conversion via N-nitrosoamide
MeOH 1MSOCl2 2 eq
0 oC then reflux 3 h84% y
HO
O
NH2 O
O
NH3 Cl
Syntheses started with ammonium chloride salt
MeO
O
NH3 ClEDC 1 eq/ HOBt 1 eq
TEA 1.1 eq, DCM 0.2 MRT overnight, 14% y
MeO
O
NH
OCl
OH
OCl
Cl
Cl1.1 eq
MeO
O
NH3 ClTEA 2 eq, DCM 0.2 MRT overnight, 94% y
MeO
O
NH
OCl
O
OCl
Cl
Cl5 eq
Na2CO3 1 M
99% yMeO
O
NH3 MeO
O
NH2Cl
Deprotonation to obtain amine
EtOAc 1MRT Overnight
99% y
MeO
O
NH2 MeO
O
NH
OCl
Cl
MeO
OCl
Cl
1.5 eq
No aqueous work up was needed, simple removal of volatiles
Conclusion: for all the amides needed, yields of 50%-99% on gram scale were obtained.
Study of amine-alcohol conversion via N-nitrosoamide
MeO
O
O7
O
Cl
Cl
MeO
O
NH7 R
O
MeO
O
N7 R
O
MeO
O
O7 Me
O
NaNO2 2+2 eq
NO
MeO
O
O7 R
O
MeO
O
O7 CF3
O
(A, desired product)
(B, desired rearrangement product)
(C, acetate)
(D, trifluoroacetate)
(E, dichloroacetate)
MeO
O
N7
O
O
F3C
( F, imidoyl trifluoroacetate)
N-Nitrosylation of amides.
0 oC, 6 hcosolvent 0.3 M
Anhydride/ acid (5:1)
Amide Solvent Conversion Product Distribution
NH
CH3
O
AcOH /Ac2O full A
NH
OCl
Cl
DCA /Ac2O
AcOH /Ac2O
DCA /EtOAc
full
82%
0%
94% (A) : 6% (B)
66% (A) : 16% (B)
none
NH
OAcOH /Ac2O full A
TFA /TFAA /EtOAc 80% F
NH
CF3
OAcOH /Ac2O 17% C
NH
O
TFA /Ac2O2 h
98% 78%(A):6%(B):14%(D)
DCA /Ac2O2 h
full 94% (A) : 6% (E)
iBA /iBAn6 h
full A
Study of amine-alcohol conversion via N-nitrosoamide
NaNO2EtOAc/TFA / TFAA
5:1:1MeO
O
NH MeO
O
N
OO
F3C
O
6 h, Ice Bath
The unexpected product raised questions about the mechanism of nitrosylation
ON
O
nitrite anion
O R
OH
O R
O
ON
OH
nitrous acidblue
in cold solution
ON
OH
HH2O
N O
nitrosonium cation
ON
OHside reactions:
H2O
NO2 NOO2
NO2
brown-red gas
NO
brown-red gas
colorless
imidoyl acetamide intermediate
NH
R'
O
O R''
O
OR'' R''
O O
H~
N R'
OR''
O
N O
N R'
O
N Ogreen
N2O4
keep reaction drythe additional NaNO2
the reaction does not occur without anhydride
CF3 is very electron-withdrawing,so the imidoly acetate
is less nucleophilic
Study of amine-alcohol conversion via N-nitrosoamide
MeO
O
O7
O
Cl
Cl
MeO
O
N7 R
O
MeO
O
O7 Me
O
MeO
O
7
MeO
O
O7 CF3
O
(A desired product)
(E, elimination)
(C, acetate)(B, trifluoroacetate)
(D, dichloroacetate)
( F, rearranged elimination)
NO
MeO
O
7
MeO
O
O7 R
OAmide
Solvent/Concentration
(mmol/ mL)Conversion Product
DistributionTemperature
/Time
24 h 88% 71%(A):16%(E)N CH3
O
0.11NO
toluene 70 ºC
toluene/ 0.11
toluene/1 eq AcOH
toluene/1 eq TEA
toluene1 eq AcOH/ 1 eq TEA
full
full
full
50 ºC/ 24 h
full
84%(A):5%(C):11%(E)
51%(A):36%(C):13%(E)
81%(A):8%(C):11%(E)
49%(A):38%(C):13%(E)
50 ºC/ 24 h
50 ºC/ 24 h
50 ºC/ 24 h
Cl
Cl
0.05
0.11
toluene
toluene
toluene 0.11 full
100 ºC/ 2 h
40 ºC/ 24 h
50 ºC/ 5 h
full
full
93%(A) :7%(E)
80%(A):6%(C):14%(E)
74%(A):3%(C):23%(E)
0.11toluene 50 ºC/ 24 h full 84%(A):2%(C):14%(E)
CF3
during nitrosylation
5 : 1 Ac2O/ TFA 0 ºC/ 6 h full
36(A):28%(C):24%(E):12%(F)
Several other solvents were also tested,full conversion was achieved in CCl4, DMF and EtOAc,DMF give more byproduct.EtOAc is the extraction solvent of nitrosylation
Study of amine-alcohol conversion via N-nitrosoamide
N R
O
N O
cyclic substitution
NN
O R
O
diazoester dissociation
N
diazoalkane
N O R
OH
recombine
N2
O R
O
Mechanism of thermal rearangement from literature
MeO
O
O7
O
Cl
Cl
MeO
O
N7 R
O
MeO
O
O7 Me
O
MeO
O
7
MeO
O
O7 CF3
O
(A desired product)
(E, elimination)
(C, acetate)(B, trifluoroacetate)
(D, dichloroacetate)
( F, rearranged elimination)
NO
MeO
O
7
MeO
O
O7 R
O
Rate determining stepelectron deficient R grouprearranges faster
Work up of nitrosylation reaction was optimized to remove anhydride and acid
Study of amine-alcohol conversion via N-nitrosoamide
MeO
O
O
O
MeO
O
OH
K2CO3 2 eq, MeOH 0.06 M22 h, RT
MeO
O
OH
K2CO3 2 eq, MeOH 0.2 MRT overnight
MeO
O
O
O
almost full conversion48% mass recovery
63% y
MeO
O
OHMeO
O
O
OCl
Cl RT, 2h65% y
TEA 2 eq, MeOH 0.2 M
Mild and selective base catalyzed hydrolysis
Study of amine-alcohol conversion via N-nitrosoamide
full conversion99% mass recovery
MeO
OCl
Cl
MeO
O
NH7
OCl
Cl
MeO
O
N7
OCl
ClNO
MeO
O
O7
OCl
Cl
NaNO2 2+2 eq,Cl2CHCOOH / Ac2O (1:1)
0 ºC, 6 h
full conversionused crude
40 ºC, 16 h, EtOAc
98% mass recoveryclean full conversion
without Ac2O
RT, 2 hcolumn purification
65% y
TEA 2 eq, MeOH 0.2 M
MeO
O
NH27
MeO
O
OH7
Dichloro group was found to be the optimal choice over 4 steps.
Easy to execute with 2 aqueous work ups and only one column. Over all yield was 63%.
1.5 eq
EtOAc 1M, 21 h, RT
Study of amine-alcohol conversion via N-nitrosoamide
ONH2 O NH2
N3 NH2Si
3 amine substrates
O
OHO
O
OO O
NH2NH
+HONH2
THF/ H2O (1:1) 0.43 M
NaOH, R.T. 4 h
Allyl Bromide 1.3 eq
Tetrabutylamonium Iodide 0.1 eqTHF 0.24 M, N2 protected30 min 0C then R.T. 24 h
Column, 2 steps y 24%TFA/ CH2Cl2 (1:1) 0.12 M
30 min RT
basic extraction quant. y
NaH 1 eq
NHO
OO
O O
Synthesis of allyl amine.
24% y overall
protect the amine first
allyl iodide intermediate has better leaving group
0.54 eq
Study of amine-alcohol conversion via N-nitrosoamide
BrBr N3 Br
O
ON3 N
N3 NH2
NaN3 1 eqDMF 0.4 M60°C, 4.5 h
Column
N
O
O
K
DMF 0.15M 70C, 5 h
H2N NH2 • H2O 3.5 eq
Ethanol 0.07 MReflux 4 h
Azide big enough to be handledy 23% Columny 71%
y 89%
Synthesis of azido amine
15% y overall
azide substitution, not selectiveexcess bromide to consume azide basic work up to avoid
formation to hydrazoic acid
Gabriel synthesis with Ing-Maske procedure
removal of DMF is important
1.5 eq
+
ClSi NH2HO NH2TBDPSO
Imidazole 2.2 eq
DCM 0.1 M0°C then RT 24 h
Synthesis of amine.
2 eq
O
OTBDPSO NH
TBDPSO NH2
Cl O
O
TEA 3.2 eq
DCM 0.16 M1 h R.T.column
2 steps y: 32%
Pd(OH)2/C 50mg/mmol
H2 , MeOH 0.2 M, R.T. 4 h y 95% 30% y overallless polar, can be column purified
2 eq
Study of amine-alcohol conversion via N-nitrosoamide
N3
Cl
Cl
O N
O
NN3
Cl
ClO
O
N3
Cl
ClO
NH
O
O
MeCl
Cl
EtOAc, RT, 24 h
NaNO2Cl2CHCOOH
Ac2O
0 °C, 6 h
N3 NH2
a mixture of compounds
EtOAc, 40°C,24 h
Attempts of conversion of amines to alcohols via standard procedure.
Cl
Cl
O N
O
NO
Cl
ClO
OO
Cl
ClO
NH
O
O
O
MeCl
Cl
EtOAc, RT, 24 h
EtOAc, 40°C,24 h
ONH2
not the expected productnot compatible
NaNO2Cl2CHCOOH
Ac2O
0 °C, 6 h
Study of amine-alcohol conversion via N-nitrosoamide
TBDPSO NH2
TBDPSO OH
y 38% over 4 stepswith trace amount of impurity
O
O
MeCl
Cl
EtOAc, RT, 24 h
TBDPSO NH
OCl
Cl
NaNO2Cl2CHCOOH
Ac2O
0 °C, 6 h
TBDPSO N
OCl
ClNO
EtOAc, 40°C,24 hTBDPSO O
OCl
Cl
TEA MeOH
silica gel chromotography
Attempts of conversion of amines to alcohols via standard procedure.
38% y
TIPSO NH2 TIPSO OH
y 49% over 4 steps
Conversion of a similar amine substrate to alcohols via standard procedure.
Summary
HN N
H
O
HNNH2
PeptidePeptide H2N OH
O
NH2
Lysine
PgHN OH
O
OH
6-HydroxynorleucineHydrazine functionalizedpeptide inhibitor
Key conversion
Synthesis of a peptide inhibitor
MeO
O
NH29 MeO
O
NH9 R
O
MeO
O
N9 R
O
N O
MeO
O
O9 R
O
MeO
O
OH9
NaNO2AcOHAc2O
K2CO3MeOH
CH3 CF3
Cl
Cl Cl
Cl
Amine-alcohol 4-step sequence
A scope of R group
model molecule
NaNO2EtOAc/TFA / TFAA
5:1:1MeO
O
NH MeO
O
N
OO
F3C
O
6 h, Ice Bath
Formation of unexpected imidoyl trifluoroacetate product
imidoyl trifluoroacetate3 Amine substrates for compatibility study of amine-alcohol 4 step sequence
ONH2 O NH2
N3 NH2Si
38% y
Summary
1. Synthesis of Small Molecules and Amino Acids for LOX InhibitionProjects:
2. Study of the Amine to Alcohol Conversion via N-Nitrosoamides
MeO
O
NH29 MeO
O
NH9 R
O
MeO
O
N9 R
O
N O
MeO
O
O9 R
O
MeO
O
OH9
NaNO2AcOHAc2O
K2CO3MeOH
model molecule
HN N
H
O
HNNH2
PeptidePeptide H2N OH
O
NH2Lysine
PgHN OH
O
OH6-Hydroxynorleucine
Key conversion
HN
NH2 NH
NH
NH2
O
phenylhydrazine semicarbazide
H2N OH
O
NBocNHBoc
HO O
O
HNN
H
O
NH
O
NHNH2
Acknowledgements• Committee Members:• Professor Jakobsche• Professor Greenaway• Professor Granados-Focil• Professor Turnbull
• Group Members:• Alexander Wall• Ashley Burke• Blaine McCarthy• Danielle Augur• Devon Fontaine• Maria Solares Bucaro• Mike Reardon• Nick MacArthur• Tony Xu • Rachel Donnelly-Cokinos
• Special Thanks:• Dr. Lin• Frank Abell
• Rene Baril• Meghan O'Rourke• Sue Dejong• Ernest Krygier• Kostika Stefo• Wendy Nason• Rhady Sorm• Kai Peng• Will Wei• Amy Zheng• Mark Tan • Everyone that helped me grow in the past
three years and the audience today.
Clark University Master’s Seminar
08/12/2016Master’s Seminar