A simple and efficient synthesis of N-[2-{(4-propionyloxyphenyl)thio}ethyl] propionamide (DIPCAP), a tyrosine-dependent cytotoxic agent as a topical antimelanoma therapy

Article abstract of DOI:10.1081/SCC-200026188

The synthesis of N-[2-{(4-propionyloxyphenyl)thio}ethyl]propionamide (DIPCAP), a tyrosine-dependent cytotoxic agent that is currently being evaluated as a topical antimelanoma treatment is described. Problems encountered in scaling up the original patent

Full text of DOI:10.1081/SCC-200026188

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A Simple and
Efficient Synthesis of
N‐[2‐{(4‐Propionyloxyphenyl)thio}
(DIPCAP), a
Tyrosine‐Dependent
Cytotoxic Agent as a Topical
Antimelanoma Therapy
Lisa D. Coutts a , Harold Meckler a & Karl F. Popp b
a Albany Molecular Research Inc. , P.O. Box
15098, 21 Corporate Circle, Albany, New York,
12212‐5098, USA
b August C. Stiefel Research Institute, Inc. , Oak
Hill, New York, USA
Published online: 12 Jan 2011.
To cite this article: Lisa D. Coutts , Harold Meckler & Karl F. Popp (2004) A Simple
and Efficient Synthesis of N‐[2‐{(4‐Propionyloxyphenyl)thio}ethyl]propionamide
(DIPCAP), a Tyrosine‐Dependent Cytotoxic Agent as a Topical Antimelanoma
Therapy, Synthetic Communications: An International Journal for Rapid
Communication of Synthetic Organic Chemistry, 34:15, 2697-2704, DOI: 10.1081/
SCC-200026188
To link to this article: http://dx.doi.org/10.1081/SCC-200026188

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SYNTHETIC COMMUNICATIONSw
Vol. 34, No. 15, pp. 2697–2704, 2004
A Simple and Efficient Synthesis of
N-[2-f(4-Propionyloxyphenyl)thiogethyl]
propionamide (DIPCAP), a Tyrosine-
Dependent Cytotoxic Agent as a Topical
Antimelanoma Therapy
2
Lisa D. Coutts,1 Harold Meckler,1, and Karl F. Popp
*
1Albany Molecular Research Inc., Albany, New York, USA
2August C. Stiefel Research Institute, Inc., Oak Hill, New York, USA
ABSTRACT
The synthesis of N-[2-f(4-propionyloxyphenyl)thiogethyl]propionamide
(DIPCAP), a tyrosine-dependent cytotoxic agent that is currently being
evaluated as a topical antimelanoma treatment is described. Problems
encountered in scaling up the original patent procedure have been elimi-
nated and DIPCAP was obtained in an overall yield of 65% from com-
mercially available starting materials.
Key Words: DIPCAP; Tyrosine-dependent; Antimelanoma; N-[2-f(4-
Propionyloxyphenyl)thiogethyl]propionamide; 2-Ethyl-2-oxazoline; 4-
Hydroxythiophenol.
*
Correspondence: Harold Meckler, Albany Molecular Research Inc., P.O. Box 15098,
21 Corporate Circle, Albany, NY 12212-5098, USA; E-mail: hmeckler@albmolecular.
com.
2697
DOI: 10.1081/SCC-200026188
0039-7911 (Print); 1532-2432 (Online)
www.dekker.com
Copyright # 2004 by Marcel Dekker, Inc.

2698
Coutts, Meckler, and Popp
INTRODUCTION
N-[2-f(4-Propionyloxyphenyl)thiogethyl]propionamide (DIPCAP, 1) is a
member of a small class of compounds known as phenolic thioether amines
that have demonstrated[1] selective in vitro cytotoxicity. This activity has
been proposed[2] to be tyrosinase-dependent, making (1) a possible candidate
for targeted topical antimelanoma therapy. Melanocytes are pigmented cells
found in the basal layer of the epidermis and are the normal precursor cells
to melanoma. One step in this conversion is proposed[3] to be tyrosinase-
mediated. DIPCAP (1) has been postulated[4] to selectively accumulate in
melanocytes and causes a reduction in tumor size through its cytotoxic
activity. The structure of 1 and its cytotoxic activity in ten different cell
lines have recently been described in the literature.[4]
RESULTS AND DISCUSSION
While the synthesis of 1 has not been fully described in the literature, the
preparation of the key intermediate, N-propionyl-4-S-cysteaminylphenol
(P-CAP, 2) and the O-acetylated analog 3 have been described (Fig. 1).[5,6] In
these reports, the reaction of 5 and 6 to afford 2 was run without any solvent.
When 6 was added to 408C melt of 5, the reaction exothermed to about 1408C,
and was then heated at 1608C overnight. The next day, the reaction mixture
was cooled to room temperature, concentrated, and purified by column chromato-
graphy. The reported acylationof2 was runatroom temperature in pyridine as the
solvent; crude 1 that was obtained required purification by tedious preparative
chromatography. Attempts to scale these procedures to 50-g quantities met
with critical processing problems: an uncontrollable, 1508C runaway exotherm
in the reaction of 5 and 6 to produce 2, and overpropionylation of 2 causing a
significant quantity of impurity 4. Since kilogram quantities of 1 were required,
these undesirable process parameters required a detailed examination. Also, no
spectral data for 1 have been reported in the literature. We now report an efficient
reaction sequence to afford 1 in overall 65% yield from commercially available,
technical grade (90% pure) 4-hydroxythiophenol (Sch. 1).
While the condensation of a thiophenol with 2-ethyl-2-oxazoline, 6, as a
neat melt at 1608C has been described in the literature,[7,8] there has been no
mention of any exothermic events occurring. To solve the runaway exotherm
problem, we encountered during the reaction of 5 and 6 to afford 2, many
solvents were examined. N,N-Dimethylformamide, dimethylsulfoxide, and
N-methylpyrrolidinone all dissolved the starting materials and allowed the
reaction to proceed to completion, but 2 was difficult to isolate from these
solvents. The reaction did not proceed in ethylene glycol, propylene glycol,

Synthesis of DIPCAP
2699
Figure 1. Structures of DIPCAP (1) and related compounds.
or poly(ethylene glycol-400) [PEG-400]. The reaction did proceed smoothly in
refluxing xylenes, but the product 2 separated from the hot reaction mixture as a
“stringy oil”, which rapidly turned dark. The only solvent in which the reaction
proceeded smoothly and afforded 2 as a white crystalline solid was toluene.
Preheating a solution of 5 in toluene (0.33 g/mL) to 908C and adding
a solution of 2-ethyl-2-oxazoline (6) in toluene (0.27 mL/mL) led to a very
controllable 108C temperature rise. The reaction mixture was then heated to
reflux for 7 hr during which time all of reactant 5 was consumed. The heat
was turned off and the reaction mixture was stirred at room temperature over-
night during which time phenol 2 crystallized. The white crystalline solid
was collected by filtration and rinsed with methylene chloride.a The solid
was dried to afford 2 in 70% yield and the purity was 98.5% (HPLC).
aThe methylene chloride rinse was found to efficiently remove the colored impurities.

2700
Coutts, Meckler, and Popp
Scheme 1. (a) Toulene, reflux, 7 hr; toulene recrystallization; 78%; (b) propionic
anhydride/pyridine/CH2Cl2, 288C for 5 hr; then 208C for 18 hr; MeOH/H2O
crystallization; 83%.
The attempted propionylation of 2 under standard conditions (propionic
anhydride or propionyl chloride in pyridine at room temperature) led to the
formation of a significant quantity of the overpropionylated by-product 4
and a brown, thin layer chromatography (TLC) baseline impurity which
imparted an orange color to the crude product. After several experiments,
it was found that the offending impurities were controlled by running the
reaction in methylene chloride, below 258C and by limiting the propionic
anhydride and pyridine quantities to 1.16 and 1.94 M equiv., respectively.
These modifications afforded crude 1 in .90% isolated yield, and product 1
as a light yellow solid that was .98% pure (HPLC). The treatment of a sol-
ution of 1 in methanol with a mixture of decolorizing carbon and activated
clay[9] removed the color. Following addition of this clarified methanol sol-
ution to water, 1 was isolated as a white crystalline product.
EXPERIMENTAL SECTION
All reactions were performed under an atmosphere of dry nitrogen.
Reagents purchased from commercial sources were used as received, unless

Synthesis of DIPCAP
2701
noted otherwise. Proton magnetic resonance spectra were obtained on a
Brucker AMX-500 MHz NMR, using tetramethylsilane as an internal refer-
ence. Infrared spectra were recorded with a Nicolet Nexus 470 FT-IR spectro-
photometer. Mass spectra analyses were performed on a Finnigan LCQ-DUO
mass spectrometer (APCI mass spectrometry). Chemical purities were deter-
mined on a Spectra-Physics HPLC system with a Waters Nova-Pak C8 column
(4.6 ꢀ 250 mm), with a gradient elution of 80 : 20 to 50 : 50 water/aceto-
nitrile, at a flow rate of 1.0 mL/min, and UV detection at 254 nm. TLC was
˚
performed using 1 ꢀ 3 in. Whatman 60 A (0.025 mm thick) silica gel plates.
TLC plates were visualized by observation under UV lamp or by staining
with a solution of ceric ammonium sulfate in 50% aqueous sulfuric acid.
Melting points were obtained by differential scanning calorimetry using a
Mettler Toledo Star 80 DSC, calibrated on an indium standard. Elemental
analyses were performed by Quantitative Technologies, Inc., Whitehouse, NJ.
N-Propionyl-4-S-cysteaminylphenol (2)
To a 22-L, four-necked, round-bottomed flask, equipped with an overhead
stirrer, water-cooled reflux condenser, a thermocouple temperature recorder,
and a pressure-equalizing addition funnel, was charged Aldrich 90% technical
grade 4-hydroxythiophenol (5, 1.99 kg, 15.7 mol) and toluene (6 L). The stirrer
was started and the solution was heated to 908C. To the pressure-equalizing
addition funnel was charged a solution of 2-ethyl-2-oxazoline (6, 1.57 kg,
1.60 L, 15.6 mol) in toluene (6 L), which was added to the hot toluene solution
of 5 over 2 hr. At this point, the reaction temperature was raised to 1108C and
held for 7 hr. The reaction progress was monitored by TLC (5% methanol in
methylene chloride eluent). After complete consumption of 5, the reaction
mixture was allowed to cool and the resulting slurry was stirred at room temp-
erature for 18 hr. The solids were collected by filtration, rinsed with methylene
chloride (1.9 L), and air-dried. Crude 2 (5.5 kg) was suspended in toluene
(16 L) in a 22-L flask equipped with an overhead stirrer and a water-cooled
distillation head. The suspension was heated to 1058C, during which time
1 L of distillate was removed. The resulting solution was hot-filtered
through filter paper and the filtrate was allowed to cool to room temperature
overnight. The solid product was collected by filtration and dried at 558C
and 1 mmHg vacuum for 18 hr to afford a 78% yield (corrected for the
purity of the starting material) of P-CAP, 2 (2.51 kg) as a white solid:
HPLC purity was 98.5% (AUC).
M.p. 84–868C. 1H NMR (500 MHz, CDCl3) d ppm ¼ 8.30 (s, 1H),
7.31–7.29 (d, J ¼ 8.6 Hz, 2H), 6.82–6.80 (d, J ¼ 8.6 Hz, 2H), 6.07–6.02
(br s, 1H), 3.44–3.40 (t, J ¼ 6.5 Hz, 2H), 2.73–2.18 (dd, J ¼ 15.3; 7.6 Hz,

2702
Coutts, Meckler, and Popp
2H), 1.15–1.12 ppm (t, J ¼ 7.6 Hz, 3H). 13C NMR (125 MHz, CDCl3): 175.2,
157.0, 134.4, 123.4, 116.6, 38.9, 35.6, 29.9, 10.0. IR (KBr) 3291, 3096, 2972,
1641, 1562, 1496, 1255, 890 cm21. APCI MS (m/z): 226 [M þ 1]þ. Anal.
calcd. for C, H, and N: 58.64, 6.71, and 6.22. Found: 58.59, 6.71, and 6.15,
respectively.
Crude N-[2-f(4-propionyloxyphenyl)thiogethyl]propionamide
(Crude 1)
To a 22-L, four-necked, round-bottomed flask equipped with an overhead
stirrer, thermocouple temperature recorder, and a nitrogen bubbler was
charged phenol 2 (2.0 kg, 8.9 mol) and methylene chloride (10 L). The
stirrer was started and, when complete solution was obtained, pyridine
(1.37 kg, 1.40 L, 17.3 mol) was added. The reaction mixture was cooled to
288C, at which point propionic anhydride (1.36 kg, 1.34 mL, 10.4 mol) was
added at a rate to keep the internal temperature below 258C. The reaction
mixture was stirred at 288C for 5 hr and then allowed to warm to 208C for
18 hr. The reaction was monitored by TLC (2% methanol in methylene chlor-
ide eluent). After complete reaction, the mixture was washed with 2 N HCl
(2 ꢀ 5 L) and saturated aqueous sodium bicarbonate solution (2 ꢀ 5 L). The
organic layer was concentrated by atmospheric pressure distillation to a
volume of 4 L. Cyclohexane (15 L) was added over 1-hr period, so the
product crystallized without caking on the agitator and thermocouple well.
The white slurry was stirred at ambient room temperature for 5 hr and the
solid product was collected by vacuum filtration. The filter cake was rinsed
with cyclohexane (2 L), air-dried, then dried at 508C and 1 mmHg vacuum
for 24 hr to afford a 92% yield of crude 1 (2.27 kg) as a light yellow solid.
The HPLC purity was 98.1% (AUC).
Purification of N-[2-f(4-
propionyloxyphenyl)thiogethyl]propionamide (1)
To a 22-L, four-necked, round-bottomed flask, equipped with an overhead
stirrer, was charged crude 1 (1.20 kg, 4.26 mol) and methanol (5.0 L). The
stirrer was started and the slurry was warmed in a 308C water bath until
complete solution was obtained. To this solution was added 75 g of a 1 : 1
mixture of decolorizing carbon (Darco G-60) and activated clay (Clarion
550) and the slurry was stirred for 1 hr. The mixture was clarified through a
diatomaceous earth pad and the filtrate was then slowly charged to 10 L of
stirred water in a clean 22-L, round-bottomed flask equipped with an overhead

Synthesis of DIPCAP
2703
stirrer. The resulting slurry was stirred for 1.5 hr at 208C. The crystalline
product was collected by vacuum filtration, rinsed with water (5.0 L), and
air-dried for 1 hr. The solid was dried at 558C and 1 mmHg vacuum for
19 hr to afford a 90% recovery of 1 (1.09 kg) as a white solid. The HPLC
purity was 99.1% (AUC).
M.p. 98–1008C. 1H NMR (500 MHz, CDCl3) d ppm ¼ 7.39–7.35
(d, J ¼ 8 Hz, 2H), 7.03–7.01 (d, J ¼ 8.4 Hz, 2H), 6.06–5.89 (br s, 1H),
3.45–3.41 (q, J ¼ 6.5 Hz, 2H), 2.60–2.55 (dd, J ¼ 15.2; 7.5 Hz, 2H), 2.17–
2.13 (dd, J ¼ 15.3; 7.6 Hz, 2H), 1.26–1.23 (t, J ¼ 7.6 Hz, 3H), 1.13–
1.10 ppm (t, J ¼ 7.6 Hz, 3H). 13C NMR (125 MHz, CDCl3): 174.1, 173.0,
149.7, 132.3, 131.2, 122.5, 38.7, 34.2, 29.7, 27.8, 9.9, 9.2. IR (KBr) 3294,
3095, 2977, 2939, 1750, 1642, 1561, 1492, 1440, 1230, 1172, 894, 807,
513 cm21. APCI MS (m/z): 282 [M þ 1]þ. Anal. calcd. for C, H, N, and S:
59.76, 6.81, 4.98, and 11.39. Found: 59.97, 6.94, 4.97, and 11.50, respectively.
ACKNOWLEDGMENT
The authors would like to thank Paul Isbester and Frank Scuderi for the
spectroscopic analyses and Sandy Birk for assistance with the preparation
of this manuscript. Additionally, we would like to thank the August
C. Stiefel Research Institute for permission to publish this study.
REFERENCES
1. Prezioso, J.A.; Epperly, M.W.; Wang, N.; Bloomer, W.D. Effects of
tyrosinase activity on the cytotoxicity of 4-S-cysteaminylphenol and
N-acetyl-4-S-cysteaminylphenol in melanoma cells. Cancer Lett. 1992,
63, 73–79.
2. Jimbow, K.; Fitzpatrick, T.B.; Wick, M.; Goldsmith, L.A., Eds.;
Biochemistry and physiology of melanin pigmentation. In Physiology
and Molecular Biology of the Skin; Oxford University Press: New York,
1991; 873–909.
3. Cooksey, C.J.; Garratt, P.J.; Land, E.J.; Pavel, S.; Ramsden, C.A.;
Riley, P.A.; Smit, N.P. Evidence of the indirect formation of the catecholic
intermediate substrate responsible for the autoactivation kinetics of tyrosi-
nase. J. Biol. Chem. 1997, 272, 26226–26235.
4. Gili, A.; Thomas, P.D.; Ota, M.; Jimbow, K. Comparison of in vitro cyto-
toxicity of N-acetyl and N-propionyl derivatives of phenolic thioether
amines in melanoma and neuroblastoma cells and the relationship to

2704
Coutts, Meckler, and Popp
tyrosinase and tyrosine hydroxylase enzyme activity. Melanoma Res.
2000, 10, 9–15.
5. Tandon, M.; Thomas, P.D.; Shokravi, M.; Singh, S.; Samra, S.; Chang, D.;
Jimbow, K. Synthesis and antitumour effect of the melanogenesis-based
antimelanoma agent, N-propionyl-4-S-cysteaminylphenol. Biochem.
Pharm. 1998, 55, 2023–2029.
6. Jimbow, K. Phenolic Amine Depigmenting and Antimelanoma Agents. US
Patent 5,395,611, March 7, 1995.
7. Wehrmeister, H.L. Reactions of aromatic thiols with oxazolines. J. Org.
Chem. 1963, 28, 2587–2589.
8. Lis, R.; Morgan, J.K.; Marisca, A.J.; Gomez, R.P.; Lind, J.M.; Davey, D.D.;
Phillips, G.B.; Sullivan, M.E. Synthesis of novel (aryloxy)propanolamines
and related compounds possessing both class II and class III antiarrhythmic
activity. J. Med. Chem. 1990, 33, 2883–2891.
9. Herr, R.J.; Meckler, H.; Scuderi, F., Jr. Observed acidities of charcoals,
clays, and common laboratory purification reagents in aqueous and
organic solutions. Org. Process Res. Dev. 2000, 4, 43–45.
Received in the USA March 31, 2004